# Boosting photosynthesis opens new opportunities for agriculture sustainability and circular economy: The BEST - CROP research and innovation action

Paolo Pesaresi, Pierre Bono, Stephane Corn, Cristina Crosatti, Sara Daniotti, Jens Due Jensen, Ivo Frébort, Eder Groli, Claire Halpin, Mats Hansson, et al.

<figure><img src="image_1.png" /></figure>

## To cite this version:

Paolo Pesaresi, Pierre Bono, Stephane Corn, Cristina Crosatti, Sara Daniotti, et al. Boosting photosynthesis opens new opportunities for agriculture sustainability and circular economy: The BEST - CROP research and innovation action. The Plant Journal, 2025, 121 (3), pp.e17264. 10.1111/tpj.17264. hal-04932089

HAL Id: hal-04932089

https://imt-mines-ales.hal.science/hal-04932089v1

Submitted on 6 Feb 2025

HAL is a multi-disciplinary open access archive for the deposit and dissemination of scientific research documents, whether they are published or not. The documents may come from teaching and research institutions in France or abroad, or from public or private research centers.

L'archive ouverte pluridisciplinaire HAL, est destinée au dépôt et à la diffusion de documents scientifiques de niveau recherche, publiés ou non, émanant des établissements d'enseignement et de recherche français ou étrangers, des laboratoires publics ou privés.

<figure><img src="image_2.png" /></figure>

Distributed under a Creative Commons Attribution 4.0 International License
The Plant Journal (2025) **121**, e17264

doi: 10.1111/tpj.17264

PERSPECTIVE

Boosting photosynthesis opens new opportunities for agriculture sustainability and circular economy: The BEST-CROP research and innovation action

Paolo Pesaresi<fnref n="1" />,* ID, Pierre Bono<fnref n="2" />, Stephane Corn<fnref n="3" /> ID, Cristina Crosatti<fnref n="4" /> ID, Sara Daniotti<fnref n="5" /> ID, Jens Due Jensen<fnref n="6" />, Ivo Frébort<fnref n="7" /> ID, Eder Groli<fnref n="8" /> ID, Claire Halpin<fnref n="9" /> ID, Mats Hansson<fnref n="10" /> ID, Goetz Hensel<fnref n="11" />,<fnref n="12" /> ID, David S. Horner<fnref n="1" /> ID, Kelly Houston<fnref n="13" /> ID, Ahmed Jahoor<fnref n="6" />, Milos Klíma<fnref n="14" />, Hannes Kollist<fnref n="15" />,<fnref n="16" /> ID, Clément Lacoste<fnref n="17" /> ID, Boubker Laidoudi<fnref n="2" />, Susanna Larocca<fnref n="18" />, Caterina Mare<fnref n="4" /> ID, Nicolas Le Moigne<fnref n="17" /> ID, Chiara Mizzotti<fnref n="1" /> ID, Tomas Morosinotto<fnref n="19" /> ID, Klaus Oldach<fnref n="20" /> ID, Laura Rossini<fnref n="21" /> ID, Sebastian Raubach<fnref n="22" /> ID, Miguel Sanchez-Garcia<fnref n="23" />, Paul D. Shaw<fnref n="22" /> ID, Rodolphe Sonnier<fnref n="17" /> ID, Alessandro Tondelli<fnref n="4" /> ID, Robbie Waugh<fnref n="13" /> ID, Andreas P.M. Weber<fnref n="11" />,<fnref n="24" /> ID, Dmitry Yarmolinsky<fnref n="15" /> ID, Alessandro Zeni<fnref n="5" /> and Luigi Cattivelli<fnref n="4" />,* ID

Received 30 September 2024; revised 20 December 2024; accepted 3 January 2025.

*For correspondence (e-mail paolo.pesaresi@unimi.it and luigi.cattivelli@crea.gov.it).

Authors are listed alphabetically (with the exception of Paolo Pesaresi and Luigi Cattivelli, coordinating editors).

<footnote n="1">¹Department of Biosciences, University of Milan, Milan 20133, Italy,</footnote>

<footnote n="2">²FRD-CODEM (Fibres Recherche Développement-Construction Durable et EcoMatériaux), Hôtel de Bureaux, Technopole de l'Aube en Champagne, 2 rue Gustave Eiffel, CS 90601, Troyes Cedex 9 10 901, France,</footnote>

<footnote n="3">³LMGC, IMT Mines Ales, Univ Montpellier, CNRS, Ales, France,</footnote>

<footnote n="4">⁴Council for Agricultural Research and Economics (CREA) – Research Centre for Genomics and Bioinformatics, Fiorenzuola d’Arda 29017, Italy,</footnote>

<footnote n="5">⁵Consorzio Italbiotec, Piazza della Trivulziana 4, Milan 20126, Italy,</footnote>

<footnote n="6">⁶Nordic Seed A/S, Kornmarken 1, Galten 8464, Denmark,</footnote>

<footnote n="7">⁷Czech Advanced Technology and Research Institute (CATRIN), Palacký University Olomouc, Šlechtitelů 27, Olomouc 783 71, Czech Republic,</footnote>

<footnote n="8">⁸S.I.S. Società Italiana Sementi, via Mirandola di Sopra 5, 40068 S. Lazzaro di S, Bologna, Italy,</footnote>

<footnote n="9">⁹Division of Plant Sciences, School of Life Sciences, University of Dundee at the James Hutton Institute, Dundee DD2 5DA, UK,</footnote>

<footnote n="10">¹⁰Department of Biology, Lund University, Lund 22362, Sweden,</footnote>

<footnote n="11">¹¹Cluster of Excellence in Plant Sciences “SMART Plants for Tomorrow’s Needs”, Heinrich Heine University Düsseldorf, Düsseldorf, Germany,</footnote>

<footnote n="12">¹²Centre for Plant Genome Engineering, Heinrich Heine University Düsseldorf, Düsseldorf, Germany,</footnote>

<footnote n="13">¹³Cell and Molecular Sciences, James Hutton Institute, Errol Road, Invergowrie, Dundee DD25DA, UK,</footnote>

<footnote n="14">¹⁴Úsovsko a.s., Klopina 33, Klopina 789 73, Czech Republic,</footnote>

<footnote n="15">¹⁵Institute of Bioengineering, University of Tartu, Tartu 50411, Estonia,</footnote>

<footnote n="16">¹⁶Institute of Plant Sciences Paris-Saclay (IPS2) Université Paris-Saclay, CNRS, INRAE, Université Evry, Université Paris Cité, Gif sur Yvette 91190, France,</footnote>

<footnote n="17">¹⁷Polymers, Composites and Hybrids (PCH), IMT Mines Ales, Ales, France,</footnote>

<footnote n="18">¹⁸SO.G.I.S. Industria Chimica SpA, Cremona, Italy,</footnote>

<footnote n="19">¹⁹Department of Biology, University of Padova, Padova, Italy,</footnote>

<footnote n="20">²⁰KWS LOCHOW GmbH, Bergen, Lower Saxony, Germany,</footnote>

<footnote n="21">²¹Department of Agricultural and Environmental Sciences-Production, Landscape, Agroenergy (DiSAA), University of Milan, Milan 20133, Italy,</footnote>

<footnote n="22">²²Information and Computational Sciences, James Hutton Institute, Errol Road, Invergowrie, Dundee DD2 5DA, UK,</footnote>

<footnote n="23">²³International Center for Agricultural Research in the Dry Areas (ICARDA), Rabat 10100, Morocco, and</footnote>

<footnote n="24">²⁴Institute for Plant Biochemistry, Heinrich Heine University Düsseldorf, Düsseldorf, Germany</footnote>
## SUMMARY

There is a need for ground-breaking technologies to boost crop yield, both grains and biomass, and their processing into economically competitive materials. Novel cereals with enhanced photosynthesis and assimilation of greenhouse gasses, such as carbon dioxide and ozone, and tailored straw suitable for industrial manufacturing, open a new perspective for the circular economy. Here we describe the vision, strategies, and objectives of BEST-CROP, a Horizon-Europe and United Kingdom Research and Innovation (UKRI) funded project that relies on an alliance of academic plant scientists teaming up with plant breeding companies and straw processing companies to use the major advances in photosynthetic knowledge to improve barley biomass and to exploit the variability of barley straw quality and composition. We adopt the most promising strategies to improve the photosynthetic properties and ozone assimilation capacity of barley: (i) tuning leaf chlorophyll content and modifying canopy architecture; (ii) increasing the kinetics of photosynthetic responses to changes in irradiance; (iii) introducing photorespiration bypasses; (iv) modulating stoma opening, thus increasing the rate of carbon dioxide fixation and ozone assimilation. We expect that by improving our targeted traits we will achieve increases in aboveground total biomass production without modification of the harvest index, with added benefits in sustainability via better resource-use efficiency of water and nitrogen. In parallel, the resulting barley straw is tailored to: (i) increase straw protein content to make it suitable for the development of alternative biolubricants and feed sources; (ii) control cellulose/lignin contents and lignin properties to develop straw-based construction panels and polymer composites. Overall, by exploiting natural- and induced-genetic variability as well as gene editing and transgenic engineering, BEST-CROP will lead to multi-purpose next generation barley cultivars supporting sustainable agriculture and capable of straw-based applications.

**Keywords:** barley, canopy photosynthesis, straw quality, circular bioeconomy, straw-based panels, composites, biolubricants, feed.

## INTRODUCTION

The continuous growth of the world population is driving an increase in the demand for food and feed up to 50% by 2050 (van Dijk et al., 2021). This, in turn, will require a boost of agricultural productivity, while simultaneously reducing the negative environmental impacts (Haughey et al., 2023), like freshwater consumption and greenhouse gas (GHG) emissions, and without increasing cultivated land area.

BEST-CROP (https://www.bestcrop.eu)—Boosting photosynthesis To deliver novel CROPs for the circular bioeconomy—is a research and innovation action aiming to contribute to these challenges in the frame of the circular and bioeconomy transition, supporting the European Green Deal. Specifically, BEST-CROP targets barley (Hordeum vulgare L.), a major crop worldwide, with 153.6 million tons of grain produced in the 2022/23 season and an almost equivalent amount of straw covering around 60 million ha of world arable land (Taner et al., 2004).

The ambition is to develop next generation multi-purpose barley plants with: (i) increased uptake of carbon dioxide (CO₂) and ozone (O₃); (ii) enhanced total biomass production without modification of the harvest index; (iii) straw composition tailored for transformation into high-value bio-based industrial products for the feed,

green chemistry (biolubricants), construction, and composites sectors (see Figure 1).

The experimental plan builds on evidence that genetic variability impacting photosynthesis (Croce et al., 2024; Flood et al., 2011), canopy architecture (Mantilla-Perez & Salas Fernandez, 2017; Sakamoto et al., 2006; Shaaf et al., 2019), and barley straw composition, in terms of lignin (Daly et al., 2019; Grove et al., 2003; Halpin, 2019; Li et al., 2003; Zhang et al., 2010) and protein content (Bellucci et al., 2017; Przulj & Momcilovic, 2001; White et al., 1981), exists within barley germplasm and mutant collections and can be exploited to design a next generation barley plant. Research conducted in different laboratories worldwide has demonstrated that genetic manipulation of model organisms has the potential to improve photosynthesis efficiency (for a review see Croce et al., 2024), O₃ uptake (Brosché et al., 2010; Morales et al., 2021; Sierla et al., 2018; Yamauchi et al., 2016), and straw quality (Araus et al., 2016; Guo et al., 2020; Halpin, 2019; Karunarathe et al., 2022; Wang, Nian, et al., 2018). The performance of advanced breeding lines, carrying the selected traits, will be evaluated under field conditions in different European agroclimatic scenarios by monitoring, among other things, total biomass production, grain yield, and O₃
<figure><img src="image_3.png" /><figcaption>**Figure 1.** Overview of the main objectives and products of BEST-CROP. Next generation barley plants with optimized canopy photosynthesis will provide high-quality straw to produce biolubricants, construction panels, and feed.</figcaption></figure>

uptake. Finally, BEST-CROP tailors the straw composition to different industrial transformation processes by increasing protein content for feeding insects capable of transforming straw into animal feed and biolubricants, as well as by modulating (up and down) the lignin content to make barley straw suitable to produce mycelium-based construction panels (Alaneme et al., 2023), structural straw board (Amziane & Collet, 2017; Li et al., 2023; Tlaiji et al., 2022), and sandwich panels, as well as polymer composites (Bourmaud et al., 2018; Mohanty et al., 2018).

During more than one century of barley/wheat (Triticum ssp.) breeding, straw traits have been selected mainly to maximize yield and minimize lodging (semidwarf plants), while no attention has been given to straw composition. Although some straw transformation processes have been proposed, straw composition has never got attention from breeders and the current straw composition might not be the most suitable for every possible transformation. By tailoring straw composition to specific industrial needs, the project innovations will contribute to replacing non-renewable with renewable resources. The production of chemical lubricants has a high environmental impact that could be reduced by substituting oil-derived lubricants with biodegradable alternatives. Moreover, transforming straw into high-nutrition feed using insects could reduce the need for dedicated protein crops. The increase in root biomass and the use of straw as raw material in the construction and composite sector promote carbon sequestration contributing to mitigate the effects of climate change. By shifting to utilizing straw-based materials which are locally and widely available, BEST-CROP

will aid in decarbonizing relevant industries and will contribute to reducing their environmental impact.

These objectives will be driven forward based on highly innovative biotechnology approaches that exploit natural- and induced-genetic variation, gene editing and genetic engineering techniques for improving the photosynthetic efficiency, and building a next generation barley plant that could be exploited directly in breeding programs, while also serving as proof-of-concept of gene function.

## OPTIMIZATION OF CANOPY PHOTOSYNTHESIS PERFORMANCE

The efficiency of solar energy conversion into biomass, during photosynthesis, is surprisingly low in crops, typically below 1% in temperate climates. For this reason, numerous strategies have recently been proposed to improve overall photosynthetic efficiency under different growth conditions (Kromdijk et al., 2016; Simkin et al., 2015, 2019; Slattery & Ort, 2019, 2021; Smith et al., 2023; South et al., 2019). These studies have explored various innovative approaches, from enhancing light harvesting to introducing new biochemical pathways aimed at increasing carbon fixation efficiency. In synergy with other EU-funded projects such as CAPITALISE (https://www.capitalise.eu/), GAIN4CROPS (http://gain4crops.eu/) and PhotoBoost (http://www.photoboost.org/), BEST-CROP aims to exploit promising strategies, selected based on proof-of-concept in *Arabidopsis thaliana* (L.) Heynh., tobacco (*Nicotiana tabacum* L.) and a few crops, demonstrating that photosynthesis can be readily improved and translated into breeding programs (Table 1).

## Improved light distribution across the canopy

Light within a canopy is not absorbed homogeneously, with the leaves more exposed to light absorbing a large fraction, leaving the leaves underneath under light limitation. The reduction of leaf chlorophyll content has been demonstrated to be highly effective in improving light penetration in high-density mass canopy (Figure 2). Pale green crops can be created by manipulating a plethora of processes, such as the biogenesis and/or accumulation of antenna proteins (Light harvesting complex, Lhc)—also known as the Truncated Light-harvesting Antenna (TLA) strategy—and pigment biosynthesis (for a review, see Cutolo et al., 2023; Table 1). For instance, increased photosynthetic performance and enhanced plant biomass accumulation were observed upon cultivation at high density under greenhouse conditions of a pale green tobacco line with downregulated expression of *cpSRP43* encoding the 43-kDa chloroplast Signal Recognition Particle (*cpSRP43*; Kirst et al., 2018; see also Table 1). *cpSRP43* is a chaperone required for post-translational targeting of Lhc to the thylakoid membranes and contributes to the biogenesis and
<table><caption>**Table 1** Main proof-of-concepts, demonstrating that photosynthesis can be readily improved, inspiring BEST-CROP experimental plan</caption><thead><tr><th>Trait</th><th>Target gene(s)</th><th>Tested species</th><th>Gain</th><th>References</th></tr></thead><tbody><tr><td>Altering the composition of the thylakoid electron transport chain</td><td></td><td></td><td></td><td></td></tr><tr><td>Reduced antenna size</td><td>Downregulation of the 43-kDa chloroplast Signal Recognition Particle (cpSRP43)</td><td>Nicotiana tabacum L.</td><td>Plants with 10% greater leaf-to-stem ratio, and 8.2% more canopy biomass accumulation under high-density cultivation conditions</td><td>Kirst et al. (2018)</td></tr><tr><td></td><td>A premature stop codon in the 43-kDa chloroplast Signal Recognition Particle (cpSRP43)</td><td>Hordeum vulgare L.</td><td>A 50% reduction in the chlorophyll content of leaves does not cause any penalty on total biomass production and grain yield under standard field conditions</td><td>Rotasperti et al. (2022)</td></tr><tr><td>Faster NPQ relaxation</td><td>Overexpression of Violaxanthin DE-epoxidase (VDE), Zeaxanthin Epoxidase (ZEP), and the PsbS subunit of Photosystem II (PSII)</td><td>N. tabacum L.</td><td>15% greater plant biomass production in natural field conditions</td><td>Kromdijk et al. (2016)</td></tr><tr><td>Additional electron sinks</td><td>Overexpression of Violaxanthin DE-epoxidase (VDE), Zeaxanthin Epoxidase (ZEP), and the PsbS subunit of Photosystem II (PSII)</td><td>Glycine max (L.) Merr.</td><td>In replicated field trials, seed yield increased by up to 33%</td><td>de Souza et al. (2022)</td></tr><tr><td></td><td>Overexpression of Flavo-di-iron proteins FlvA-FlvB from Physcomitrium patens</td><td>Arabidopsis thaliana L.</td><td>Protect photosystems under fluctuating light</td><td>Yamamoto et al. (2016)</td></tr><tr><td></td><td>Overexpression of Flavo-di-iron proteins FlvA-FlvB from Physcomitrium patens</td><td>Oryza sativa L.</td><td>Increased resistance to PSI photodamage under fluctuating light</td><td>Wada et al. (2018)</td></tr><tr><td></td><td>Overexpression of cyanobacterial Flavo-di-iron proteins Flv1/Flv3</td><td>A. thaliana L.</td><td>Plants with 10-30% higher shoot dry weight</td><td>Tula et al. (2020)</td></tr><tr><td></td><td>Overexpression of Synechocystis Flavo-di-iron proteins Flv2-Flv4</td><td>A. thaliana L.; N. tabacum L.</td><td>Plants with increased tolerance toward high irradiation, salinity, oxidants, and drought</td><td>Vicino et al. (2021)</td></tr><tr><td>Photorespiration bypasses</td><td>Introduction of glycolate catabolic pathway from Escherichia coli</td><td>A. thaliana L.</td><td>Plants with at least 50% more root and shoot biomass, faster growth and more soluble sugars</td><td>Kebeish et al. (2007)</td></tr><tr><td></td><td>Introduction of three alternative glycolate catabolic pathways in combination with the downregulation of the Plastidal glycolate/glycerate translocator 1 (PLGG1)</td><td>N. tabacum L.</td><td>Plants with up to 24% increased biomass accumulation and improved light-use efficiency of photosynthesis by 17%</td><td>South et al. (2019)</td></tr><tr><td></td><td>Introduction of a catabolic pathway allowing complete oxidation of glycolate into CO<sub>2</sub> catalyzed by three rice enzymes, that is, glycolate oxidase, oxalate oxidase, and catalase</td><td>O. sativa L.</td><td>Plants with significant increases in photosynthesis efficiency, nitrogen content, and up to 35% more biomass production</td><td>Shen et al. (2019)</td></tr><tr><td>Excluding stomatal limitation</td><td>Overexpression of the plasma membrane H<sup>+</sup>-ATPase 1 (OSA1)</td><td>O. sativa L.</td><td>Plant with a 33% increase in grain yield and a 46% increase in N use efficiency</td><td>Zhang et al. (2021)</td></tr><tr><td>Modifying canopy architecture</td><td></td><td>O. sativa L.</td><td>Plants with more erect leaf angle, increased aboveground biomass, and grain yield at high planting densities.</td><td>Sakamoto et al. (2006)</td></tr><tr><td>More upright leaves</td><td>Loss of function mutant in DWARF4 gene encoding a cytochrome P450, CYP90B1, involved in brassinosteroid biosynthesis</td><td>Zea mays L.</td><td>Plants exhibit progressively more erect leaf angles from lower to upper leaves, increased fraction of penetrated PAR and net photosynthesis, increased grain yield per ha at high planting density</td><td>Tian et al. (2024)</td></tr></tbody></table>
<figure><img src="image_4.png" /><figcaption>**Figure 2.** Phenotypic characteristics expected to be observed in advanced breeding lines upon implementation of BEST-CROP experimental plan. Pale green barley lines (left side), combined with erect leaves will allow a better light penetration through the canopy (yellow arrow), with respect to dark green control plants (right side). These features will be stacked with faster adaptation to rapid light intensity changes (NPQ, red arrow), enhanced gas exchange, and photorespiratory bypasses, allowing an increased rate of CO₂ fixation and higher O₃ assimilation (light blue arrow). Image is courtesy of BEST-CROP partner, UMIL.</figcaption></figure>

maintenance of the thylakoid membranes and their associated electron transport chains (Klimyuk et al., 1999; Schuenemann et al., 1998). The corresponding *A. thaliana* mutant is known as *chaos* [chlorophyll a/b binding protein harvesting-organelle specific (Klimyuk et al., 1999)]. In the chloroplast stroma, cpSRP43 interacts with 54-KDa chloroplast Signal Recognition Particle (cpSRP54) to form the heterodimeric complex cpSRP. Lhc precursors imported into the stroma from the cytosol are N-terminally processed and bound by cpSRP to form a soluble cpSRP-Lhc complex termed the transit complex, which maintains Lhcs in an integration-competent state (for a review see Ziehe et al., 2018). The transit complex is needed to dock and integrate the Lhc proteins into the thylakoid membranes, with the help of the cpSRP receptor homolog, cpFtsY (Kogata et al., 1999; Tu et al., 1999) and the integral thylakoid membrane protein Albino3 (Alb3), a translocase that physically interacts with cpSRP43 (Moore et al., 2000). Recently, cpSRP43 was also shown to efficiently chaperone

and stabilize the glutamyl-tRNA reductase (GluTR), a rate-limiting enzyme in tetrapyrrole biosynthesis, enabling Lhc thylakoid insertion to be coordinated with Chl biosynthesis (Wang, Liang, et al., 2018). More recently, the barley mutant *happy under the sun 1* (*hus1*), which carries a premature stop codon in the corresponding *HvcpSRP43* gene and is characterized by a 50% reduction in the chlorophyll content of leaves, was shown to accumulate biomass and grains at levels comparable to those observed for the control cultivar Sebastian, when grown under field conditions at standard density (Rotasperti et al., 2022). These findings demonstrate that crops can indeed decrease their investment in antenna proteins and chlorophyll biosynthesis significantly, without detrimental effects on productivity (Table 1). In barley, a rich variety of mutants exists with a reduced amount of chlorophyll (Hansson et al., 2024). BEST-CROP will exploit the slight and variable reductions of leaf chlorophyll content observed in *chlorina* mutants. These are light green viable mutants, which can be kept in homozygous stocks. More severe mutants, such as yellow *xantha* mutants, white *albina* mutants, and light green *viridis* mutants, are lethal and not directly suitable for plant breeding. Still, they can be used to identify important genes to be explored for development of plants with a reduced leaf chlorophyll content.

In the "smart canopy" model, reduced chlorophyll content is combined with an optimized plant architecture comprising more erect leaf angles to improve the distribution of photosynthetically active radiation within the crop canopy, increase photosynthetic efficiency and reduce competition among neighboring plants (Ort et al., 2015); these features are designed to allow higher planting densities and increase grain yield (Figure 2). For example, the selection of maize (*Zea mays* L.) hybrids with upright leaves contributed to reaching current densities of over 80 000 plants per hectare in the US Corn Belt compared to 30 000 plants per hectare in the 1930s, which largely account for yield increases in the past decades (Assefa et al., 2018; Duvick, 2005; Tian et al., 2011; see Mantilla-Perez & Salas Fernandez, 2017 for a review). In grass leaves, the sheath wraps around the stem while the leaf blade or lamina projects away from the stem to facilitate light capture: the leaf insertion angle is defined as the angle between the blade and the vertical stem axis and is regulated by multiple genetic and hormonal factors with brassinosteroids playing a major role. Breakthrough studies demonstrated that specific mutations in orthologous brassinosteroid biosynthesis genes *DWARF4* in rice (*Oryza sativa* L.) and *Leaf Angle Architecture of Smart Canopy 1* (*LAC1*) in maize result in reduced leaf angles and increased grain productivity under high-density planting (Sakamoto et al., 2006; Tian et al., 2024). In particular, a detailed analysis of the maize *lac1* mutant under field conditions showed that optimized reduction of leaf angles at different canopy
levels leads to increased penetration of sunlight to the lower canopy and improved photosynthesis (Tian et al., 2024). A similar result was obtained in sugarcane (Saccharum spp. hybrid) where the fine-tuning of leaf angle through genome editing of the *LIGULELESS* 1 gene (*LG1*) has optimized light capture and increased dry biomass yield up to 18% without having to add more fertilizer (Brant et al., 2024). In barley, plants with a reduced leaf angle and an overall erect architecture are especially found among the *Brachytic*, *Erectoides*, and *Breviaristatum* mutant groups (Hansson et al., 2024; see also Table 1).

## Faster responses to changes in irradiance

Under full sunlight, plants dissipate the excess of potentially damaging absorbed light energy by inducing the photo-protecting mechanism termed nonphotochemical quenching (NPQ). This process is essential to avoid the formation of reactive oxygen species (ROS) that would damage the photosynthetic apparatus. However, NPQ activation/deactivation kinetics are too slow to adapt to the rapid fluctuations of light intensity within crop canopies, due to changing cloud cover and leaf movement. Optimized plant response to these dynamics has been shown to improve photosynthetic efficiency and productivity with Violaxanthin DE-epoxidase (VDE), Zeaxanthin Epoxidase (ZEP), and the PsbS subunit of Photosystem II (PSII) playing major roles. Kromdijk et al. (2016) demonstrated, for instance, that upregulations of *VDE*, *ZEP*, and *PbsS* significantly accelerate the kinetics of photo-protection induction and relaxation in tobacco, and this resulted in an increased efficiency of CO₂ assimilation in fluctuating light conditions, which translated into a 14–21% increase in shoot biomass production in replicated field trials (Figure 2; Table 1). More recently, the same change in an elite soybean (*Glycine max* L. Merr.) variety increased seed yield by up to 33% in small-scale field trials (de Souza et al., 2022), pointing to NPQ as a novel potential target for increasing photosynthesis efficiency and crop yield. However, these results were not replicated in other species such as Arabidopsis (Garcia-Molina & Leister, 2020) and potato (*Solanum tuberosum*; Lehretz et al., 2022), indicating that the complexity of light environmental conditions requires tailored and species-specific optimization of the response. BEST-CROP will attempt to ameliorate NPQ kinetics by exploiting barley natural genetic variability collected at the Cranachan database (https://barley.hutton.ac.uk/), as well as barley mutant populations available at the James Hutton Institute (Caldwell et al., 2004; Schreiber et al., 2019).

An additional strategy to minimize the over-reduction of photosynthetic electron transporters in response to light dynamics and the consequent generation of ROS evolved in cyanobacteria, algae, non-vascular plants, and gymnosperms but not in angiosperms. This photoprotective strategy relies on Flavo-di-iron proteins (FLVs; Alboresi

et al., 2019), which accept electrons downstream of Photosystem I (PSI) to reduce oxygen to water, decreasing the potential formation of ROS. FLV role is particularly important after an abrupt change in illumination intensity, which causes an imbalance between the photosynthetic electron transport, which instantaneously responds to changes in illumination and NADPH consumption that, instead, depends on the metabolic activity and thus requires several minutes to adjust. Consistent with this activity, mutants depleted in FLV show strong photosensitivity when exposed to light fluctuations, as experienced under field conditions.

Recently, FLV proteins have been shown functional when expressed in Arabidopsis (Yamamoto et al., 2016) and rice (Wada et al., 2018), meaning that their activity does not require specific accessory components absent in angiosperms. Moreover, FLV expression in Arabidopsis (Tula et al., 2020) and tobacco (Vicino et al., 2021) was shown to induce a higher tolerance to abiotic stresses, even though the impact on yield under field conditions was never fully assessed (Table 1). BEST-CROP aims at expressing heterologous FLVs in barley and verifying whether this strategy could improve crop productivity under natural light fluctuations, especially in conditions such as drought stress, where CO₂ fixation is limited, and the availability of additional electron acceptors could be beneficial.

## Carbon-positive photorespiration bypass routes

To develop a carbon-positive photorespiration bypass pathway, enzymes that do not occur naturally in barley are introduced through *Agrobacterium*-mediated transformation. This will alleviate the impact of the double activity of RuBisCo that not only catalyzes the carboxylation of ribulose 1,5-bisphosphate but also its oxygenation (for a review see Smith et al., 2023). The oxygenation reaction reduces the photosynthetic efficiency and, thus, the plant's yield. The oxygenation activity of RuBisCo produces 2-phosphoglycolate (2PG), which must be recycled by photorespiration. This results in a yield loss of 20–50% of the previously assimilated carbon. BEST-CROP aims to reduce the yield loss due to photorespiration by 50% by using alternative bypass pathways for photorespiration. Independent research groups have shown that such bypasses can significantly increase yield under laboratory and field conditions in various crops and model plants and offer a promising solution to this challenge (Kebeish et al., 2007; Maier et al., 2012; Shen et al., 2019; South et al., 2019; see also Table 1).

Two alternative bypasses of photorespiration are exploited in BEST-CROP (Figure 3). First, the complete oxidation of 2-phosphoglycolate to CO₂ by the sequential action of phosphoglycerate phosphatase native to barley chloroplasts, glycolate dehydrogenase from
<figure><img src="image_5.png" /><figcaption>**Figure 3.** Summary of the two alternative bypasses of photorespiration exploited in BEST-CROP. (1) In the first bypass, the complete oxidation of 2-phosphoglycolate to CO₂ will be obtained through the sequential action of phosphoglycerate phosphatase (PGH) native to barley chloroplasts, glycolate dehydrogenase (GDH) from *Chlamydomonas reinhardtii* mitochondria, malate synthase (MS) from *Cucurbita maxima* (pumpkin) peroxisomes, endogenous NADP malic enzyme, and finally endogenous pyruvate dehydrogenase (PDH). In addition, RNAi-mediated reduction (indicated by dotted lines) of the plastid glycolate glycerate translocator 1 (PLGG1) reduces glycolate export from the plastids. (2) The second bypass will allow to achieve carbon-neutral conversion of glycolate to oxaloacetate through a plastid-targeted version of the beta-hydroxyaspartate shunt (BHAC). This includes glycolate dehydrogenase (GDH) from *C. reinhardtii* mitochondria, aspartate:glyoxylate aminotransferase (AGAT), β-hydroxyaspartate aldolase (BHAA), β-hydroxyaspartate dehydratase (BHAD), and iminosuccinate reductase (ISR). BASS6, bile acid sodium symporter. Endogenous genes are in black, while heterologous genes are in red. Image created in https://BioRender.com.</figcaption></figure>

*Chlamydomonas reinhardtii* mitochondria, malate synthase from *Cucurbita maxima* Duch. (pumpkin) peroxisomes, endogenous NADP malic enzyme, and finally endogenous pyruvate dehydrogenase. In addition, RNAi-mediated reduction of the Plastidial glycolate glycerate translocator 1 (PLGG1) reduces glycolate export from the plastids. Variations of this strategy have been successfully used in several crops and model plants and are considered a low-risk baseline option (Kebeish et al., 2007; Maier et al., 2012; Shen et al., 2019; South et al., 2019). Second, we will achieve carbon-neutral conversion of glycolate to oxaloacetate through a plastid-targeted version of the beta-hydroxyaspartate shunt (BHAC). This includes glycolate dehydrogenase from *C. reinhardtii* mitochondria, aspartate: glyoxylate aminotransferase, β-hydroxyaspartate aldolase, β-hydroxyaspartate dehydratase, and iminosuccinate reductase. This pathway was developed as part of the EU H2020 GAIN4CROPS project, and proof-of-concept for a peroxisome-targeted version in Arabidopsis was provided (Roell et al., 2021). Unpublished work suggests a higher efficiency of a plastid-targeted beta-hydroxyaspartate shunt.

Based on this, we will introduce a plastid-targeted version in barley by replacing the peroxisome-targeting signals with plastid transit peptides. This strategy carries a higher risk with a potentially high gain and complements the lower-risk bypass described above.

## Excluding stomatal limitations for CO₂ and O₃ uptake

Stomatal pores formed by the guard cells balance plant water loss by evapotranspiration with CO₂ uptake for photosynthetic carbon assimilation. Vegetation also absorbs significant amounts of air pollutants, such as ozone, mainly through stomatal pores (Brosché et al., 2010). The central role of stomata in controlling plant gas exchange processes makes them an attractive target to enhance photosynthetic efficiency and drought tolerance as well as to manage ozone uptake in crops. Several strategies employing stomata for improving crop productivity have been suggested; these include altering stomatal density and size (Caine et al., 2023; Hughes et al., 2017), accelerating stomatal movements (McAusland et al., 2016; Papanatsiou et al., 2019), and increasing stomatal conductance for
elevated CO₂ uptake by overexpressing proton pumps (Zhang et al., 2021; Table 1). Enhanced stomatal conductance will also ensure improved ozone uptake, although it is important to maintain stomatal responsiveness to stress factors, such as drought, in these plants. Such a strategy could be instrumental in developing crops that help to improve the air quality in regions with a high level of air pollution (Diener & Mudu, 2021; Sicard et al., 2018). In BEST-CROP, we aim to develop barley lines with moderately increased stomatal conductance, to improve CO₂ and ozone uptake while maintaining stomatal responsiveness to drought conditions. We will target barley proton pumps providing energy for stomatal opening; we will either overexpress proton pumps or introduce mutations that disrupt their autoinhibition mechanism. Such barley lines will have enhanced stomatal conductance ensuring maximal supply of CO₂ for photosynthesis and elevated ozone uptake by mesophyll (Kollist et al., 2000). At the same time, these lines will maintain an intact stomatal closure in response to drought stress hormone, abscisic acid (ABA), as demonstrated in Arabidopsis (Figure 4). Alternatively, lines with enhanced stomatal conductance but unaffected ABA/drought responses can be obtained by modulating activities of the proteins involved in CO₂ sensing in guard cells (Takahashi et al., 2023; Yeh et al., 2023).

To achieve this, we will identify gain-of-function mutants for HIGH LEAF TEMPERATURE 1 (HT1; Hörak

et al., 2016) and knockouts for the barley MITOGEN-ACTIVATED PROTEIN KINASE 12 homolog (MPK12; Töldsepp et al., 2018) in mutant collections. Stomatal traits in these novel barley lines will be thoroughly characterized by using the custom-made whole plant gas exchange devices (Hörak et al., 2017; PlantInvent Ltd).

Cumulative uptake of ozone and its possible toxicity will be estimated upon exposure to ozone concentrations observed in Europe's air-polluted regions, together with measurements of photosynthesis and stomatal behavior in various environmental conditions. As a potential remedy to mitigate ozone toxicity, overexpression of enzymes involved in scavenging reactive oxygen species, such as superoxide dismutase, can be applied (van Camp et al., 1994).

## IMPROVING PHOTOSYNTHESIS BY EXPLOITING INDUCED MUTANTS IN COMBINATION WITH GENE EDITING AND GENETIC ENGINEERING

Currently, most biotechnological approaches to improve photosynthetic performance rely on mutants as well as on gene editing and genetic engineering approaches. This is possible due to the knowledge gained in more than three decades of functional genomics studies on photosynthesis using model organisms in combination with the ability to model photosynthetic systems to identify critical proteins and enzymes that control photosynthetic efficiency. For instance, the improvement of canopy photosynthesis via

<figure><img src="image_6.png" /><figcaption>**Figure 4.** The gain-of-function mutation in the proton pump AtAHA1 increases stomatal conductance and ozone (O₃) uptake but does not affect the stomatal responsiveness to abscisic acid (ABA) in Arabidopsis.</figcaption></figure>

The 3 to 4 weeks-old plants were incubated in the gas exchange device (Kollist et al., 2007) and were subjected to spraying with 5 µM ABA (a) or 3-min 450 nM L⁻¹ ozone pulse (b). The *ost2-2D* mutant (Merlot et al., 2007) was compared with wild type plants (Col-0). The starts of the treatments are shown by the arrows. In another experiment, the Col-0 and *ost2-2D* plants were exposed to continuous 220 nM L⁻¹ ozone. The cumulative dose of absorbed ozone is shown (c). The values are averages ± SE (n = 3-4).
the faster recovery of NPQ was predicted, initially, through a dynamic system model of canopy photosynthesis (Zhu et al., 2004), and later validated in both soybean and tobacco (de Souza et al., 2022; Kromdijk et al., 2016). Similarly, modeling of photorespiratory bypass suggested that decreased expression of *PLGG1*, a glycolate/glycerate transporter (Pick et al., 2013), could result in an increased benefit of photorespiratory bypass and further increased photosynthetic CO₂ uptake rate (Xin et al., 2015), which was later experimentally confirmed in tobacco (South et al., 2019) and rice (Shen et al., 2019). Exploiting intraspecific natural genetic diversity to improve photosynthesis is, instead, still in its infancy. Limitations are imposed by the minor variation in the basic physiological and biochemical mechanisms of the photosynthetic engine within species, as shown by the fact that the key components of photosynthesis [i.e., the subunits of PSII, PSI, and the Calvin-Benson-Bassham cycle (CBB)] are highly conserved. Higher rates of photosynthesis have been shown, indeed, to correlate with higher amounts of Cyt *b*<sub>6f</sub> complex or RuBisCo per unit area, and not by the identification of Cyt *b*<sub>6f</sub> or RuBisCo with higher activity (Miller et al., 2017). Similarly, the accelerated kinetics of NPQ and the improved efficiency of CBB have been obtained by overexpression approaches aimed at increasing the accumulation of specific enzymes in several species, including Arabidopsis, tobacco, rice, soybean, and wheat (for a review see Croce et al., 2024).

On the contrary, a wide diversity of photosynthetic properties and the main components of the photosynthetic apparatus is found in nature across species, and examples of increased photosynthetic carbon assimilation and biomass production in model plants and crops, obtained by overexpression of genes from other species, such as the bifunctional cyanobacterial *sedoheptulose-1,7-biphosphatase (SBPase)/fructose-1,6-biphosphatase (FBPase)* and algal *Cyt c*<sub>6</sub> (López-Calcagno et al., 2020), as well as *FLVs* (Tula et al., 2020; Vicino et al., 2021; Wada et al., 2018; Yamamoto et al., 2016), have been reported. This evidence, together with the improved photosynthetic performance obtained through engineered synthetic pathways to bypass photorespiration (for a review see Smith et al., 2023), highlight the relevance of mutants, as well as of gene editing and genetic engineering approaches for increasing crop production. In particular, the gene editing technology, namely CRISPR/Cas, is still evolving, with new enhancements in editing precision, multiplex editing, and gain-of-function strategies, although challenges remain in optimizing CRISPR delivery methods and transformation efficiency (Ahmar et al., 2024). Recently, the European Union has taken steps toward embracing New Genomic techniques (NGTs), with a new regulation passing its first vote in the European Parliament (Katsarova, 2024), although NGTs will still have to go through several approvals before they can be used for crop breeding also in Europe.

Currently, the adoption of these different biotechnological strategies led to increases in biomass accumulation ranging from 10 to 68%, although most of the studies were conducted in greenhouses or small-scale field trials (Khaipho-Burch et al., 2023). Moreover, no data are available on the adoption of these approaches in different combinations, albeit it is anticipated that altering crucial pathways in plant metabolism, including light conversion and carbon fixation, will inherently have interdependent effects that require additional investigation, through predictive mathematical models and experimental validation of predictions. Noteworthy, even a modest improvement of photosynthesis can lead to a marked increase in productivity according to the Blackman model (Blackman, 1919).

BEST-CROP is aimed to fill this gap, by combining these strategies into barley elite germplasms and by evaluating their performance under field conditions, taking care to adhere to well-established testing methods. This includes standard definitions of yields, obtained through field trials replicated across plots, geographical locations, and years, where agriculture practices closely match the conditions of the farms that will ultimately produce the crop (Khaipho-Burch et al., 2023). It is reasonable to expect that under highly dynamic environmental conditions, in the presence of environmental stressors and pathogens, the increase in productivity of the novel barley lines will be less than the ones observed under controlled conditions and closer to what breeders consider true breakthroughs in crop productivity, that is, yield increases of the order of 5-10%, although combined improvements could potentially yield an additive outcome, maximizing the overall impact on productivity.

## IMPROVING PHOTOSYNTHESIS IN CONNECTION WITH THE OPTIMIZATION OF STRAW QUALITY FOR THE CIRCULAR BIOECONOMY

In the last decade, several research programs have investigated the possibility of valorizing annual plants in various industrial sectors. The BFF project (Biomass For the Future; https://anr.fr/ProjetIA-11-BTBR-0006), for instance, aimed to optimize miscanthus (*Miscanthus Andersson*) and *Sorghum* production and develop new end-uses of these bio-masses for energy (anaerobic digestion) and materials (polymer composites for automotive and construction materials). AGRIMAX (Multiple high-value products from crop and food-processing waste; https://agrimax-project.eu), instead, had the objective to develop and demonstrate the production of multiple, high-value products (packaging, food additives, fertilizers) from various crops and food-processing wastes. In addition, GRACE (GRowing Advanced industrial Crops on marginal lands for biorEnergies; https://www.grace-bbi.eu/) had the ambition to optimize various value chains for the bio-based industry from miscanthus and hemp (*Cannabis sativa* L.) grown on
marginal lands, that is, polluted or non-arable. Despite stimulating and interesting outcomes, existing major crops such as barley (60 million ha of world arable land), and wheat (240 million ha in the world) were not at the center of these projects, which seems critical given the huge amount of residual straw involved. On the other hand, research projects investigating novel biotechnological strategies able to increase crop photosynthesis performance and total biomass production and grain yield (see also recent EU-funded projects such as CAPITALISE, GAIN4CROPS, and PhotoBoost) did not take into consideration the need to tailor the quality of the residual straw to be transformed into new bio-based products.

In BEST-CROP, the optimization of canopy photosynthesis performance will be coupled with the improvement of barley straw quality, closing the loop on resource use, and waste generation in agro-ecological systems. While cereal straw is a relevant postharvest by-product that can be used as an ideal ground cover to prevent soil erosion or incorporated into the soil to improve soil organic matter, it is estimated that, at a grain yield >6 ton/ha, around 60% of barley straw can be removed from soil without depleting soil organic carbon (Johnson et al., 2006). In addition, straw removal from fields prevents nitrogen sequestration by the decomposing straw (Fontaine et al., 2020), further supporting the use of straw for developing high-value bio-based materials and compounds.

## Barley straw with different cellulose/lignin contents and lignin properties for the construction and biocomposite sectors

Over the last two decades, there has been a widespread global campaign toward the development and utilization of more sustainable manufacturing materials obtained from plant fibers to ensure a successful transition from a linear economy model to a circular, sustainable bioeconomy (Alaneme et al., 2023; Butu et al., 2020).

Currently, the use of non-wood plant fibers as supplements or as direct substitutes for wood in particleboards and as reinforcements in biocomposites, that is, a broad class of materials in which natural fibers, used as reinforcement, are combined with a petroleum- or bio-based matrix to form a composite material with superior properties, have been identified as sustainable alternatives to conventional synthetic materials because of their competitive properties and environmentally friendly characteristics (Ahmad et al., 2022). In the case of agricultural residues (e.g., cereal straws, bagasse) for instance, their use should allow reduced consumption of raw materials from petroleum and/or forestry resources and are rapidly renewable.

Some examples of plant-based particleboards and biocomposites are already available in the building and construction sector, food packaging, sports, and leisure and automotive industries, driven by the major

contribution of these sectors to environmental issues (Figure 5). These industries are, indeed, responsible for a large percentage of carbon emissions and the generation of non-biodegradable waste, which in turn has adverse effects on ecosystems (Haneef et al., 2017; Peñaloza, 2017; Siddique et al., 2016; Yang et al., 2021). Despite the many factors favoring the use of plant-based particleboards and biocomposites, there is still some way to go before they are considered reliable alternatives to conventional materials in terms of market uptake. Indeed, their industrial use requires reproducibility and consistency in the morphological and mechanical properties of plant fibers, which are influenced by various cell wall characteristics, including the concentration of cellulose and non-cellulosic components (i.e., lignin, hemicelluloses, pectins, proteins), microstructure (microfibrillar angle, MFA of cellulose microfibrils), and hydrogen bonds and crosslinks between cell wall components (Bourmaud et al., 2018; Mohanty et al., 2018). One reason for apparent lack of consistency is that plant cell walls and fibers from different crops (and to a lesser extent from different cultivars of the same crop) can have wide diversity in morphology, composition, and mechanical properties. This relates to their differing function within plants, for example the soft seed fibers of cotton help the seeds to disperse and are made of pure cellulose, while stem structural fibers can be flexible with low lignin content (e.g., the phloem bast fibers of crops like flax, hemp, and ramie) or inflexible with relatively high lignin for example in the straw of cereal crops or in wood. Little research has been performed to directly link cell wall properties in a given species and cultivar with the performance of biocomposites produced from that plant material. This means that it is currently not possible to infer, except in the most general terms, how a plant material will perform or how it might be improved for specific biocomposite applications. Better understanding of the relationships between physical properties of barley straw-based materials and cell wall properties is thus necessary and this is precisely what is proposed in BEST-CROP to inform the use of barley straw in different applications (Figures 5 and 6).

Flax is a crop currently being used in biocomposites due to its long bast fibers with particularly low lignin and high cellulose content. At the scale of flax elementary fiber cells or fiber bundles, the mechanical properties result from the complex biomolecular assemblies of cellulosic and non-cellulosic components and their organization within the cell walls. In particular, it has been shown by the selective removal of non-cellulosic components from flax fibers that these play a key role in the mechanical behavior of the fibers (Lefeuvre et al., 2015). The authors assumed that non-cellulosic components, that is, matrix and structuring polysaccharides, contribute to the cohesion of cellulose microfibrils and to load transfer within cell
<figure><img src="image_7.png" /><figcaption>**Figure 5.** Examples of high-value bio-based industrial products that can be obtained from the transformation of barley straw. (a) The use of BSFL (black soldier fly larvae) can transform barley straw into high-quality protein and fat-rich biomass suitable for animal feed and biolubricant production; (b) Straw with various lignin content and properties can be used as raw material for manufacturing mycelium-based insulation panels, and (c) straw-based panels and composites. Images are courtesy of BEST-CROP partners: FRD-CODEM and IMT Mines Alès (straw-based panels and composites); MOGU srl (mycelium-based insulation panels); SO.G.I.S. Industria Chimica SpA (animal feed and biolubricants); S.I.S. Società Italiana Sementi (barley straw).</figcaption></figure>

<figure><img src="image_8.png" /><figcaption>**Figure 6.** Physical properties of straw-based materials. Mechanical, flammability, thermal and acoustic insulation, and hygroscopicity properties of mycelium-based insulation panels and straw-based panels and composites to be studied in BEST-CROP in relation to the different barley cultivars selected and the intrinsic characteristics of the cell walls.</figcaption></figure>

walls, which can greatly affect the tensile stiffness and strength of the fibers.

Variations in stiffness can occur even between differ-
ent genotypes of the same crop, for example in a study of
stem fragments from six different miscanthus genotypes
with contrasting biochemical compositions, variations in
stiffness were related to their biochemical composition,

that is, lignin, hemicelluloses, but also *p*-coumaric and
ferulic acids that are proposed to play a role in both cross-
linking between cell wall components and inter-cellular
cohesion (Chupin et al., 2020).

At the composite scale, using stem fragments from
different miscanthus and sorghum genotypes with varying
biochemical and microstructural features of the cell walls
was also found to have a significant influence on the
mechanical performance of composite materials. In partic-
ular, improvements in the tensile stiffness and strength of
miscanthus-reinforced composites was shown to be corre-
lated with increases in cellulose, lignin, and p-coumaric
acid content (Brancourt-Hulmel et al., 2021; Chupin
et al., 2017; Di Giuseppe et al., 2021). Variations in bio-
chemical composition and histological organization among
genotypes are also responsible for different breakage
mechanisms of stem fragments during composite proces-
sing, leading to variations in their mechanical properties
related to their microstructure, and the distribution of fiber
size and shape within the composites (Vo et al., 2017).
Moreover, it has been shown for miscanthus-based con-
crete that sugar-based molecules, in particular xylose and
glucose, can hinder the cement hydration process due to
their release into the cement-water-sand mixture and their
adsorption onto cement particles, which is detrimental to
the resulting mechanical strength of concrete blocks (Boix
et al., 2020). These studies on plant fibers and their use as
reinforcements in composite and concrete materials high-
light the need to better identify the key biological traits and
related cell wall characteristics that should be optimized
for better selection of high-performance plant fibers and
genotypes for material and structural applications.

Flammability of lignocellulosic fibers or stem frag-
ments is also strongly related to their biochemical compo-
sition, in particular to the quantities and nature of
polysaccharides and lignin, although their fire properties
cannot be easily deduced due to the existing interactions
between these components (Dorez et al., 2014). Lignin is a
high-charring polyphenol that decomposes slowly over
a wide temperature range, while cellulose decomposes
more rapidly with a peak mass loss centered at around
360°C and leaving only a small residue. When present in
significant amounts and depending on their chemical
structure, extractives can also influence the flammability.
Therefore, the energy released in case of combustion may
vary to some extent. Moreover, the above-mentioned
parameters influence the fire behavior of composite mate-
rials incorporating these bioresources. For example, in bio-
based concretes, in which stem fragments are the only
organic components likely to burn, the risk of ignition
greatly varies depending on the cell wall characteristics of
the bioresource used (Lopes et al., 2024).

BEST-CROP addresses this challenge by modulating
lignin/cellulose content and lignin properties to determine
the most advantageous composition for efficient imple-
mentation of barley straw in high-value bio-based indus-
trial products for the construction, and composites sectors.
Milled barley straws in the form of fibers, granulates, and
flour, will be used as substrates for inoculation and coloni-
zation by fungal strains, and the manufacture of straw-
derived mycelium-based construction panels targeting

thermal and acoustic insulation for wall, floor, and ceiling
tiles (Alaneme et al., 2023; Attias et al., 2020; see also Fig-
ure 6). BEST-CROP will also consider the valorization of
barley straw for the manufacture of straw-based particle
boards with the use of bio-based binders with enhanced
thermal insulation and mechanical properties (Amziane &
Collet, 2017; Li et al., 2023; Mahieu et al., 2021; Nakanishi
et al., 2018; Tlaiji et al., 2022; Uemura Silva et al., 2021), as
well as the use of straw as reinforcements in thermoplastic
processes for the manufacture of polymer composites
(Bourmaud et al., 2018; Mohanty et al., 2018). To this end,
BEST-CROP will exploit the natural variability in straw lig-
nin content observed to vary from 18.96 to 27.8% when
hundreds of elite spring barleys were phenotyped (Halpin
et al., unpublished). Moreover, prior work demonstrates
how lignin content can be reduced by suppressing lignin
biosynthesis genes (Table 2). For instance, a range of
transgenic barley plants have been produced where most
genes on the lignin biosynthesis pathway have been indi-
vidually suppressed or knocked out using RNAi or CRISPR-
Cas9 mutagenesis (e.g., Daly et al., 2019; Shafiei
et al., 2023). Genes manipulated include phenylalanine
ammonia-lyase (PAL), coumarate 3-hydroxylase (C3H),
cafe ic acid 3-O-methyltransferase (COMT), 4-coumarate-CoA
ligase (4CL), cinnamoyl-CoA reductase (CCR), ferulate 5-
hydroxylase (F5H), and cinnamyl alcohol dehydrogenase
(CAD). Lignin composition and structure were dramatically
altered in all lines and lignin content was either unchanged
or reduced. Downregulation of F5H substantially altered
the structure of lignin by changing the ratio of syringyl-to-
guaiacyl monomers in the polymer, but the mechanical
properties and yield were unchanged illustrating the plas-
ticity of the polymer and opportunities to tailor it to
improve lignocellulosic biomass as a feedstock for green
chemistry and specific straw applications (Shafiei
et al., 2023). For example, COMT RNAi lines, like F5H RNAi
lines, had dramatically reduced syringyl-to-guaiacyl mono-
mer ratio (reduced by 50%), a 15-fold increase in the
amount of 5-hydroxyguaiacyl units, but also had moder-
ately reduced lignin content (up to 15% reduction) and
some improvement in digestibility (Daly et al., 2019).
Improved digestibility should mean that sugars can be
more easily hydrolysed from the straw biomass as a sub-
strate for growing black soldier fly larvae (see next section)
or by mycelium-generating fungi producing architectural
panels. However, defining the best ideotype of barley for
industrial purposes in terms of straw composition (cellu-
lose, lignin, and other cell wall components) must await
the results of BEST-CROP after straws of different composi-
tions are tested in various applications. To manipulate lig-
nin even further and, for example, to increase lignin
content, different approaches can be exploited. The chal-
cone synthase colourless2 (C2) gene, controls the flux
toward flavonoid biosynthesis in maize vegetative tissues
and, when silenced, flux is redirected in lignin monomer synthesis causing strikingly higher lignin content in leaves (Eloy et al., 2017). Similarly, some *brittle culm (bc)* mutants of rice with altered cellulose deposition have increased levels of lignin, for example, the *bc1* mutant affected in a *COBRA-like* gene (Li et al., 2003), *bc10* a mutant in a Domain of Unknown Function 266 (DUF266)-containing type-II membrane protein (Zhou et al., 2009), and *bc12* a kinesin-4 protein (Zhang et al., 2010). Another novel way to manipulate cell wall properties is to alter the amount of ferulic acid involved in cross-linking between arabinoxylan and lignin, and the amount of *p*-coumaric acid also bound to arabinoxylan and lignin. A natural mutation that achieves this by knocking out a *p*-coumaroyl *acyltransferase* (PAT) gene in barley grain has been discovered, recently (Houston et al., 2023).

## Barley straw with increased protein amount for the feed and green chemistry sectors

Low-grade organic waste, as in the case of cereal straws, can also be converted at a high rate, by the fast-growing black soldier fly larvae (BSFL; *Hermetia illucens* L.), into high-quality protein (41–54% of dry matter) and fat-rich biomass (11.8–41.7% of dry matter) suitable for animal feeding and biolubricant production, contributing to recycling nutrients from the environment in the frame of the circular bioeconomy (Oonincx & de Boer, 2012; Seyedalmoosavi et al., 2022). The EU Commission has set a target of 20% of chemicals from renewable sources by 2030 (COM 2021/800/EC—Sustainable Carbon Cycles), and oleo-chemistry is the most appropriate technology to foster the transition from mineral oil chemistry to chemistry from renewables. Palm oil, an essential raw material for oleo-chemicals production, is under issue because of deforestation and, in general, it is considered a non-sustainable or a scarcely sustainable raw material. Insects can be a useful alternative, and BSFL accumulate large amounts of fat with a prevalence of saturated fatty acids (76% of total fat, including C12 and C16 chains; Renna et al., 2024), making its oil composition similar to that of palm oil. In addition, the aminoacidic composition of BSFL proteins makes them suitable as an alternative protein source with respect to soybean meal and fishmeal. Leucine, lysine, and valine are the most abundant amino acids in BSFL proteins, and they are more abundant than in soybean and fishmeal, while the less abundant essential amino acids, methionine, and tryptophan, are comparable to soybean (Lu et al., 2022). Thanks to its aminoacidic profile, BSFL has been used as a partial replacement for soybean and fishmeal in pig, broiler, and fish diets (Chia et al., 2019; Dabbou et al., 2018; Nogales-Mérida et al., 2019; Yu et al., 2019).

Larvae of BSF are highly efficient in converting organic waste, including agricultural residues, into

biomass. However, residues such as the lignocellulosic cereal straw do not contain enough protein to support BSFL fast growth (Fuso et al., 2021) and enriching them by adding vegetable proteins (e.g., soybean), strongly increases costs. BEST-CROP aims at developing advanced barley cultivars with increased straw protein content, thus improving its nutritional quality and nutrient balance as a component of the BSFL feeding substrate.

Natural genetic variation for straw protein accumulation remains largely unexplored, with limited available data estimating a 2–6% crude protein content (Keno et al., 2021; Wamatu et al., 2019). Recently, ICARDA barley breeding programs evaluated crude straw protein content as a valuable selection parameter on over 200 breeding lines, confirming a straw protein content ranging from 3 to 6.5%, with a heritability of 0.51 and a weak negative correlation with aboveground biomass (Sanchez-Garcia et al., unpublished; Table 2). Similarly, little is known about nitrogen partitioning in the different organs of barley plants (Barmeier et al., 2021) and the differences between malting and feeding barley, selected for low/moderate and high grain protein content, respectively, in their ability to mobilize nitrogen from the stem to grains. Genes involved in nitrogen use efficiency (NUE) or protein remobilization from leaves to developing grains, can be identified in close crop species from literature data, and they can be considered as possible targets for improving barley straw protein content. For instance, loss of function mutations in the rice abnormal cytokinin response1 repressor1 (OsARE1) gene have been associated with improved N utilization (Wang, Nian, et al., 2018). Moreover, CRISPR/Cas9-mediated editing of the barley ortholog HvARE1 caused delayed senescence and increased content of total chlorophyll in the flag leaf at the grain filling stage. Under low nitrogen conditions, barley are1 mutants showed high nitrogen content in shoots (Karunarathne et al., 2022). BT2, a member of the Bric-a-Brac/Tramtrack/Broad gene family, represses the expression of nitrate transporters, thus playing a central role in the NUE network. Mutations of this gene in rice increased NUE by 20% compared with wild type under low N conditions (Araus et al., 2016). The rice Lysine-Histidine-type Transporter 1 (OsLHT1) is involved in the translocation of amino acids from vegetative to reproductive organs for grain yield and quality. The concentrations of total N in the flag leaf at maturation were higher in OsLht1 mutants than wild type rice plants, but a reduction of panicle length, seed setting rate, and total grain weight was also observed (Guo et al., 2020). BEST-CROP will investigate allelic variation for these genes within available barley germplasm collections and TILLING populations, and will generate novel mutants through CRISPR/Cas9, with the final aim of improving the protein content of barley straw and promoting its use as substrate for BSFL growth.
<table><caption>**Table 2** Main proof-of-concepts, demonstrating that straw quality can be modified</caption><thead><tr><th>Trait</th><th>Target gene(s)/germoplasm(s)</th><th>Tested Species</th><th>Level of accumulation</th><th>References</th></tr></thead><tbody><tr><td>Straw/stover with different cellulose/lignin contents and lignin properties</td><td></td><td></td><td></td><td></td></tr><tr><td>Reduced lignin</td><td>Downregulation or mutation of 4-coumarate-CoA ligase (4CL)</td><td>Oryza sativa L.; Sorghum bicolor (L.) Moench</td><td>16-20% lignin reduction in both species; 17% increase in sugar yield on saccharification demonstrated in sorghum</td><td>Saballos et al. (2008, 2012) and Gui et al. (2011)</td></tr><tr><td></td><td>Downregulation or mutation of cinnamoyl-CoA reductase (CCR)</td><td>Zea mays L.; Lolium perenne L.</td><td>Lignin content reduction by 9-37%, with 14-15% improved digestibility; up to 40% improved glucose yield on saccharification in maize</td><td>Tu et al. (2010), Tamasloukht et al. (2011) and Smith et al. (2017)</td></tr><tr><td>Increased lignin</td><td>Mutation in chalcone synthase (CHS)</td><td>Z. mays L.</td><td>Increased leaf lignin and reduced leaf saccharification</td><td>Eloy et al. (2017)</td></tr><tr><td></td><td>Mutation in brittle culm1 (bc1) encoding COBRA-like protein</td><td>O. sativa L.</td><td>Increased lignin and reduced cellulose in developing sclerenchyma</td><td>Li et al. (2003)</td></tr><tr><td></td><td>Mutant (bc10) in a Domain of Unknown Function 266 (DUF266)-containing type-II membrane protein</td><td>O. sativa L.</td><td>Increased lignin and reduced levels of cellulose and arabinogalactan proteins (AGPs)</td><td>Zhou et al. (2009)</td></tr><tr><td></td><td>Mutation in brittle Culm12 (bc12) encoding kinesin-4 protein</td><td>O. sativa L.</td><td>Increased lignin with no change in cellulose</td><td>Zhang et al. (2010)</td></tr><tr><td>Altered lignin structure and composition</td><td>Downregulation of caffeic acid O-methyltransferase (COMT)</td><td>L. perenne L.; Z. mays L.; Hordeum vulgare L.</td><td>Generally: Lignin syringyl-to-guaicyl ratio reduced; 5-hydroxyguaiacyl lignin units increased; Improvement in digestibility and/or saccharification</td><td>Pichon et al. (2006), Tu et al. (2010) and Daly et al. (2019)</td></tr><tr><td></td><td>Downregulation of ferulate-5-hydroxylase (F5H)</td><td>O. sativa L.; H. vulgare L.</td><td>Lignin syringyl-to-guaicyl ratio reduced by up to 85%</td><td>Takeda et al. (2017) and Shafiei et al. (2023)</td></tr><tr><td></td><td>Downregulation or mutation of cinnamyl alcohol dehydrogenase (CAD)</td><td>Many grasses for example, Z. mays L.; Festuca arundinacea Schreb.</td><td>Generally reduction in syringyl lignin units and syringyl-to-guaicyl ratio; New lignin monomers detected (sinapaldehyde, coniferaldehyde); Digestibility, saccharification, or increased ethanol yield reported in some studies</td><td>Halpin et al. (1998), Chen et al. (2003), Fornalé et al. (2012) and Barriere et al. (2013)</td></tr><tr><td>Cell wall crosslinks</td><td>Downregulation of feruloyl transferases (FAEs)</td><td>O. sativa L.</td><td>19% reduction in ferulic acid in cell walls</td><td>Piston et al. (2010)</td></tr><tr><td></td><td>Overexpression of feruloyl-CoA monolignol transferase (FMT)</td><td>O. sativa L.</td><td>Increased lignin-associated ferulate esters</td><td>Karlen et al. (2016)</td></tr><tr><td></td><td>Mutation in BAHD p-coumaroyl arabinoxylan transferase (HvAt10)</td><td>H. vulgare L.</td><td>Reduction in esterified p-coumaric acid and increase in ferulic acid in grain cell walls</td><td>Houston et al. (2023)</td></tr><tr><td>Straw with increased protein content</td><td>Exploit natural variation for the trait (e.g., 200 spring advanced breeding lines from ICARDA—Morocco)</td><td>H. vulgare L.</td><td>Crude protein content varies between 3 and 6.5% of straw weight</td><td>Sanchez-Garcia et al., unpublished</td></tr></tbody></table>

(continued)
<table><caption>**Table 2.** (continued)</caption><thead><tr><th colspan="5">Optimization of barley straw</th></tr><tr><th>Trait</th><th>Target gene(s)/germoplasm(s)</th><th>Tested Species</th><th>Level of accumulation</th><th>References</th></tr></thead><tbody><tr><td>Nitrogen content</td><td>Mutation in abnormal cytokinin response 1 repressor 1 (HvARE1)</td><td>H. vulgare L.</td><td>Four times more nitrogen content in shoots of are1 mutant grown under low nitrogen conditions</td><td>Karunaratne et al. (2022)</td></tr><tr><td>Nitrogen use efficiency</td><td>Mutation in Bric-a-Brac/Tramtrack/Broad gene (OsBT2)</td><td>O. sativa</td><td>Mutations of this gene in rice increased NUE by 20% compared with wild type under low N conditions</td><td>Araus et al. (2016)</td></tr><tr><td>Nitrogen translocation</td><td>Mutation in Lysine-Histidine-type Transporter 1 (OsLHT1)</td><td>O. sativa</td><td>At maturity, only 40% of total N was translocated to reproductive tissues in mutants (62% in wild type plants)</td><td>Guo et al. (2020)</td></tr></tbody></table>

## PERSPECTIVES ON INNOVATIVE TECHNOLOGIES AND ENVIRONMENTAL AND ECONOMIC BENEFITS

### Impacts from increased barley productivity

By increasing the productivity of barley (both grain and biomass) up to 5–10%, BEST-CROP could contribute to almost 8 Mt more grain and 5 Mt more removable straw per year if this was replicated globally, potentially contributing to food security. Improving the photosynthetic efficiency of barley will also offer major advantages for the environment, given that around 4% of arable land is used for barley cultivation, globally. Agriculture has, indeed, an active role in the emission of greenhouse gasses to the atmosphere (Kabange et al., 2023). According to the Green House Gases (GHG) inventories of the US Environmental Protection Agency (EPA), GHG emissions in agriculture accounted for almost 22% of total global GHG emissions, a non-negligible amount (EPA, 2024). The new barley lines developed by BEST-CROP are expected to convert more CO₂ to biomass and soil carbon, which will directly contribute to decreasing agricultural contribution to greenhouse gas emissions. The impact of agriculture on climate change and GHG emissions is largely driven by the production of ammonia for agriculture, which currently takes 2–3% of the total world energy (Pfromm, 2017), thus saving nitrogen use is key to reducing energy consumption and GHG release. Further, nitrogen runoff from agricultural land is a major pollutant. Therefore, reducing the use of nitrogen fertilizer will help to minimize this problem. With respect to this issue, the adoption of pale green barley lines in BEST-CROP is of critical importance since reduced leaf chlorophyll content has been associated with a 9% decrease in nitrogen inputs (Walker et al., 2018). Moreover, photorespiration lowers crop nitrogen use efficiency because ammonia (NH₃) is released by the glycine decarboxylase reaction in mitochondria during photorespiration. Reducing the rate of photorespiration and/or restricting photorespiratory nitrogen release by introducing photorespiratory bypasses will contribute to the unproductive loss of nitrogen from plants. In this context, it is interesting to note that approximately 90% of the C3-photosynthesis leaf NH3 assimilation machinery is busy with re-assimilation of photorespiratory ammonia, while only approximately 10% is involved in net N-assimilation (Keys, 2006). Pale green leaves in crops will also have a major role in climate change mitigation since they will increase the surface albedo. Modeling experiments have shown that higher cropland albedo may effectively mitigate the magnitude of future heatwaves and global warming in general (Kala et al., 2022) by lowering near-surface air temperatures (Seneviratne et al., 2018) and transpiration water loss, leading to a 25% reduction of leaf-level water use (Głowacka et al., 2018). Furthermore, BEST-CROP aims to increase stomatal O₃ assimilation without yield penalty. This will
mitigate O₃ spikes and improve air quality. Noteworthy, some of the biotechnological strategies adopted in BEST-CROP have been already shown to work also in other crops including, among others, wheat, rice, tomato (Solanum lycopersicum L.), soybean, and maize, thus cultivation of crops with enhanced photosynthesis performance and ozone uptake is expected to provide a major contribution to the sustainability of agriculture and mitigation of climate change.

## Impacts from barley straw-based construction panels and composites

Straw with various lignin content and properties can be used as raw material for different manufacturing pipelines. This includes mycelium-based and straw-based panels and composites for the construction and composite sectors. In the past 20 years, the production volume of wood-based panels for building and construction has almost doubled from approximately 180 million m³ in 2000 to over 368 million m³ in 2020 (FAOSTAT-Forestry Database, https://www.xresearch.biz/shop/biocomposites-market). Also, the global biocomposite market is expected to grow from USD 24 726.7 million in 2022 to USD 74 593.5 million in 2030 (XResearch Company, 2023). The market attractiveness for these materials and the actual and projected increase in production volumes offer excellent prospects for the use of other biomasses as alternatives to wood and synthetic materials. In particular, the valorization of residual straws will have a considerable impact on the circular bioeconomy and climate change mitigation. Various lignocellulosic sources and agricultural production processes, including traditional plant fibers (e.g., flax, hemp, jute, sisal kenaf) and agro and forestry residues (e.g., cereal straws, corn stover, grasses such as miscanthus; Mahieu et al., 2019; Mohanty et al., 2018; Neitzel et al., 2023; Pędzik et al., 2021), have shown clear potential as alternative raw materials for the manufacture of particle boards and composite materials with excellent mechanical, insulation and fire properties (Andrew et al., 2024; Bekhta et al., 2023; Bourmaud et al., 2018; Lee et al., 2022; Rosa Latapie et al., 2023). Furthermore, annual production volumes of residual cereal straws are consistent with the huge demand for materials in these sectors. The sustainable solutions exploited in BEST-CROP will enter the economically relevant building insulation material market that has been valued globally at USD 27.84 billion in 2020, and it is projected to register a CAGR (Compound Annual Growth Rate) of 4.35% during the period 2021–2026.

## Impacts from barley straw-derived biolubricants and proteins

By gaining oils from BSFL fed with protein-enriched barley straw, BEST-CROP will deliver sustainable solutions for the oleochemical industry as well as proteins for feed. This

strategic chemical sector is facing significant difficulties in raw material supply due to competition from biodiesel/biofuels and deforestation regulations, and sustainability rules, leading to uncertainties over future availability of palm and soybean oils. Oil from BSFL will offer sustainable and local solutions to substitute plant and fossil-fuel derived oils, and possibly increase guarantees on supply security. Moreover, several other substances can be obtained from BSLF oil finding application in a variety of industrial sectors, like building industry, cosmetics, pharmaceuticals and animal feeding.

In addition to being biodegradable and non-toxic, biolubricants have been shown to exhibit superior properties over conventional lubricants (Syahir et al., 2017). In this context, the demand for biolubricants, such as lubrorefrigerant oils and metalworking lubricants, is expected to increase soon (for applications such as hydraulic oils, engine oils, gear oils, cutting fluids, electrical appliances, and turbomachinery) as an alternative to conventional lubricants, especially in uses entailing leakages (Sarma & Vinu, 2022). The global biolubricant market was estimated at 1.9 billion USD in the year 2020 and is projected to reach 2.5 billion USD by 2026, growing at a CAGR of 5.2% over the analysis period (Grand View Research, 2023). Moreover, proteins obtained from BSFL-based conversion of barley straw will help to cover the existing shortage in local protein production. Being sustainable and free from indirect land-usage change (ILUC) risk, these proteins could be used to feed fish, non-ruminant animals and, in the case of hydrolysates, free amino acids can also be used for ruminants.

## CONCLUDING REMARKS

Current environmental and societal issues, together with existing and future policies, are driving industries to develop eco-efficient, bio-based compounds, and materials in various sectors, such as chemical industry, building & construction, transportation, and sports & leisure. Besides the market size and growth expectations, the current availability of sustainable raw materials offers great opportunities. To this end, a variety of lignocellulosic sources and agricultural production processes are being considered, including traditional plant fibers and agricultural and forestry residues. The deployment of crop-straw and plant-based products will promote the creation of new industrial facilities, offering new growth opportunities, particularly for farm-based rural areas, where the diversification of economic activities and new industrial outlets are crucial for generating added value in agriculture-related fields, while contributing to the decarbonization of corresponding industrial sectors. The multipurpose barley cultivars delivered by BEST-CROP will support straw-based applications and innovative technologies that can be arranged, even on a small scale, in any
rural area. This is a huge advantage for the supply chain, allowing the entire production process to be covered in a single compact plant—from raw material to final product. Finally, BEST-CROP knowledge and innovations in terms of genetic improvement of photosynthesis and straw quality, together with straw transformation technologies could also be transferred to other major crops, first of all wheat, making the optimization of canopy photosynthesis and valorization of straw in different industrial sectors a powerful lever for the circular bioeconomy and climate change mitigation.

## CONFLICT OF INTEREST STATEMENT

The authors have not declared a conflict of interest.

## ACKNOWLEDGEMENT

Open access publishing facilitated by Universita degli Studi di Milano, as part of the Wiley - CRUI-CARE agreement.

## DATA AVAILABILITY STATEMENT

The data that support the findings of this study are available on request from the corresponding author. The data are not publicly available due to privacy or ethical restrictions.

## REFERENCES

* Ahmad, H., Chhipi-Shrestha, G., Hewage, K. & Sadiq, R. (2022) A comprehensive review on construction applications and life cycle sustainability of natural fiber biocomposites. *Sustainability, 14*, 15905.

* Ahmar, S., Usman, B., Hensel, G., Jung, K.-H. & Gruszka, D. (2024) CRISPR enables sustainable cereal production for a greener future. *Trends in Plant Science, 29*, 179-195. Available from: https://doi.org/10.1016/j.tplants.2023.10.016

* Alaneme, K.K., Anaele, J.U., Oke, T.M., Kareem, S.A., Adediran, M., Ajibuwa, O.A. et al. (2023) Mycelium based composites: a review of their bio-fabrication procedures, material properties and potential for green building and construction applications. *Alexandria Engineering Journal, 83*, 234-250. Available from: https://doi.org/10.1016/j.aej.2023.10.012

* Alboresi, A., Storti, M. & Morosinotto, T. (2019) Balancing protection and efficiency in the regulation of photosynthetic electron transport across plant evolution. *The New Phytologist, 221*, 105-109. Available from: https://doi.org/10.1111/nph.15372

* Amziane, S. & Collet, F. (Eds.). (2017) *Bio-aggregates based building materials*. State-of-the-Art Report of the RILEM Technical Committee 236-288. Dordrecht: Springer. Available from: https://doi.org/10.1007/978-94-024-1031-0

* Andrew, J.J., Sain, M., Ramakrishna, S., Jawaid, M. & Dhakal, H.N. (2024) Environmentally friendly fire retardant natural fibre composites: a review. *International Materials Review, 69*(5-6), 267-308. Available from: https://doi.org/10.1177/09506608241266302

* Araus, V., Vidal, E.A., Puelma, T., Alamos, S., Mieulet, D., Guiderdoni, E. et al. (2016) Members of BTB gene family of scaffold proteins suppress nitrate uptake and nitrogen use efficiency. *Plant Physiology, 171*, 1523-1532.

* Assefa, Y., Carter, P., Hinds, M., Bhalla, G., Schon, R., Jeschke, M. et al. (2018) Analysis of long term study indicates both agronomic optimal plant density and increase maize yield per plant contributed to yield gain. *Scientific Reports, 8*, 1-11. Available from: https://doi.org/10.1038/s41598-018-23362-x

* Attias, N., Danai, O., Abitbol, T., Tarazi, E., Ezov, N., Pereman, I. et al. (2020) Mycelium bio-composites in industrial design and architecture: comparative review and experimental analysis. *Journal of Cleaner Production, 246*, 119037. Available from: https://doi.org/10.1016/j.jclepro.2019.119037

* Barmeier, G., Hu, Y. & Schmidhalter, U. (2021) Partitioning and translocation of dry matter and nitrogen during grain filling in spring barley varieties and their roles in determining malting quality. *Frontiers in Plant Science, 12*, 1-15. Available from: https://doi.org/10.3389/fpls.2021.722871

* Barriere, Y., Chavignneau, H., Delaunay, S. et al. (2013) Different mutations in the ZmCAD2 gene underlie the maize brown-midrib1 (bm1) phenotype with similar effects on lignin characteristics and have potential interest for bioenergy production. *Maydica, 58*, 6-20.

* Bekhta, P., Kozak, R., Gryc, V., Pipiska, T., Sedliacik, J., Reh, R. et al. (2023) Properties of lightweight particleboard made with sunflower stalk particles in the core layer. *Industrial Crops and Products, 205*, 117444. Available from: https://doi.org/10.1016/j.indcrop.2023.117444.%0A

* Bellucci, A., Tondelli, A., Fangel, J.U., Torp, A.M., Xu, X., Willats, W.G.T. et al. (2017) Genome-wide association mapping in winter barley for grain yield and culm cell wall polymer content using the high-throughput CoMPP technique. *PLoS One, 12*, e0173313. Available from: https://doi.org/10.1371/journal.pone.0173313

* Blackman, V.H. (1919) The compound interest law and plant growth. *Annals of Botany, os-33*, 353-360. Available from: https://doi.org/10.1093/oxfordjournals.aob.a089727

* Boix, E., Gineau, E., Narciso, J.O., Höfte, H., Mouille, G. & Navard, P. (2020) Influence of chemical treatments of miscanthus stem fragments on polysaccharide release in the presence of cement and on the mechanical properties of bio-based concrete materials. *Cement and Concrete Composites, 105*, 1-8. Available from: https://doi.org/10.1016/j.cemconcomp.2019.103429.%0A

* Bourmaud, A., Beaugrand, J., Shah, D.U., Placet, V. & Bailey, C. (2018) Towards the design of high-performance plant fibre composites. *Progress in Materials Science, 97*, 347-408. Available from: https://doi.org/10.1016/j.pmatsci.2018.05.005

* Brancourt-Hulmel, M., Raverdy, R., Girones, J., Arnoult, S., Mignot, E., Griyeaux, Y. et al. (2021) Variability of stem solidness among miscanthus genotypes and its role on mechanical properties of polypropylene composites. *GCB Bioenergy, 13*, 1576-1585. Available from: https://doi.org/10.1111/gcbb.12818

* Brant, E.J., Eid, A., Kannan, B., Baloglu, M.C. & Altpeter, F. (2024) The extent of multiallelic, co-editing of LIGULELESS1 in highly polyploid sugarcane tunes leaf inclination angle and enables selection of the ideotype for biomass yield. *Plant Biotechnology Journal, 22*, 2660-2671. Available from: https://doi.org/10.1111/pbi.14380

* Brosché, M., Merilo, E., Mayer, F., Pechter, P., Puzorjova, I., Brader, G. et al. (2010) Natural variation in ozone sensitivity among *Arabidopsis thaliana* accessions and its relation to stomatal conductance. *Plant, Cell & Environment, 33*, 914-925. Available from: https://doi.org/10.1111/j.1365-3040.2010.02116.x

* Butu, A., Rodino, S., Miu, B. & Butu, M. (2020) Mycelium-based materials for the ecodesign of bioeconomy. *Digest Journal of Nanomaterials and Biostructures, 15*, 1129-1140. Available from: https://doi.org/10.15251/DJNB.2020.154.1129

* Caine, R.S., Harrison, E.L., Sloan, J., Flis, P.M., Fischer, S., Khan, M.S. et al. (2023) The influences of stomatal size and density on rice abiotic stress resilience. *The New Phytologist, 237*, 2180-2195. Available from: https://doi.org/10.1111/nph.18704

* Caldwell, D.G., McCallum, N., Shaw, P., Muehlbauer, G.J., Marshall, D.F. & Waugh, R. (2004) A structured mutant population for forward and reverse genetics in Barley (*Hordeum vulgare* L.). *The Plant Journal, 40*, 143-150. Available from: https://doi.org/10.1111/j.1365-313X.2004.02190.x

* Chen, L., Auh, C., Dowling, P., Bell, J., Chen, F., Hopkins, A. et al. (2003) Improved forage digestibility of tall fescue (*Festuca arundinacea*) by transgenic down-regulation of cinnamyl alcohol dehydrogenase. *Plant Biotechnology Journal, 1*, 437-449. Available from: https://doi.org/10.1046/j.1467-7652.2003.00040.x

* Chia, S.Y., Tanga, C.M., van Loon, J.J. & Dicke, M. (2019) Insects for sustainable animal feed: inclusive business models involving smallholder farmers. *Current Opinion in Environment Sustainability, 41*, 23-30. Available from: https://doi.org/10.1016/j cosust.2019.09.003

* Chupin, L., Arnoult, S., Brancourt-Hulmel, M., Lapierre, C., Gineau, E. & Navard, P. (2017) Polyethylene composites made from below-ground miscanthus biomass. *Industrial Crops and Products, 109*, 523-528. Available from: https://doi.org/10.1016/j.indcrop.2017.09.007
Chupin, L., Soccalingame, L., de Ridder, D., Gineau, E., Mouille, G., Arnout, S. et al. (2020) Thermal and dynamic mechanical characterization of miscanthus stem fragments: effects of genotypes, positions along the stem and their relation with biochemical and structural characteristics. *Industrial Crops and Products*, **156**, 112863. Available from: https://doi.org/10.1016/j.indcrop.2020.112863

Croce, R., Carmo-Silva, E., Cho, Y.B., Ermakova, M., Harbinson, J., Lawson, T. et al. (2024) Perspectives on improving photosynthesis to increase crop yield. *Plant Cell*, **36**, 3944–3973. Available from: https://doi.org/10.1093/plcell/koae132

Cutolo, E.A., Guardini, Z., Dall'Osto, L. & Bassi, R. (2023) A paler shade of green: engineering cellular chlorophyll content to enhance photosynthesis in crowded environments. *The New Phytologist*, **239**, 1567–1583. Available from: https://doi.org/10.1111/nph.19064

Dabbou, S., Gai, F., Biasato, I., Capucchio, M.T., Biasibetti, E., Dezzutto, D. et al. (2018) Black soldier fly defatted meal as a dietary protein source for broiler chickens: effects on growth performance, blood traits, gut morphology and histological features. *Journal of Animal Science and Biotechnology*, **9**, 49. Available from: https://doi.org/10.1186/s40104-018-0266-9

Daly, P., McClellan, C., Maluk, M., Oakey, H., Lapierre, C., Waugh, R. et al. (2019) RNAi-suppression of barley caffeic acid O-methyltransferase modifies lignin despite redundancy in the gene family. *Plant Biotechnology Journal*, **17**, 594–607. Available from: https://doi.org/10.1111/pbi.13001

de Souza, A.P., Burgess, S.J., Doran, L., Manukyan, L., Hansen, J., Maryn, N. et al. (2022) Soybean photosynthesis and crop yield is improved by accelerating recovery from photoprotection. *Science*, **377**, 851–854. Available from: https://doi.org/10.1126/science.adc9831

Di Giuseppe, E., Girones, J., Vo, L.T.T., Gineau, E., Lapierre, C., Brancourt-Hulme, M. et al. (2021) Polysaccharides and phenolics of miscanthus belowground cell walls and their influence on polyethylene composites. *Carbohydrate Polymers*, **251**, 1–8. Available from: https://doi.org/10.1016/j.carbpol.2020.117086

Diener, A. & Mudu, P. (2021) How can vegetation protect us from air pollution? A critical review on green spaces' mitigation abilities for air-borne particles from a public health perspective—with implications for urban planning. *Science of the Total Environment*, **796**, 148605. Available from: https://doi.org/10.1016/j.scitotenv.2021.148605

Dorez, G., Ferry, L., Sonnier, R., Taguet, A. & Lopez-Cuesta, J.-M. (2014) Effect of cellulose, hemicellulose and lignin contents on pyrolysis and combustion of natural fibers. *Journal of Analytical and Applied Pyrolysis*, **107**, 323–331. Available from: https://doi.org/10.1016/j.jaap.2014.03.017

Duvick, D.N. (2005) Genetic progress in yield of United States maize (Zea mays L.). *Maydica*, **50**, 193–202.

Eloy, N.B., Voarend, W., Lan, W., Saleme, M.L.S., Cesarino, I., Vanholme, R. et al. (2017) Silencing CHALCONE SYNTHASE in maize impedes the incorporation of tricin into lignin and increases lignin content. *Plant Physiology*, **173**, 998–1016. Available from: https://doi.org/10.1104/pp.16.01108

EPA. (2024) Inventory of U.S. greenhouse gas emissions and sinks: 1990-2022. U.S. Environmental Protection Agency, EPA 430R-24004. Available at: www.epa.gov/ghgemissions/inventory-us-greenhouse-gas-emissions-and-sinks

Flood, P.J., Harbinson, J. & Aarts, M.G.M. (2011) Natural genetic variation in plant photosynthesis. *Trends in Plant Science*, **16**, 327–335. Available from: https://doi.org/10.1016/j.tplants.2011.02.005

Fontaine, D., Eriksen, J. & Sørensen, P. (2020) Cover crop and cereal straw management influence the residual nitrogen effect. *European Journal of Agronomy*, **118**, 126100. Available from: https://doi.org/10.1016/j.eja.2020.126100

Fornalé, S., Capellades, M., Encina, A., Wang, K., Irar, S., Lapierre, C. et al. (2012) Altered lignin biosynthesis improves cellulosic bioethanol production in transgenic maize plants down-regulated for cinnamyl alcohol dehydrogenase. *Molecular Plant*, **5**, 817–830. Available from: https://doi.org/10.1093/mp/ssr097

Fuso, A., Barbi, S., Macavei, L.I., Luparelli, A.V., Maistrello, L., Montorsi, M. et al. (2021) Effect of the rearing substrate on total protein and amino acid composition in black soldier fly. *Food*, **10**, 1773.

Garcia-Molina, A. & Leister, D. (2020) Accelerated relaxation of photoprotection impairs biomass accumulation in Arabidopsis. *Nature Plants*, **6**, 9–12. Available from: https://doi.org/10.1038/s41477-019-0572-z

Głowacka, K., Kromdijk, J., Kucera, K., Xie, J., Cavanagh, A.P., Leonelli, L. et al. (2018) Photosystem II subunit S overexpression increases the efficiency of water use in a field-grown crop. *Nature Communications*, **9**, 868.

Grand View Research. (2023) Biolubricants market size, share, & trends analysis report by source (vegetable oil, animal oil), by application (transportation, industrial), by end use (industrial, commercial transportation), by region, and segment forecasts, 2023–2030. Grand View Research.

Grove, A.V., Hepton, J. & Hunt, C.W. (2003) Chemical composition and ruminal fermentability of barley grain, hulls, and straw as affected by planting date, irrigation level, and variety. *The Professional Animal Scientists*, **19**, 273–280. Available from: https://doi.org/10.15232/S1080-7446(15)31422-4

Gui, J., Shen, J. & Li, L. (2011) Functional characterization of evolutionarily divergent 4-coumarate:coenzyme a ligases in rice. *Plant Physiology*, **157**, 574–586. Available from: https://doi.org/10.1104/pp.111.178301

Guo, N., Gu, M., Hu, J., Qu, H. & Xu, G. (2020) Rice OsLHT1 functions in leaf-to-panicle nitrogen allocation for grain yield and quality. *Frontiers in Plant Science*, **11**, 1150. Available from: https://doi.org/10.3389/fpls.2020.01150

Halpin, C. (2019) Lignin engineering to improve saccharification and digestibility in grasses. *Current Opinion in Biotechnology*, **66**, 223–229. Available from: https://doi.org/10.1016/j.copbio.2019.02.013

Halpin, C., Holt, K., Chojecki, J., Oliver, D., Chabbert, B., Monties, B. et al. (1998) Brown-midrib maize (bm1)—a mutation affecting the cinnamyl alcohol dehydrogenase gene. *The Plant Journal*, **14**, 545–553. Available from: https://doi.org/10.1046/j.1365-313X.1998.00153.x

Haneef, M., Ceseracciu, L., Canale, C., Bayer, I.S., Heredia-Guerrero, J.A. & Athanassiou, A. (2017) Advanced materials from fungal mycelium: fabrication and tuning of physical properties. *Scientific Reports*, **7**, 41292. Available from: https://doi.org/10.1038/srep41292

Hansson, M., Youssef, H.M., Zakhrabekova, S., Stuart, D., Svensson, J.T., Dockter, C. et al. (2024) A guide to barley mutants. *Hereditas*, **161**, 1–55. Available from: https://doi.org/10.1186/s41065-023-00304-w

Haughey, E., Neogi, S., Portugal-Pereira, J., van Diemen, R. & Slade, R.B. (2023) Sustainable intensification and carbon sequestration research in agricultural systems: a systematic review. *Environmental Science & Policy*, **143**, 14–23. Available from: https://doi.org/10.1016/j.envsci.2023.02.018

Hörak, H., Kollist, H. & Merilo, E. (2017) Fern stomatal responses to ABA and CO₂ depend on species and growth conditions. *Plant Physiology*, **174**, 672–679. Available from: https://doi.org/10.1104/pp.17.00120

Hörak, H., Sierla, M., Toldsepp, K., Wang, C., Wang, Y.S., Nuhkat, M. et al. (2016) A dominant mutation in the h1 kinase uncovers roles of MAP kinases and GHR1 in CO₂-induced stomatal closure. *Plant Cell*, **28**, 2493–2509. Available from: https://doi.org/10.1105/tpc.16.00131

Houston, K., Learmonth, A., Hassan, A.S., Lahnstein, J., Looseley, M., Little, A. et al. (2023) Natural variation in HvAT10 underlies grain cell wall-esterified phenolic acid content in cultivated barley. *Frontiers in Plant Science*, **14**, 1095862. Available from: https://doi.org/10.3389/fpls.2023.1095862

Hughes, J., Hepworth, C., Dutton, C., Dunn, J.A., Hunt, L., Stephens, J. et al. (2017) Reducing stomatal density in barley improves drought tolerance without impacting on yield. *Plant Physiology*, **174**, 776–787. Available from: https://doi.org/10.1104/pp.16.01844

Johnson, J.M.F., Allmaras, R.R. & Reicosky, D.C. (2006) Estimating source carbon from crop residues, roots and rhizodeposits using the national grain-yield database. *Agronomy Journal*, **98**, 622–636. Available from: https://doi.org/10.2134/agronj2005.0179

Kabange, N.R., Kwon, Y., Lee, S.M., Kang, J.W., Cha, J.K., Park, H. et al. (2023) Mitigating greenhouse gas emissions from crop production and management practices, and livestock: a review. *Sustainability*, **15**, 15889. Available from: https://doi.org/10.3390/su152215889

Kala, J., Hirsch, A.L., Ziehn, T., Perkins-Kirkpatrick, S.E., de Kauwe, M.G. & Pitman, A. (2022) Assessing the potential for crop albedo enhancement in reducing heatwave frequency, duration, and intensity under future climate change. *Weather and Climate Extremes*, **35**, 100415. Available from: https://doi.org/10.1016/j.wace.2022.100415

Karlen, S.D., Zhang, C., Peck, M.L., Smith, R.A., Padmakshan, D., Helmich, K.E. et al. (2016) Monolignol ferulate conjugates are naturally incorporated into plant lignins. *Science Advances*, **2**, e1600393. Available from: https://doi.org/10.1126/sciadv.1600393
Karunarathne, S.D., Han, Y., Zhang, X.Q. & Li, C. (2022) CRISPR/Cas9 gene editing and natural variation analysis demonstrate the potential for HvARE1 in improvement of nitrogen use efficiency in barley. *Journal of Integrative Plant Biology, 64*, 756-770. Available from: https://doi.org/10.1111/jipb.13214

Katsarova, I. (2024) *Plants produced by certain new genomic techniques*. Legis. train 08.2024, Eur. Parliam. Available at: https://www.europarl.europa.eu/legislative-train/theme-a-european-green-deal/file-plants-produced-by-certain-new-genomic-techniques

Kebeish, R., Niessen, M., Thiruveedhi, K., Bari, R., Hirsch, H.J., Rosenkranz, R. et al. (2007) Chloroplastic photorespiratory bypass increases photosynthesis and biomass production in *Arabidopsis thaliana*. *Nature Biotechnology*, 25, 593-599. Available from: https://doi.org/10.1038/nbt1299

Keno, M.T., Tolemariam, T., Demeke, S., Wamatu, J., Alkhtib, A. & Janssens, G.P.J. (2021) Effect of barley variety on feed intake, digestibility, body weight gain and carcass characteristics in fattening lambs. *Animals*, 11, 1773. Available from: https://doi.org/10.3390/ani11016773

Keys, A.J. (2006) The re-assimilation of ammonia produced by photosynthesis and the nitrogen economy of C₃ higher plants. *Photosynthesis Research*, 87, 165-175. Available from: https://doi.org/10.1007/s11120-005-9024-x

Khaipho-Burch, M., Cooper, M., Crossa, J., de Leon, N., Holland, J., Lewis, R. et al. (2023) Scale up trials to validate modified crops' benefits. *Nature*, 621, 470-473. Available from: https://doi.org/10.1038/d41586-023-02895-w

Kirst, H., Shen, Y., Vamvaka, E., Betterle, N., Xu, D., Warek, U. et al. (2018) Downregulation of the CpSRP43 gene expression confers a truncated light-harvesting antenna (TLA) and enhances biomass and leaf-to-stem ratio in *Nicotiana tabacum* canopies. *Planta*, 248, 139-154. Available from: https://doi.org/10.1007/s00425-018-2889-7

Klimyuk, V.I., Persello-Cartieaux, F., Havaux, M., Contard-David, P., Schueinemann, D., Meiherhoff, K. et al. (1999) A chromodomain protein encoded by the Arabidopsis CAO gene is a plant-specific component of the chloroplast signal recognition particle pathway that is involved in LHCP targeting. *Plant Cell*, 11, 87-99. Available from: https://doi.org/10.1105/tpc.11.1.87

Kogata, N., Nishio, K., Hirohashi, T., Kikuchi, S. & Nakai, M. (1999) Involve-ment of a chloroplast homologue of the signal recognition particle recep-tor protein, FtsY, in protein targeting to thylakoids. *FEBS Letters*, 329, 329-333. Available from: https://doi.org/10.1016/S0014-5793(99)00305-1

Kollist, H., Moldau, H., Mortensen, L., Rasmussen, S.K. & Jrgensen, L.B. (2000) Ozone flux to plasmalemma in barley and wheat is controlled by stomata rather than by direct reaction of ozone with Cell Wall ascorbate. *Journal of Plant Physiology*, 156, 645-651. Available from: https://doi.org/10.1016/S0176-1617(00)80226-6

Kollist, T., Moldau, H., Rasulov, B., Oja, V., Rämä, H., Hüve, K. et al. (2007) A novel device detects a rapid ozone-induced transient stomatal closure in intact Arabidopsis and its absence in abi2 mutant. *Physiologia Plantarum*, 129, 796-803. Available from: https://doi.org/10.1111/j.1399-3054.2006.00851.x

Kromdijk, J., Glowacka, K., Leonelli, L., Gabilly, S.T., Iwai, M., Niyogi, K.K. et al. (2016) Improving photosynthesis and crop productivity by accelerating recovery from photoprotection. *Science*, 354, 857-861. Available from: https://doi.org/10.1126/science.aai887

Lee, S.H., Lum, W.C., Boon, J.G., Kristak, L., Antov, P., Pedzik, M. et al. (2022) Particleboard from agricultural biomass and recycled wood waste: a review. *Journal of Materials Research and Technology*, 20, 4630-4658. Available from: https://doi.org/10.1016/j.jmrt.2022.08.166

Lefevre, A., Le, A., Bourmaud, A., Kervoelen, A., Morvan, C. & Bailey, C. (2015) Analysis of the role of the main constitutive polysaccharides in the flax fibre mechanical behaviour. *Industrial Crops and Products*, 76, 1039-1048. Available from: https://doi.org/10.1016/j.indcrop.2015.07.062

Lehretz, G.G., Schneider, A., Leister, D. & Sonnewald, U. (2022) High non-photochemical quenching of VPZ transgenic potato plants limits CO₂ assimilation under high light conditions and reduces tuber yield under fluctuating light. *Journal of Integrative Plant Biology*, 64, 1821-1832. Available from: https://doi.org/10.1111/jipb.13320

Li, Y., Qian, Q., Zhou, Y., Yan, M., Sun, L., Zhang, M. et al. (2003) BRITTLE CULM1, which encodes a COBRA-like protein, affects the mechanical properties of rice plants. *Plant Cell*, 15, 2020-2031.

Li, Y., Zhu, N. & Chen, J. (2023) Straw characteristics and mechanical straw building materials: a review. *Journal of Materials Science*, 58, 2361-2380. Available from: https://doi.org/10.1007/s10853-023-08153-8

Lopes, T., Labeni, S., Sonnier, R., Ferry, L., Regazzi, A., Uwizeyimana, P. et al. (2024) Ignition of biobased concretes. *Construction and Building Materials*, 440, 137423. Available from: https://doi.org/10.1016/j.conbuildmat.2024.137423

López-Calcagno, P.E., Brown, K.L., Simkin, A.J., Fisk, S.J., Vialet-Chabrand, S., Lawson, T. et al. (2020) Stimulating photosynthetic processes increases productivity and water-use efficiency in the field. *Nature Plants*, 6, 1054-1063. Available from: https://doi.org/10.1038/s41777-020-0740-1

Lu, S., Taethaisong, N., Meethip, W., Surakhunthod, J., Sinpru, B., Sroichak, T. et al. (2022) Nutritional composition of black soldier fly larvae (Hermetia illucens L.) and its potential uses as alternative protein sources in animal diets: a review. *Insects*, 13, 831. Available from: https://doi.org/10.3390/insects13090831

Mahieu, A., Alix, S. & Leblanc, N. (2019) Properties of particleboards made of agricultural by-products with a classical binder or self-bound. *Industrial Crops and Products*, 130, 371-379. Available from: https://doi.org/10.1016/j.indcrop.2018.12.094

Mahieu, A., Vivet, A., Poilane, C. & Leblanc, N. (2021) Performance of particleneboards based on annual plant byproducts bound with bio-adhesives. *International Journal of Adhesion and Adhesives*, 107, 102847. Available from: https://doi.org/10.1016/j.ijadhadh.2021.102847

Maier, A., Fahnenstich, H., von Caemmerer, S., Engqvist, M.K.M., Weber, A.P.M., Flügge, U.I. et al. (2012) Transgenic introduction of a glycolate oxidative cycle into *A. thaliana* chloroplasts leads to growth improvement. *Frontiers in Plant Science*, 3, 38. Available from: https://doi.org/10.3389/fpls.2012.00038

Mantilla-Perez, M.B. & Salas Fernandez, M.G. (2017) Differential manipulation of leaf angle throughout the canopy: current status and prospects. *Journal of Experimental Botany*, 68, 5699-5717. Available from: https://doi.org/10.1093/jxb/erx378

McAusland, L., Vialet-Chabrand, S., Davey, P., Baker, N.R., Brendel, O. & Lawson, T. (2016) Effects of kinetics of light-induced stomatal responses on photosynthesis and water-use efficiency. *The New Phytologist*, 211, 1209-1220. Available from: https://doi.org/10.1111/nph.14000

Merlot, S., Leonhardt, N., Fenzi, F., Valon, C., Costa, M., Piette, L. et al. (2007) Constitutive activation of a plasma membrane H⁺-ATPase prevents abscisic acid-mediated stomatal closure. *The EMBO Journal*, 26, 3216-3226. Available from: https://doi.org/10.1038/sj.emboj.7601750

Miller, M.A.E., O'Cualain, R., Selley, J., Knight, D., Karim, M.F., Hubbard, S.J. et al. (2017) Dynamic acclimation to high light in *Arabidopsis thaliana* involves widespread reengineering of the leaf proteome. *Frontiers in Plant Science*, 8, 1239. Available from: https://doi.org/10.3389/fpls.2017.01239

Mohanty, A.K., Vivekanandhan, S., Pin, J.M. & Misra, M. (2018) Composites from renewable and sustainable resources: challenges and innovations. *Science*, 362, 536-542. Available from: https://doi.org/10.1126/science.aat9072

Moore, M., Harrison, M.S., Peterson, E.C. & Henry, R. (2000) Chloroplast Oxa1p homolog Albino3 is required for post-translational integration of the light harvesting chlorophyll-binding protein into thylakoid membranes. *The Journal of Biological Chemistry*, 275, 1529-1532. Available from: https://doi.org/10.1074/jbc.275.3.1529

Morales, L.O., Shapiguzov, A., Safronov, O., Leppälä, J., Vaahtera, L., Yarmolinsky, D. et al. (2021) Ozone responses in Arabidopsis: beyond stomatal conductance. *Plant Physiology*, 186, 180-192. Available from: https://doi.org/10.1093/plphys/kiab097

Nakanishi, E.Y., Cabral, M.R., Fiorelli, J., dos Santos, V., Christoforo, A.L. & Savastano Junior, H. (2018) Study of the production process of 3-layer sugarcane-bamboo-based particleboards. *Construction and Building Materials*, 183, 618-625. Available from: https://doi.org/10.1016/j.conbuildmat.2018.06.202

Neitzel, N., Eder, M., Hosseinpourpia, R., Walther, T. & Adamopoulos, S. (2023) Chemical composition, particle geometry, and micro-mechanical strength of barley husks, oat husks, and wheat bran as alternative raw materials for particleboards. *Materials Today Communications*, 36, 106602. Available from: https://doi.org/10.1016/j.mtcomm.2023.106602

Nogales-Mérida, S., Gobbi, P., Józefiak, D., Mazurkiewicz, J., Dudek, K., Rawski, M. et al. (2019) Insect meals in fish nutrition. *Reviews in Agriculture*, 11, 1080-1103. Available from: https://doi.org/10.1111/raq.12281
Oonincx, D.G.A.B. & de Boer, I.J.M. (2012) Environmental impact of the pro-
duction of mealworms as a protein source for humans—a life cycle
assessment. *PLoS One*, 7, e51145. Available from: https://doi.org/10.
1371/journal.pone.0051145

Ort, D.R., Merchant, S.S., Alric, J., Barkan, A., Blankenship, R.E., Bock, R.
et al. (2015) Redesigning photosynthesis to sustainably meet global food
and bioenergy demand. *Proceedings of the National Academy of Sci-
ences of the United States of America*, **112**, 8529–8536. Available from:
https://doi.org/10.1073/pnas.1424031112

Papanatsiou, M., Petersen, J., Henderson, L., Wang, Y., Christie, J.M. & Blatt, M.R. (2019) Optogenetic manipulation of stomatal kinetics improves carbon assimilation, water use, and growth. *Plant Science*, **363**, 1456–1459.

Pedzik, M., Janiszewska, D. & Rogozinski, T. (2021) Alternative lignocellu-
losic raw materials in particleboard production: a review. *Industrial
Crops and Products*, **174**, 114162. Available from: https://doi.org/10.
1016/j.indcrop.2021.114162

Peñaloza, D. (2017) *The role of biobased building materials in the climate impacts of construction*. Doctoral thesis, KTH Stockholm, Sweden. Available at: https://kth.diva-portal.org/smash/get/diva2:1096048/FULLTEXT01.pdf

Pfromm, P.H. (2017) Towards sustainable agriculture: fossil-free ammonia. *Journal of Renewable and Sustainable Energy*, 9, 034702. Available from: https://doi.org/10.1063/1.4985090

Pichon, M., Deswartes, C., Gerentes, D., Guillaumie, S., Lapierre, C., Top-
pan, A. et al. (2006) Variation in lignin and cell wall digestibility in caffeic
acid O-methyltransferase down-regulated maize half-sib progenies in
field experiments. *Molecular Breeding*, **18**, 253–261. Available from:
https://doi.org/10.1007/s11032-006-9033-2

Pick, T.R., Bräutigam, A., Schulz, M.A., Obata, T., Fernie, A.R. & Weber, A.P.M. (2013) PLGG1, a plasticid glycolate glycerate transporter, is required for photorespiration and defines a unique class of metabolite transporters. *Proceedings of the National Academy of Sciences of the United States of America*, **110**, 3185–3190. Available from: https://doi.org/10.1073/pnas.1215142110

Piston, F., Uauy, C., Fu, L., Langston, J., Labavitch, J. & Dubcovsky, J. (2010) Down-regulation of four putative arabinoxylan feruloyl transferase genes from family PF02458 reduces ester-linked ferulate content in rice cell walls. *Planta*, **231**, 677–691.

Przulj, N. & Momcilovic, V. (2001) Genetic variation for dry matter and nitro-
gen accumulation and translocation in two-rowed spring barley II.
Nitrogen translocation. *European Journal of Agronomy*, **15**, 255–265.
Available from: https://doi.org/10.1016/S1161-0301(01)00108-3

Renna, M., Gasco, L., Livorsi, L., Mele, M., Conte, G., Meneguz, M. et al. (2024) Growth performance, proximate composition and fatty acid profile of black soldier fly larvae reared on two grape pomace varieties. *Animal*, **18**, 101240. Available from: https://doi.org/10.1016/j.animal.2024.101240

Roell, M.S., von Borzykowski, L.S., Westhoff, P., Plett, A., Paczia, N., Claus, P. et al. (2021) A synthetic C4 shuttle via the β-hydroxyaspartate cycle in C3 plants. *Proceedings of the National Academy of Sciences of the United States of America*, **118**, e2022307118. Available from: https://doi.org/10.1073/pnas.2022307118

Rosa Latapie, S., Lagouin, M., Sabathier, V. & Abou-Chakra, A. (2023) From aggregate to particleboard: a new multi-scale model approach to thermal conductivity in bio-based materials. *Journal of Building Engineering*, **78**, 107664. Available from: https://doi.org/10.1016/j.jobe.2023.107664

Rotasperti, L., Tadini, L., Chiara, M., Crosatti, C., Guerra, D., Tagliani, A. et al. (2022) The barley mutant happy under the sun 1 (hus1): an additional contribution to pale green crops. *Environmental and Experimental Botany*, **196**, 104795. Available from: https://doi.org/10.1016/j.envexpbot.2022.104795

Saballos, A., Sattler, S.E., Sanchez, E., Foster, T.P., Xin, Z., Kang, C. et al. (2012) Brown midrib2 (Bmr2) encodes the major 4-coumarate: coenzyme A ligase involved in lignin biosynthesis in sorghum (Sorghum bicolor (L.) Moench). *The Plant Journal*, **70**, 818–830. Available from: https://doi.org/10.1111/j.1365-313X.2012.04933.x

Saballos, A., Vermerris, W., Rivera, L. & Ejeta, G. (2008) Allelic association, chemical characterization and saccharification properties of brown mid-rib mutants of sorghum (Sorghum bicolor (L.) Moench). *Bioenergy Research*, **1**, 193–204. Available from: https://doi.org/10.1007/s12155-008-9025-7

Sakamoto, T., Morinaka, Y., Ohnishi, T., Sunohara, H., Fujioka, S., Ueguchi-
Tanaka, M. et al. (2006) Erect leaves caused by brassinosteroid deficiency
increase biomass production and grain yield in rice. *Nature Biotechnol-
ogy*, **24**, 105–109. Available from: https://doi.org/10.1038/nbt1173

Sarma, R.N. & Vinu, R. (2022) Current status and future prospects of bioluc
bricants: properties and applications. *Lubricants*, **10**, 1–21. Available
from: https://doi.org/10.3390/lubricants10040070

Schreiber, M., Barakate, A., Uzrek, N., Macaulay, M., Sourdille, A., Morris, J. et al. (2019) A highly mutagenised barley (cv. Golden Promise) TIL-LING population coupled with strategies for screening-by-sequencing. *Plant Methods*, **15**, 99. Available from: https://doi.org/10.1186/s13007-019-0486-9

Schuenemann, D., Gupta, S., Persello-Cartieaux, F., Klimyk, V.I., Jones, J.D., Nussaume, L. et al. (1998) A novel signal recognition particle targets light-harvesting proteins to the thylakoid membranes. *Proceedings of the National Academy of Sciences of the United States of America*, **95**, 10312–10316. Available from: https://doi.org/10.1073/pnas.95.17.10312

Seneviratne, S.I., Wartenburger, R., Guillod, B.P., Hirsch, A.L., Vogel, M.M.,
Brovkin, V. et al. (2018) Climate extremes, land-climate feedbacks and
land-use forcing at 1.5°C. *Philosophical Transactions of the Royal Society
A - Mathematical Physical and Engineering Sciences*, **376**, 20160450.
Available from: https://doi.org/10.1098/rsta.2016.0450

Seyedalmoosavi, M.M., Mielenz, M., Veldkamp, T., Das, G. & Metges, C.C. (2022) Growth efficiency, intestinal biology, and nutrient utilization and requirements of black soldier fly (Hermetia illucens) larvae compared to monogastric livestock species: a review. *Journal of Animal Science and Biotechnology*, **13**, 31. Available from: https://doi.org/10.1186/s40104-022-00682-7

Shaaf, S., Bretani, G., Biswas, A., Fontana, I.M. & Rossini, L. (2019) Genetics of barley tiller and leaf development. *Journal of Integrative Plant Biol*, **61**, 226–256. Available from: https://doi.org/10.1111/jipb.12757

Shafiei, R., Hooper, M., McClellan, C., Oakey, H., Stephens, J., Lapierre, C. et al. (2023) Downregulation of barley ferulate 5-hydroxylase dramatically alters straw lignin structure without impact on mechanical properties. *Frontiers in Plant Science*, **13**, 1125003. Available from: https://doi.org/10.3389/fpls.2022.1125003

Shen, B.R., Wang, L.M., Lin, X.L., Yao, Z., Xu, H.W., Zhu, C.H. et al. (2019) Engineering a new chloroplastic photorespiratory bypass to increase photosynthetic efficiency and productivity in rice. *Molecular Plant*, **12**, 199–214. Available from: https://doi.org/10.1016/j.molp.2018.11.013

Sicard, P., Agathokleous, E., Araminiene, V., Carrari, E., Hoshika, Y., de Marco, A. et al. (2018) Should we see urban trees as effective solutions to reduce increasing ozone levels in cities? *Environmental Pollution*, **243**, 163–176. Available from: https://doi.org/10.1016/j.envpol.2018.08.049

Siddique, R., Singh, K., Kunal, P., Singh, M., Corinaldesi, V. & Rajor, A. (2016) Properties of bacterial rice husk ash concrete. *Construction and Building Materials*, **121**, 112–119. Available from: https://doi.org/10.1016/j.conbuildmat.2016.05.146

Sierla, M., Hörak, H., Overmyer, K., Waszczak, C., Yarmolinsky, D., Maierhofer, T. et al. (2018) The receptor-like pseudokinase GHR1 is required for stomatal closure. *Plant Cell*, **30**, 2813–2837. Available from: https://doi.org/10.1105/tpc.18.00441

Simkin, A.J., López-Calcagno, P.E. & Raines, C.A. (2019) Feeding the world: improving photosynthetic efficiency for sustainable crop production. *Journal of Experimental Botany*, **70**, 1119–1140. Available from: https://doi.org/10.1093/jxb/ery445

Simkin, A.J., McAusland, L., Headland, L.R., Lawson, T. & Raines, C.A. (2015) Multigene manipulation of photosynthetic carbon assimilation increases CO₂ fixation and biomass yield in tobacco. *Journal of Experimental Botany*, **66**, 4075–4090. Available from: https://doi.org/10.1093/jxb/erv204

Slattery, R.A. & Ort, D.R. (2019) Carbon assimilation in crops at high tem-
peratures. *Plant, Cell & Environment*, **42**, 2750–2758. Available from:
https://doi.org/10.1111/pce.13572

Slattery, R.A. & Ort, D.R. (2021) Perspectives on improving light distribution and light use efficiency in crop canopies. *Plant Physiology*, **185**, 34–48. Available from: https://doi.org/10.1093/plphys/kiaa006

Smith, E.N., van Aalst, M., Tosens, T., Niinemets, Ü., Stich, B., Morosinotto, T. et al. (2023) Improving photosynthetic efficiency toward food security: strategies, advances, and perspectives. *Molecular Plant*, **16**, 1547–1563. Available from: https://doi.org/10.1016/j.molp.2023.08.017
Smith, R.A., Cass, C.L., Mazaheri, M., Sekhon, R.S., Heckwolf, M., Kaeppler, H. *et al.* (2017) Suppression of CINNAMOYL-CoA REDUCTASE increases the level of monolignol ferulates incorporated into maize lignins. *Bio-technology for Biofuels*, **10**, 109. Available from: https://doi.org/10.1186/s13068-017-0793-1

South, P.F., Cavanagh, A.P., Liu, H.W. & Ort, D.R. (2019) Synthetic glycolate metabolism pathways stimulate crop growth and productivity in the field. *Science*, **363**, eaat9077.

Syahir, A.Z., Zulkifli, N.W.M., Masjuki, H.H., Kalam, M.A., Alabdulkarem, A., Gulzar, M. *et al.* (2017) A review on bio-based lubricants and their applications. *Journal of Cleaner Production*, **168**, 997–1016. Available from: https://doi.org/10.1016/j.jclepro.2017.09.106

Takahashi, Y., Bosmans, K.C., Hsu, P. *et al.* (2023) Stomatal CO₂/bicarbonate sensor consists of two interacting protein kinases, Raf-like HT1 and non-kinase-activity requiring MPK12/MPK4. *Science Advances*, **6161**, 1–13. Available from: https://doi.org/10.1126/sciadv.abq616

Takeda, Y., Koshiba, T., Tobimatsu, Y., Suzuki, S., Murakami, S., Yamamura, M. *et al.* (2017) Regulation of CONIFERALDEHYDE 5-HYDROXYLASE expression to modulate cell wall lignin structure in rice. *Planta*, **246**, 337–349. Available from: https://doi.org/10.1007/s00425-017-2692-x

Tamasloukht, B., Lam, W.Q., Martinez, Y., Tozo, K., Barbier, O., Jourda, C. *et al.* (2011) Characterization of a cinnamoyl-CoA reductase 1 (CCR1) mutant in maize: effects on lignification, fibre development, and global gene expression. *Journal of Experimental Botany*, **62**, 3837–3848. Available from: https://doi.org/10.1093/jxb/err077

Taner, A., Muzaffer, A. & Fazil, D. (2004) *Barley: post-harvest operations*. Rome: Food and Agriculture Organization of the United Nations. Available at: http://www.fao.org/3/a-au997e.pdf

Tian, F., Bradbury, P.J., Brown, P.J., Hung, H., Sun, Q., Flint-Garcia, S. *et al.* (2011) Genome-wide association study of leaf architecture in the maize nested association mapping population. *Nature Genetics*, **43**, 159–162. Available from: https://doi.org/10.1038/ng.746

Tian, J., Wang, C., Chen, F. *et al.* (2024) Maize smart-canopy architecture enhances yield at high densities. *Nature*, **632**, 576–584. Available from: https://doi.org/10.1038/s41586-024-07669-6

Tlaiji, G., Ouldboukhitine, S., Pennec, F. & Biwole, P. (2022) Thermal and mechanical behavior of straw-based construction: a review. *Construction and Building Materials*, **316**, 125915. Available from: https://doi.org/10.1016/j.conbuildmat.2021.125915

Toldsepp, K., Zhang, J., Takahashi, Y., Sindarovska, Y., Hórak, H., Ceciliato, P.H.O. *et al.* (2018) Mitogen-activated protein kinases MPK4 and MPK12 are key components mediating CO₂-induced stomatal movements. *The Plant Journal*, **96**, 1018–1035. Available from: https://doi.org/10.1111/tpj.14087

Tu, C., Schuenemann, D. & Hoffman, N.E. (1999) Chloroplast FtsY, chloroplast signal recognition particle, and GTP are required to reconstitute the soluble phase of light-harvesting chlorophyll protein transport into thylakoid membranes*. *The Journal of Biological Chemistry*, **274**, 27219–27224. Available from: https://doi.org/10.1074/jbc.274.38.27219

Tu, Y., Rochfort, S., Liu, Z., Ran, Y., Griffith, M., Badenhorst, P. *et al.* (2010) Functional analyses of Caffeic acid O-methyltransferase and Cinnamoyl-CoA-reductase genes from perennial ryegrass (*Pennisetum perenne*). *Plant Cell*, **22**, 3357–3373. Available from: https://doi.org/10.1105/tpc.109.072827

Tula, S., Shahinnia, F., Melzer, M., Rutten, T., Gómez, R., Lodeyro, A.F. *et al.* (2020) Providing an additional electron sink by the introduction of cyanobacterial flavodiirons enhances growth of *A. thaliana* under various light intensities. *Frontiers in Plant Science*, **11**, 902. Available from: https://doi.org/10.3389/fpls.2020.00902

Uemura Silva, V., Nascimento, M.F., Resende Oliveira, P., Panzera, T.H., Rezende, M.O., Silva, D.A.L. *et al.* (2021) Circular vs. linear economy of building materials: a case study for particleboards made of recycled wood and biopolymer vs. conventional particleboards. *Construction and Building Materials*, **285**, 122906. Available from: https://doi.org/10.1016/j.conbuildmat.2021.122906

van Camp, W., Willekens, H., Bowler, C., van Montagu, M., Inzé, D., Reupold-Popp, P. *et al.* (1994) Elevated levels of superoxide dismutase protect transgenic plants against ozone damage. *Nature Biotechnology*, **12**, 165–168.

van Dijk, M., Morley, T., Rau, M.L. & Saghai, Y. (2021) A meta-analysis of projected global food demand and population at risk of hunger for the

period 2010–2050. *Nature Food*, **2**, 494–501. Available from: https://doi.org/10.1038/s43016-021-00322-9

Vicino, P., Carrillo, J., Gómez, R., Shahinnia, F., Tula, S., Melzer, M. *et al.* (2021) Expression of flavodiiron proteins Flv2-Flv4 in chloroplasts of arabidopsis and tobacco plants provides multiple stress tolerance. *International Journal of Molecular Sciences*, **22**, 1178. Available from: https://doi.org/10.3390/ijms22031178

Vo, L.T.T., Girones, J., Beloli, C., Chupin, L., di Giuseppe, E., Vidal, A.C. *et al.* (2017) Processing and properties of sorghum stem fragment-polyethylene composites. *Industrial Crops and Products*, **107**, 386–398. Available from: https://doi.org/10.1016/j.indcrop.2017.05.047

Wada, S., Yamamoto, H., Suzuki, Y., Yamori, W., Shikanai, T. & Makino, A. (2018) Flavodiiron protein substitutes for cyclic electron flow without competing CO₂ assimilation in rice. *Plant Physiology*, **176**, 1509–1518. Available from: https://doi.org/10.1104/pp.17.01335

Walker, B.J., Drewry, D.T., Slattery, R.A., VanLoocke, A., Cho, Y.B. & Ort, D.R. (2018) Chlorophyll can be reduced in crop canopies with little penalty to photosynthesis. *Plant Physiology*, **176**, 1215–1232. Available from: https://doi.org/10.1104/pp.17.01401

Wamatu, J., Alkhtib, A. & Gardi, M.W. (2019) Leveraging traditional crops for food and feed: a case of hulless barley (*Hordeum vulgare*) landraces in Ethiopia. *Journal of Experimental Biology and Agricultural Sciences*, **7**, 42–50. Available from: https://doi.org/10.18006/2019.7(1).42.50

Wang, P., Liang, F.C., Wittmann, D., Siegel, A., Shan, S. & Grimm, B. (2018) Chloroplast SRP43 acts as a chaperone for enzyme in tetrapyrrole biosynthesis. *PNAS*, **115**, E3588-E3596. Available from: https://doi.org/10.1073/pnas.1719645115

Wang, Q., Nian, J., Xie, X., Yu, H., Zhang, J., Bai, J. *et al.* (2018) Genetic variations in ARE1 mediate grain yield by modulating nitrogen utilization in rice. *Nature Communications*, **9**, 735. Available from: https://doi.org/10.1038/s41467-017-02781-w

White, L.M., Hartman, G.P. & Bergman, J.W. (1981) In vitro digestibility, crude protein, and phosphorus content of straw of winter wheat, spring wheat, barley, and oat cultivars in eastern Montana. *Agronomy Journal*, **73**, 117–121. Available from: https://doi.org/10.2134/agronj1981.00021962007300010026x

Xin, C.P., Tholen, D., Devloo, V. & Zhu, X.G. (2015) The benefits of photorepiratory bypasses: how can they work? *Plant Physiology*, **167**, 574–585. Available from: https://doi.org/10.1104/pp.114.248013

XResearch Company. (2023) *Biocomposites market 2022–Segments analysis, industry trends, manufacturers analysis, opportunities and market size & forecast 2030*. Available at: www.xresearch.biz/shop/biocomposites-market

Yamamoto, H., Takahashi, S., Badger, M.R. & Shikanai, T. (2016) Artificial remodelling of alternative electron flow by flavodiiron proteins in Arabidopsis. *Nature Plants*, **2**, 16012. Available from: https://doi.org/10.1038/n plants.2016.12

Yamauchi, S., Takemiya, A., Sakamoto, T., Kurata, T., Tsutsumi, T., Kinoshita, T. *et al.* (2016) The plasma membrane H⁺-ATPase AHA1 plays a major role in stomatal opening in response to blue light. *Plant Physiology*, **171**, 2731–2743. Available from: https://doi.org/10.1104/pp.16.01581

Yang, L., Park, D. & Qin, Z. (2021) Material function of mycelium-based bio-composite: a review. *Frontiers in Materials*, **8**, 737377. Available from: https://doi.org/10.3389/fmats.2021.737377

Yeh, C., Wang, Y., Takahashi, Y., Kuusk, K., Paul, K., Arjus, T. *et al.* (2023) MPK12 in stomatal CO₂ signaling: function beyond its kinase activity. *The New Phytologist*, **239**, 146–158. Available from: https://doi.org/10.1111/nph.18913

Yu, M., Li, Z., Chen, W., Rong, T., Wang, G. & Ma, X. (2019) *Hermetia illucens* larvae as a potential dietary protein source altered the microbiota and modulated mucosal immune status in the colon of finishing pigs. *Journal of Animal Science and Biotechnology*, **10**, 50. Available from: https://doi.org/10.1186/s40104-019-0358-1

Zhang, M., Wang, Y., Chen, X., Xu, F., Ding, M., Ye, W. *et al.* (2021) Plasma membrane H⁺-ATPase overexpression increases rice yield via simultaneous enhancement of nutrient uptake and photosynthesis. *Nature Communications*, **12**, 735. Available from: https://doi.org/10.1038/s41467-021-20964-4

Zhang, M., Zhang, B., Qian, Q., Yu, Y., Li, R., Zhang, J. *et al.* (2010) Brittle culm 12, a dual-targeting kinesin-4 protein, controls cell-cycle progression and wall properties in rice. *The Plant Journal*, **63**, 312–328. Available from: https://doi.org/10.1111/j.1365-313X.2010.04238.x
Zhou, Y., Li, S., Qian, Q., Zeng, D., Zhang, M., Guo, L. et al. (2009) BC10, a DUF266-containing and Golgi-located type II membrane protein, is required for cell-wall biosynthesis in rice (Oryza sativa L.). *The Plant Journal*, **57**, 446-462. Available from: https://doi.org/10.1111/j.1365-313X. 2008.03703.x

Zhu, X.G., Ort, D.R., Whitmarsh, J. & Long, S.P. (2004) The slow reversibility of photosystem II thermal energy dissipation on transfer from high to

low light may cause large losses in carbon gain by crop canopies: a theoretical analysis. *Journal of Experimental Botany*, **55**, 1167–1175. Available from: https://doi.org/10.1093/jxb/erh141

Ziehe, D., Dünschede, B. & Schünemann, D. (2018) Molecular mechanism of SRP-dependent light-harvesting protein transport to the thylakoid membrane in plants. *Photosynthesis Research*, **138**, 303–313. Available from: https://doi.org/10.1007/s11120-018-0544-6