bluuebunny/crossref_metadata_2025 huggingface dataset:
Characterization of thePristionchus pacificus“epigenetic toolkit” reveals the evolutionary loss of the histone methyltransferase complex PRC2
Authors: Brown Audrey , Meiborg Adriaan B. , Franz-Wachtel Mirita , Macek Boris , Gordon Spencer , Rog Ofer , Weadick Cameron J , Werner Michael S.
Published: December 2023

Comparative approaches have revealed both divergent and convergent paths to achieving shared developmental outcomes. Thus, only through assembling multiple case studies can we understand biological principles. Yet, despite appreciating the conservation – or lack thereof – of developmental networks, the conservation of epigenetic mechanisms regulating these networks is poorly understood. The nematodePristionchus pacificushas emerged as a model system of plasticity and epigenetic regulation as it exhibits a bacterivorous or omnivorous morph depending on its environment. Here, we determined the “epigenetic toolkit” available toP. pacificusas a resource for future functional work on plasticity, and as a comparison withC. elegansto investigate the conservation of epigenetic mechanisms. Broadly, we observed a similar cast of genes with putative epigenetic function betweenC. elegansandP. pacificus. However, we also found striking differences. Most notably, the histone methyltransferase complex PRC2 appears to be missing inP. pacificus.We described the deletion/pseudogenization of the PRC2 genesmes-2andmes-6and concluded that both were lost in the last common ancestor ofP. pacificusand a related speciesP. arcanus.Interestingly, we observed the enzymatic product of PRC2 (H3K27me3) by mass spectrometry and immunofluorescence, suggesting that a currently unknown methyltransferase has been co-opted for heterochromatin silencing. Altogether, we have provided an inventory of epigenetic genes inP. pacificusto enable reverse-genetic experiments related to plasticity, and in doing so have described the first loss of PRC2 in a multicellular organism.
A ‘through-DNA’ mechanism for metal uptake-vs.-efflux regulation
Authors: Chakraborty Udit Kumar , Park Youngchan , Sengupta Kushal , Jung Won , Joshi Chandra P. , Francis Danielle H. , Chen Peng
Published: December 2023

Transition metals like Zn are essential for all organisms including bacteria, but fluctuations of their concentrations in the cell can be lethal. Organisms have thus evolved complex mechanisms for cellular metal homeostasis. One mechanistic paradigm involves pairs of transcription regulators sensing intracellular metal concentrations to regulate metal uptake and efflux. Here we report that Zur and ZntR, a prototypical pair of regulators for Zn uptake and efflux inE. coli, respectively, can coordinate their regulation through DNA, besides sensing cellular Zn2+concentrations. Using a combination of live-cell single-molecule tracking andin vitrosingle-molecule FRET measurements, we show that unmetallated ZntR can enhance the unbinding kinetics of Zur from DNA by directly acting on Zur-DNA complexes, possibly through forming heteromeric ternary and quaternary complexes that involve both protein-DNA and protein-protein interactions. This ‘through-DNA’ mechanism may functionally facilitate the switching in Zn uptake regulation when bacteria encounter changing Zn environments; it could also be relevant for regulating the uptake-vs.-efflux of various metals across different bacterial species and yeast.
Fungi and Bacteria Occupy Distinct Spatial Niches within Carious Dentin
Authors: Sulyanto Rosalyn M. , Beall Clifford J. , Ha Kasey , Montesano Joseph , Juang Jason , Dickson John R. , Hashmi Shahr B. , Bradbury Seth , Leys Eugene J. , Edgerton Mira , Ho Sunita P. , Griffen Ann L.
Published: December 2023

The role of bacteria in the etiology of dental caries is long established, while the role of fungi has only recently gained more attention. The microbial invasion of dentin in advanced caries especially merits additional research. We evaluated the fungal and bacterial community composition and spatial distribution within carious dentin. Amplicon 16S rRNA gene sequencing together with quantitative PCR was used to profile bacterial and fungal species in caries-free children (n=43) and 4 stages of caries progression from children with severe early childhood caries (n=32). Additionally, healthy (n=10) and carious (n=10) primary teeth were decalcified, sectioned, and stained with Grocott’s methenamine silver, periodic acid Schiff (PAS) and calcofluor white (CW) for fungi. Immunolocalization was also performed using antibodies against fungal β-D-glucan, gram-positive bacterial lipoteichoic acid,Streptococcus mutans, andCandida albicans. We also performed field emission scanning electron microscopy (FESEM) to visualize fungi and bacteria within carious dentinal tubules. Bacterial communities observed included a high abundance ofS. mutansand theVeillonella parvulagroup, as expected. There was a higher ratio of fungi to bacteria in dentin-involved lesions compared to less severe lesions with frequent preponderance ofC. albicans,C. dubliniensis, and in one caseC. tropicalis. Grocott’s silver, PAS, CW and immunohistochemistry (IHC) demonstrated the presence of fungi within carious dentinal tubules. Multiplex IHC revealed that fungi and gram-positive bacteria primarily occupied separate dentinal tubules, with rare instances of colocalization. Similar findings were observed with multiplex immunofluorescence using anti-S. mutansand anti-C. albicansantibodies. Electron microscopy showed monomorphic bacterial and fungal biofilms within distinct dentin tubules. We demonstrate a previously unrecognized phenomenon in which fungi and bacteria occupy distinct spatial niches within carious dentin and seldom co-colonize. The potential significance of this phenomenon in caries progression warrants further exploration.
Using Unstructured Crowd-sourced Data to Evaluate Urban Tolerance of Terrestrial Native Species within a California Mega-City
Authors: Curti Joseph N. , Barton Michelle , Flores Rhay G. , Lechner Maren , Lipman Alison , Montgomery Graham A. , Park Albert Y. , Rochel Kirstin , Tingley Morgan W.
Published: December 2023

In response to biodiversity loss and biotic community homogenization in urbanized landscapes, City managers around the world are increasingly working to conserve and increase urban biodiversity. Accordingly, around the world, previously extirpated species are (re)colonizing and otherwise infiltrating urban landscapes, while once abundant species are in various states of decline. Tracking the occurrence of traditionally urban intolerant species and loss of traditionally urban tolerant species should be a management goal of urban areas, but we generally lack tools to study this phenomenon. To address this, we first used species’ occurrences from iNaturalist, a large collaborative dataset of species observations, to measure an urban association index (UAI) for 967 native animal species that occur in the city of Los Angeles. On average, the occurrence of native species was negatively associated with our composite measure of urban intensity, with the exception of snails and slugs, which instead occur more frequently in areas of increased urban intensity. Next, we assessed 8,348 0.25 x 0.25 mile grids across the City of Los Angeles to determine the average grid-level UAI scores (i.e., a summary of the UAIs present in a grid cell, which we term Community Urban Tolerance Index or CUTI). We found that areas of higher urban intensity host more urban tolerant species, but also that taxonomic groups differ in their aggregate tolerance of urban areas, and that spatial patterns of tolerance vary between groups (e.g., mammals are not the same as birds). The framework established here was designed to be iteratively reevaluated by city managers of Los Angeles in order to track the progress of initiatives to preserve and encourage urban biodiversity, but can be rescaled to sample different regions within the city or different cities altogether to provide a valuable tool for city managers globally.
SNPs in genes related to the repair of damage to DNA in clinical isolates ofM. tuberculosis: a transversal and longitudinal approach
Authors: Pérez-Martínez DE. , Zenteno-Cuevas R
Published: December 2023

The presence of SNPs in genes related to DNA damage repair inM. tuberculosiscan trigger hypermutagenic phenotypes with a higher probability of generating drug resistance. The aim of this research was to compare the presence of SNPs in genes related to DNA damage repair between sensitive and DR isolates, as well as to describe the dynamics in the presence of SNPs inM. tuberculosisisolated from recently diagnosed TB patients of the state of Veracruz, Mexico. The presence of SNPs in the coding regions of 65 genes related to DNA damage repair was analyzed. Eighty-six isolates from 67 patients from central Veracruz state, Mexico, were sequenced. The results showed several SNPs in 14 genes that were only present in drug-resistant genomes. In addition, by following of 15 patients, it was possible to describe three different dynamics of appearance and evolution of non-synonymous SNPs in genes related to DNA damage repair: 1) constant fixed SNPs, 2) population substitution, and 3) gain of fixed SNPs. Further research is required to discern the biological significance of each of these pathways and their utility as markers of DR or for treatment prognosis.
Highly Multiplexed Fluorescence Microscopy with Spectrally Tunable Semiconducting Polymer Dots
Authors: Guo Ziyu , Poudel Chetan , Sarfatis Margaret C. , Yu Jiangbo , Wong Madeline , Chiu Daniel T. , Vaughan Joshua C.
Published: December 2023

Current studies of biological tissues require visualizing diverse cell types and molecular interactions, creating a growing need for versatile techniques to simultaneously probe numerous targets. Traditional multiplexed imaging is limited to around five targets at once. Emerging methods utilizing sequential rounds of staining, imaging, and signal removal can probe tens of targets but require specialized hardware, time-consuming workflows, and face some challenges with sample distortion and artifacts. Here we present a new method for highly-multiplexed fluorescence microscopy using semiconducting polymer dots (Pdots) in a single round of staining and imaging. Pdots are small, bright, and photostable fluorescent probes with a wide range of tunable Stokes shifts (20–450 nm). Multiple series of Pdots with varying excitation wavelengths allow for fast (<1 minute) and single-round imaging of up to 21 targets in brain and kidney. This method is based on a simple immunofluorescence workflow, efficient use of spectral space, standard hardware, and straightforward analysis, making it widely applicable for bioimaging laboratories.
DNDesign: Enhancing Physical Understanding of Protein Inverse Folding Model via Denoising
Authors: Lee Youhan , Kim Jaehoon
Published: December 2023

Based on the central dogma that protein structure determines its functionality, an important approach for protein sequence design is to identify promising sequences that fold into pre-designed structures based on domain knowledge. Numerous studies have introduced deep generative model-based inverse-folding, which utilizes various generative models to translate fixed backbones to corresponding sequences. In this work, we reveal that denoising training enables models to deeply capture the protein energy landscape, which previous models do not fully leverage. Based on this, we propose a novel Denoising-enhanced protein fixed backbone design (DNDesign), which combines conventional inverse-folding networks with a novel plug-in module, which learns the physical understanding via denoising training and transfers the knowledge to the entire network. Through extensive experiments, we demonstrate that DNDesign can easily be integrated into state-of-the-art models and improve performance in multiple modes, including auto-regressive, non-auto-regressive, and scaled-up scenarios. Furthermore, we introduce a fixed backbone conservation analysis based on potential energy changes, which confirms that DNDesign ensures more energetically favorable inverse-folding.
Rapid TetOn-mediated gene expression in neurons across the lifespan with uTTOP
Authors: Salmon Christopher K , Quesseveur Gael , Leo Cheong Gee Hung , Chierzi Sabrina , Green Miranda , Rosen Michael P , Gao Andy YL , Kacerovsky J Benjamin , Murai Keith K
Published: December 2023

Conditional expression of genes of interest is essential for interrogation of cellular development and function. Although tools exist for conditional gene expression, techniques for rapid-onset, temporally precise expression are lacking. The doxycyclineinducible TetOn expression system allows for this in numerous organ systems, however, transgenic TetOn expression cassettes become silenced in the nervous system during postnatal development. Here, we circumvent this silencing with uTTOP:in uteroelectroporation of Transposable TetOn Plasmids. When electroporated as transposable elements that integrate into the genome, the TetOn system allowed for robust DOX-dependent induction of expression across the postnatal lifespan of the mouse. We demonstrated induction in neurons of sensorimotor and retrospleninal cortex, hippocampus and the olfactory bulb. Latency to peak induction was ≤12 hours, a several fold increase in induction kinetics over existing methodology forin vivoconditional expression. To demonstrate the utility of uTTOP, we induced ectopic expression of Sonic hedgehog in adult mouse layer 2/3 cortical neurons, demonstrating that its expression can diversify expression of Kir4.1 in surrounding astrocytes. The rapid induction kinetics of uTTOP allowed us to show that Kir4.1 upregulation significantly lags onset of Shh expression by ∼2 days, a difference in expression time course that is likely not resolvable with current methods. Together, these data demonstrate that uTTOP is a powerful and flexible system for conditional gene expression in multiple brain areas across the mouse lifespan.
EPAS1 Attenuates Atherosclerosis Initiation at Disturbed Flow Sites through Endothelial Fatty Acid Uptake
Authors: Pirri Daniela , Tian Siyu , Tardajos-Ayllon Blanca , Irving Sophie , Donati Francesco , Allen Scott P. , Mammoto Tadanori , Vilahur Gemma , Miller Clint , Schenkel Torsten , Chambers Emily V. , Dunning Mark , Serbanovic-Canic Jovana , Botrè Francesco , Mammoto Akiko , Xu Suowen , Osto Elena , Han Weiping , Fragiadaki Maria , Evans Paul C.
Published: December 2023

Background

Atherosclerotic plaques form unevenly due to disturbed blood flow, causing localized endothelial cell (EC) dysfunction. Obesity exacerbates this process, but the underlying molecular mechanisms are unclear. The transcription factor EPAS1 (HIF2A) has regulatory roles in endothelium, but its involvement in atherosclerosis remains unexplored. This study investigates the potential interplay between EPAS1, obesity, and atherosclerosis.

Methods

Responses to shear stress were analysed using cultured porcine aortic EC exposed to flowin vitrocoupled with metabolic and molecular analyses, and by en face immunostaining of murine aortic EC exposed to disturbed flowin vivo. Obesity and dyslipidemia were induced in mice via exposure to high-fat diet or through Leptin gene deletion. The role of Epas1 in atherosclerosis was evaluated by inducible endothelial Epas1 deletion, followed by hypercholesterolemia induction (AAV-PCSK9; high-fat diet).

Results

En facestaining revealed EPAS1 enrichment at sites of disturbed blood flow that are prone to atherosclerosis initiation. Obese mice exhibited substantial reduction in endothelial EPAS1 expression, correlating with hyperlipidaemia. Sulforaphane, a compound with known atheroprotective effects, restored EPAS1 expression and concurrently reduced plasma triglyceride levels in obese mice. Consistently, triglyceride derivatives (free fatty acids) suppressed EPAS1 in cultured EC by upregulating the negative regulator PHD3. Clinical observations revealed that reduced plasma EPAS1 correlated with increased endothelial PHD3 in obese individuals. Functionally, endothelial EPAS1 deletion increased lesion formation in hypercholesterolemic mice, indicating an atheroprotective function. Mechanistic insights revealed that EPAS1 protects arteries by maintaining endothelial proliferation by positively regulating CD36 and LIPG expression to increase fatty acid beta-oxidation.

Conclusions

Endothelial EPAS1 attenuates atherosclerosis at sites of disturbed flow by maintaining EC proliferative via fatty acid uptake and metabolism. This endothelial repair pathway is inhibited in obesity, suggesting a novel triglyceride-PHD3 modulation pathway suppressing EPAS1 expression. These findings have implications for therapeutic strategies addressing vascular dysfunction in obesity.
Exploratory study of pulsed electric field ablation on atherosclerotic plaque in a rabbit model
Authors: Xuying Ye , Jiashen Hu , Shisheng Cao , Xinyu Xu , Zhixiao Xue , Chengzhi Lu , Huijuan Yin
Published: December 2023

New understanding of the pathogenesis of atherosclerotic diseases has led to the emergence of new therapeutic approaches. We explored the potential therapeutic effects of pulsed field potential ablation (PFA), a non-thermal ablation technique with high tissue selectivity, on atherosclerotic plaques. Carotid arteries of 30 high-fat rabbits were dilated with a balloon to obtain atherosclerotic plaques. PFA was administered on the carotid atherosclerotic plaques with 1000V/cm, 2000V/cm, and 1000V/cm ablation followed by rapamycin infusion. There were no visible changes in blood vessels after acute ablation, but apoptosis and polarity of cells were observed in atherosclerotic plaques. At 7 and 30 days after ablation, the density of lipid deposition in the plaque was significantly reduced, and multiple layers of new arranged anterograde smooth muscle cells appeared, replacing the original atherosclerotic plaque. The residual atherosclerotic structure is sandwiched between the new smooth muscle layer and the original smooth muscle layer, which makes vascular wall thicker and makes vascular wall elasticity increased. Rapamycin delays the vascular remodeling process. Conclusion: PFA ablation can reduce lipid deposition in atherosclerotic plaques, cause vascular remodeling, and enhance vascular elasticity. We believe that it may be a potential method for the treatment of atherosclerotic plaques.
Clinical-grade whole genome sequencing-based haplarithmisis enables all forms of preimplantation genetic testing
Authors: Janssen Anouk E.J. , Koeck Rebekka M. , Essers Rick , van Dijk Wanwisa , Drüsedau Marion , Meekels Jeroen , Yaldiz Burcu , van de Vorst Maartje , Cao Ping , de Koning Bart , Hellebrekers Debby M.E.I. , Stevens Servi J.C. , Sun Su Ming , Heijligers Malou , de Munnik Sonja A. , van Uum Chris M.J. , Achten Jelle , Hamers Lars , Naghdi Marjan , Vissers Lisenka E.L.M. , van Golde Ron J.T. , de Wert Guido , Dreesen Jos C.F.M. , de Die-Smulders Christine , Coonen Edith , Brunner Han G. , van den Wijngaard Arthur , Paulussen Aimee D.C. , Esteki Masoud Zamani
Published: December 2023

High-throughput sequencing technologies have increasingly led to discovery of disease-causing genetic variants, primarily in postnatal multi-cell DNA samples. However, applying these technologies to preimplantation genetic testing (PGT) in nuclear or mitochondrial DNA from single or few-cells biopsied fromin vitrofertilised (IVF) embryos is challenging. PGT aims to select IVF embryos without genetic abnormalities. Although genotyping-by-sequencing (GBS)-based haplotyping methods enabled PGT for monogenic disorders (PGT-M), structural rearrangements (PGT-SR), and aneuploidies (PGT-A), they are labour intensive, only partially cover the genome and are troublesome for difficult loci and consanguineous couples. Here, we devised a simple, scalable and universal whole genome sequencing haplarithmisis-based approach enabling all forms of PGT in a single assay. In a comparison to state-of-the-art GBS-based PGT for nuclear DNA (37 embryos, 18 families, 25 indications), shallow sequencing-based PGT (10 embryos, 3 families), and PCR-based PGT for mitochondrial DNA (10 embryos, 2 families), our approach alleviates technical limitations by decreasing whole genome amplification artifacts by 68.4%, increasing breadth of coverage by 4-fold, and reducing wet-lab turn-around-time by 2.5-fold. Importantly, this method enables trio-based PGT-A for aneuploidy origin, an approach we coin PGT-AO, detects translocation breakpoints, and nuclear and mitochondrial single nucleotide variants and indels in base-resolution.
LITERATURE REVIEW THE EFFECT OF MINDFULNESS INTERVENTIONS ON BLOOD PRESSURE AND STRESS IN HYPERTENSION PATIENTS
Authors: Mu’afiro Adin , Pandin Moses Glorino Rumambo , Nursalam
Published: December 2023

Background

Hypertension is a significant contributor to both illness and death on a global scale. One method of managing blood pressure is through adopting a healthy lifestyle and implementing blood pressure control measures, with or without the use of medication. The objective of the review paper is to elucidate the impact of mindfulness therapies on both blood pressure and stress levels in individuals diagnosed with hypertension.

Method

The research approach employed is literature observation, wherein relevant papers published between January 2019 and December 2023 are sought by conducting searches in four databases: Scopus and Science Direct. The search criteria include publications written in English, available in open access, and with full text. The study investigates the impact of mindfulness therapies on blood pressure and stress levels in individuals with hypertension, using Proquest and CINAHL as research databases

Results

A total of 14 papers have undergone evaluation. The primary emphasis lies in mindfulness techniques aimed at lowering blood pressure and alleviating stress. A mindfulness intervention was administered in group settings, with a duration of 2 hours per week for a period of 8 weeks. Subsequently, participants engage in physical activity inside the confines of their own homes, dedicating a minimum of 45 minutes per day for a duration of 6 to 10 days per week. They also monitor their blood pressure and stress levels. Blood pressure measurements are derived from the mean of three measurements done thrice day. Following the implementation of the mindfulness intervention, there was a notable reduction in both systolic and diastolic blood pressure among the intervention group, as compared to the first measurement. The investigation revealed substantial disparities between the experimental and control groups in the mean scores of positive stress (P = 0.001) and negative stress (P = 0.001).

Conclusion

Mindfulness can be a straightforward and cost-effective method for enhancing blood pressure and reducing stress.
A novel in vitro tubular model to recapitulate features of distal airways: The bronchioid
Authors: Maurat Elise , Raasch Katharina , Leipold Alexander M. , Henrot Pauline , Zysman Maeva , Prevel Renaud , Trian Thomas , Krammer Tobias , Bergeron Vanessa , Thumerel Matthieu , Nassoy Pierre , Berger Patrick , Saliba Antoine-Emmanuel , Andrique Laetitia , Recher Gaëlle , Dupin Isabelle
Published: December 2023

Background

Airflow limitation is the hallmark of obstructive pulmonary diseases, with the distal airways representing a major site of obstruction. Although numerousin vitromodels of bronchi already exist, there is currently no culture system for obstructive diseases that reproduces the architecture and function of small airways. Here, we aimed to engineer a model of distal airways to overcome the limitations of current culture systems.

Methods

We developed a so-called bronchioid model by encapsulating human bronchial adult stem cells derived from clinical samples in a tubular scaffold made of alginate gel.

Results

This template drives the spontaneous self-organization of epithelial cells into a tubular structure. Fine control of the level of contraction is required to establish a model of the bronchiole, which has a physiologically relevant shape and size. 3D imaging, gene expression and single-cell RNA-seq analysis of bronchioids made of bronchial epithelial cells revealed tubular organization, epithelial junction formation and differentiation into ciliated and goblet cells. Ciliary beating is observed, at a decreased frequency in bronchioids made of cells from COPD patients. The bronchioid can be infected by rhinovirus. An air-liquid interface is introduced that modulates gene expression.

Conclusion

Here, we provide a proof of concept of a perfusable bronchioid with proper mucociliary and contractile functions. The key advantages of our approach, such as the air-liquid interface, lumen accessibility, recapitulation of pathological features and possible assessment of clinically relevant endpoints, will make our pulmonary organoid-like model a powerful tool for preclinical studies.
Rapid modeling of an ultra-rare epilepsy variant in wild-type mice by in utero prime editing
Authors: Robertson Colin D. , Davis Patrick , Richardson Ryan R. , Iffland Philip H. , Vieira Daiana C. O. , Steyert Marilyn , McKeon Paige N. , Romanowski Andrea J. , Crutcher Garrett , Jašarević Eldin , Wolff Steffen B. E. , Mathur Brian N. , Crino Peter B. , Bale Tracy L. , Dick Ivy E. , Poulopoulos Alexandros
Published: December 2023

Generating animal models for individual patients within clinically-useful timeframes holds great potential toward enabling personalized medicine approaches for genetic epilepsies. The ability to rapidly incorporate patient-specific genomic variants into model animals recapitulating elements of the patient’s clinical manifestations would enable applications ranging from validation and characterization of pathogenic variants to personalized models for tailoring pharmacotherapy to individual patients. Here, we demonstrate generation of an animal model of an individual epilepsy patient with an ultra-rare variant of the NMDA receptor subunit GRIN2A, without the need for germline transmission and breeding. Using in utero prime editing in the brain of wild-type mice, our approach yielded high in vivo editing precision and induced frequent, spontaneous seizures which mirrored specific elements of the patient’s clinical presentation. Leveraging the speed and versatility of this approach, we introduce PegAssist, a generalizable workflow to generate bedside-to-bench animal models of individual patients within weeks. The capability to produce individualized animal models rapidly and cost-effectively will reduce barriers to access for precision medicine, and will accelerate drug development by offering versatile in vivo platforms to identify compounds with efficacy against rare neurological conditions.
Blue light irradiation induces pollen tube rupture in various flowering plants
Authors: Sugi Naoya , Susaki Daichi , Mizuta Yoko , Kinoshita Tetsu , Maruyama Daisuke
Published: December 2023

Pollen tubes exhibit one of the fastest apical growth rates among plant cells. Maintaining the proper balance between turgor pressure and cell wall synthesis at the pollen tube tip is crucial for this rapid growth, and any disruption can result in pollen tube rupture. In our study, we reveal that exposure to short-wavelength visible light, specifically blue light, induces pollen tube rupture. The frequency of pollen tube rupture increases in an intensity-dependent manner. Additionally, we observed Ca2+influx after blue light irradiation, accompanying with either pollen tube rupture or a temporary halt in elongation. These findings offer insights into the interplay between pollen tube integrity maintenance and Ca2+influx at the pollen tube tip, presenting a novel and efficient method to control pollen tube burst.

Subject Areas

(1) growth and development

(11) new methodology
SIRT5variants from patients with mitochondrial disease are associated with reduced SIRT5 stability and activity, but not with neuropathology
Authors: Yuan Taolin , Kumar Surinder , Skinner Mary , Victor-Joseph Ryan , Abuaita Majd , Keijer Jaap , Zhang Jessica , Kunkel Thaddeus J. , Liu Yanghan , Petrunak Elyse M. , Saunders Thomas L. , Lieberman Andrew P. , Stuckey Jeanne A. , Neamati Nouri , Al-Murshedi Fathiya , Alfadhel Majid , Spelbrink Johannes N. , Rodenburg Richard , de Boer Vincent C. J. , Lombard David B.
Published: December 2023

SIRT5 is a sirtuin deacylase that represents the major activity responsible for removal of negatively-charged lysine modifications, in the mitochondrial matrix and elsewhere in the cell. In benign cells and mouse models, under basal non-stressed conditions, the phenotypes of SIRT5 deficiency are generally quite subtle. Here, we identify two homozygousSIRT5variants in human patients suffering from severe mitochondrial disease. Both variants, P114T and L128V, are associated with reduced SIRT5 protein stability and impaired biochemical activity, with no evidence of neomorphic or dominant negative properties. The crystal structure of the P114T enzyme was solved and shows only subtle deviations from wild-type. Via CRISPR-Cas9, we generate a mouse model that recapitulates the human P114T mutation; homozygotes show reduced SIRT5 levels and activity, but no obvious metabolic abnormalities, neuropathology or other gross evidence of severe disease. We conclude that these humanSIRT5variants most likely represent severe hypomorphs, and are likely not the primary pathogenic cause of the neuropathology observed in the patients.
Trade-off between light deprivation and desiccation in intertidal seagrasses due to periodic tidal inundation and exposure: insights from a data-calibrated model
Authors: Wang Xinyan , Adams Matthew P. , Shao Dongdong
Published: December 2023

Many seagrass species thrive in shallow intertidal zones globally, adapting to periodic tidal inundation and exposure with distinctive physiological traits and offering crucial ecosystem services. However, predicting the responses of intertidal seagrasses to external stressors is hampered by the complexity of the dynamic and harsh environments they occupy. Consequently, intertidal seagrass growth models, especially those incorporating dynamic physiological responses, are scarce in the literature. Our study comprehensively collated relevant data from the literature to parameterize the relationship between air exposure, seagrass leaf water content and photosynthetic efficiency to inform new growth rate functions for generalisable intertidal seagrass growth models. We tested the applicability of these model formulations for scenarios with varying physiological process assumptions, seagrass species, tidal conditions, meadow elevations and water turbidity. We found that neglecting air-exposed physiological responses (i.e., leaf water content loss and reduced photosynthetic efficiency) can substantially overestimate seagrass growth rates. We also observed a trade-off between light deprivation and desiccation on intertidal seagrass growth under specific tidal ranges and turbidity conditions. This can yield an “optimal” elevation where combined stressors of desiccation (increasing with meadow elevation) and light deprivation (decreasing with meadow elevation) are minimized. The predicted optimal elevation, i.e., the most suitable habitat for intertidal seagrass, moves upward as water turbidity increases. Our study provides both conceptual and quantitative guidance for ecological modellers to include air exposure responses of intertidal seagrasses in coastal ecosystem models. The model also helps to evaluate the viability of intertidal seagrass habitats and inform site selection for seagrass restoration.
A 3-component module maintains sepal flatness in Arabidopsis
Authors: Xu Shouling , He Xi , Trinh Duy-Chi , Zhang Xinyu , Wu Xiaojiang , Qiu Dengying , Zhou Ming , Xiang Dan , Roeder Adrienne H. K. , Hamant Olivier , Hong Lilan
Published: December 2023

As in origami, morphogenesis in living systems heavily relies on tissue curving and folding, through the interplay between biochemical and biomechanical cues. In contrast, certain organs maintain their flat posture over several days. Here we identified a pathway, which is required for the maintenance of organ flatness, taking the sepal, the outermost floral organ, in Arabidopsis as a model system. Through genetic, cellular and mechanical approaches, our results demonstrate that global gene expression regulator VERNALIZATION INDEPENDENCE 4 (VIP4) fine-tunes the mechanical properties of sepal cell walls and maintains balanced growth on both sides of the sepals, mainly by orchestrating the distribution pattern of AUXIN RESPONSE FACTOR 3 (ARF3).vip4mutation results in softer cell walls and faster cell growth on the adaxial sepal side, which eventually cause sepals to bend outward. Downstream of VIP4, ARF3 works through modulating auxin signaling to down-regulate pectin methylesterase VANGUARD1, resulting in decreased cell wall stiffness. Our work unravels a 3-component module, which relates hormonal patterns to organ curvature, and actively maintains sepal flatness during its growth.
A maternal-to-zygotic-transition gene block on the zebrafish sex chromosome
Authors: Wilson Catherine A. , Postlethwait John H.
Published: December 2023

Wild zebrafish (Danio rerio) have a ZZ/ZW chromosomal sex determination system with the major sex locus on the right arm of chromosome-4 (Chr4R) near the largest heterochromatic block in the genome, suggesting the hypothesis that the Chr4R transcriptome might be different from the rest of the genome. We conducted an RNA-seq analysis of adult ZW ovaries and ZZ testes and identified four regions of Chr4 with different gene expression profiles. Unique in the genome, protein-coding genes in a 41.7 Mb section (Region-2) were expressed in testis but silent in ovary. The AB lab strain, which lacks sex chromosomes, verified this result, showing that testis-biased gene expression in Region-2 depends on gonad biology, not on sex-determining mechanism. RNA-seq analyses in female and male brain and liver validated few transcripts from Region-2 in somatic cells, but without sex-specificity. Region-2 corresponds to the heterochromatic portion of Chr4R and its content of genes and repetitive elements distinguishes it from the rest of the genome. In Region-2, protein-coding genes lack human orthologs; it has zinc finger genes expressed early in zygotic genome activation; it has maternal 5S rRNA genes, maternal spliceosome genes, a concentration of tRNA genes, and an distinct set of repetitive elements. The colocalization of 1) genes silenced in ovaries but not in testes that are 2) expressed in embryos briefly at the onset of zygotic genome activation; 3) maternal-specific genes for translation machinery; 4) maternal-specific spliceosome components; and 4) adjacent genes encoding miR-430, which mediates maternal transcript degradation, suggest that this is a Maternal-to-Zygotic-Transition Gene Regulatory Block.

ARTICLE SUMMARY

The wild zebrafish sex chromosome has a region, unique in the genome, that contains protein-coding genes silenced in ovaries but expressed in testes and transiently in the embryo as it begins to express its own genes. This region also contains maternal-specific genes encoding the protein-synthesis machinery used specifically by developing embryos, and molecules that target for degradation messenger RNAs that the mother stored in her eggs. This region defines a distinct maternal-to-zygotic-transition gene block.
Bioluminescence-Based Determination of Cytosolic Accumulation of Antibiotics inEscherichia coli
Authors: Dash Rachita , Holsinger Kadie A. , Chordia Mahendra D. , Sharifian Gh. Mohammad , Pires Marcos M.
Published: December 2023

Antibiotic resistance is an alarming public health concern that affects millions of individuals across the globe each year. A major challenge in the development of effective antibiotics lies in their limited ability to permeate into cells, noting that numerous susceptible antibiotic targets reside within the bacterial cytosol. Consequently, improving cellular permeability is often a key consideration during antibiotic development, underscoring the need for reliable methods to assess the permeability of molecules across cellular membranes. Currently, methods used to measure permeability often fail to discriminate between arrival within the cytoplasm and the overall association of molecules with the cell. Additionally, these techniques typically possess throughput limitations. In this work, we describe a luciferase-based assay designed for assessing the permeability of molecules into the cytosolic compartment of Gram-negative bacteria. Our findings demonstrate a robust system that can elucidate the kinetics of intracellular antibiotics accumulation in live bacterial cells in real time.
Mutational signature decomposition with deep neural networks reveals origins of clock-like processes and hypoxia dependencies
Authors: Serrano Colome Claudia , Canal Anton Oleguer , Seplyarskiy Vladimir , Weghorn Donate
Published: December 2023

DNA mutational processes generate patterns of somatic and germline mutations. A multitude of such mutational processes has been identified and linked to biochemical mechanisms of DNA damage and repair. Cancer genomics relies on these so-called mutational signatures to classify tumours into subtypes, navigate treatment, determine exposure to mutagens, and characterise the origin of individual mutations. Yet, state-of-the-art methods to quantify the contributions of different mutational signatures to a tumour sample frequently fail to detect certain mutational signatures, work well only for a relatively high number of mutations, and do not provide comprehensive error estimates of signature contributions. Here, we present a novel approach to signature decomposition using artificial neural networks that addresses these problems. We show that our approach, SigNet, outperforms existing methods by learning the prior frequencies of signatures and their correlations present in real data. Unlike any other method we tested, SigNet achieves high prediction accuracy even with few mutations. We used this to generate estimates of signature weights for more than 7500 tumours for which only whole-exome sequencing data are available. We then identified systematic differences in signature activity both as a function of epigenetic covariates and over the course of tumour evolution. This allowed us to decipher the origins of signatures SBS3, SBS5 and SBS40. We further discovered novel associations of mutational signatures with hypoxia, including strong positive correlations with the activities of clock-like and defective DNA repair mutational processes. These results provide new insights into the interplay between tumour biology and mutational processes and demonstrate the utility of our novel approach to mutational signature decomposition, a crucial part of cancer genomics studies.
ProteinNPT: Improving Protein Property Prediction and Design with Non-Parametric Transformers
Authors: Notin Pascal , Weitzman Ruben , Marks Debora S. , Gal Yarin
Published: December 2023

Protein design holds immense potential for optimizing naturally occurring proteins, with broad applications in drug discovery, material design, and sustainability. How-ever, computational methods for protein engineering are confronted with significant challenges, such as an expansive design space, sparse functional regions, and a scarcity of available labels. These issues are further exacerbated in practice by the fact most real-life design scenarios necessitate the simultaneous optimization of multiple properties. In this work, we introduce ProteinNPT, a non-parametric trans-former variant tailored to protein sequences and particularly suited to label-scarce and multi-task learning settings. We first focus on the supervised fitness prediction setting and develop several cross-validation schemes which support robust perfor-mance assessment. We subsequently reimplement prior top-performing baselines, introduce several extensions of these baselines by integrating diverse branches of the protein engineering literature, and demonstrate that ProteinNPT consistently outperforms all of them across a diverse set of protein property prediction tasks. Finally, we demonstrate the value of our approach for iterative protein design across extensive in silico Bayesian optimization and conditional sampling experiments.
Contextual and Combinatorial Structure in Sperm Whale Vocalisations
Authors: Sharma Pratyusha , Gero Shane , Payne Roger , Gruber David F. , Rus Daniela , Torralba Antonio , Andreas Jacob
Published: December 2023

Sperm whales(Physeter macrocephalus)are long-lived and highly social mammals that engage in complex group behaviours, including navigation, foraging, and child-rearing. During these behaviours, sperm whales communicate primarily using sequences of short bursts of clicks with varying inter-click intervals, known as codas. Past research has identified around 150 discrete coda types globally, with 21 in the Caribbean. A subset of these have been shown to encode information about caller and clan identity. However, almost everything else about the sperm whale communication system, including basic questions about its structure and information-carrying capacity, remains unknown. In this study, we show that codas exhibitcontextualandcombinatorialstructure with key similarities to aspects of human language and other primate communication systems. First, we report previously undescribed variations in coda structure that are sensitive to the conversational context in which they occur. We call theserubatoandornamentation, by analogy to musical terminology. These variations are systematically controlled and imitated across individual whales. Second, we show that coda types are not defined by arbitrary sequences of inter-click intervals, but instead form a combinatorial coding system in which rubato and ornamentation combine with two categorical, context-independent features that we callrhythmandtempoto give rise to a large inventory of distinguishable codas. In a dataset of 8,719 codas from the sperm whales of the Eastern Caribbean clan, this ‘sperm whale phonetic alphabet’ makes it possible to systematically explain observed variability in coda structure. Sperm whale vocalisations are more expressive and structured than previously believed, and are built from a repertoire comprising nearly an order of magnitude more distinguishable codas. These results show contextsensitive and combinatorial vocalisation systems extend beyond humans, and can appear in an organism with a divergent evolutionary lineage and vocal apparatus.
Selective lipid recruitment by an archaeal DPANN symbiont from its host
Authors: Ding Su , Hamm Joshua N. , Bale Nicole J. , Sinninghe Damsté Jaap S. , Spang Anja
Published: December 2023

The symbiontCa.Nanohaloarchaeum antarcticus is obligately dependent on its hostHalorubrum lacusprofundifor lipids and other metabolites due to its lack of certain biosynthetic genes. However, it remains unclear which specific lipids or metabolites are acquired from its host, and how the host responds to infection. Here, we explored the lipidome dynamics of theCa.Nha. antarcticus –Hrr. lacusprofundisymbiotic relationship during co-cultivation. By using a comprehensive untargeted lipidomic methodology, our study reveals thatCa.Nha. antarcticus selectively recruits 110 lipid species from its host, i.e. nearly two-thirds of the total number of host lipids. Lipid profiles of co-cultures displayed shifts in abundances of bacterioruberins and menaquinones and changes in degree of bilayer-forming glycerolipid unsaturation. This likely results in increased membrane fluidity and improved resistance to membrane disruptions, consistent with compensation for higher metabolic load and mechanical stress on host membranes when in contact withCa.Nha. antarcticus cells. Notably, our findings differ from previous observations of other DPANN symbiont-host systems, where no differences in lipidome composition were reported. Altogether, our work emphasizes the strength of employing untargeted lipidomics approaches to provide details into the dynamics underlying a DPANN symbiont-host system.
CRISPR-HOLMES-based NAD+detection
Authors: Zhuang Songkuan , Hu Tianshuai , Zhou Hongzhong , He Shiping , Li Jie , Zhang Yuehui , Gu Dayong , Xu Yong , Chen Yijian , Wang Jin
Published: December 2023

Studies have indicated that intracellular nicotinamide adenine dinucleotide (NAD+) level is associated with the occurrence and development of many diseases. While traditional NAD+detection techniques are time-consuming and may require large and expensive instruments. We recently found that the CRISPR-Cas12a protein can be inactivated by AcrVA5-mediated acetylation and reactivated by CobB, using NAD+as the co-factor. Therefore, here in this study, we created a CRISPR-Cas12a-based one-step HOLMES(NAD) system for rapid and convenient NAD+detection with the employment of both acetylated Cas12a and CobB. In HOLMES(NAD), acetylated Cas12a losses itstrans-cleavage activates and can be reactivated by CobB at the presence of NAD+, cutting ssDNA reporters to generate fluorescence signals. HOLMES(NAD) shows both sensitivity and specificity in NAD+detection and can be used for quantitative determination of intracellular NAD+concentrations. Therefore, HOLMES(NAD) not only provides a convenient and rapid approach for target NAD+quantitation but also expands the application scenarios of HOLMES to non-nucleic acid detection.
Metabolomics of Mouse Embryonic CSF Following Maternal Immune Activation
Authors: Petrova Boryana , Lacey Tiara E , Culhane Andrew J , Cui Jin , Raskin Alexander , Misra Aditya , Lehtinen Maria K , Kanarek Naama
Published: December 2023

The cerebrospinal fluid (CSF) serves various roles in the developing central nervous system (CNS), from neurogenesis to lifelong cognitive functions. Changes in CSF composition due to inflammation can impact brain function. We recently identified an abnormal cytokine signature in embryonic CSF (eCSF) following maternal immune activation (MIA), a mouse model of autism spectrum disorder (ASD). We hypothesized that MIA leads to other alterations in eCSF composition and employed untargeted metabolomics to profile changes in the eCSF metabolome in mice after inducing MIA with polyI:C. We report these data here as a resource, include a comprehensive MS1and MS2reference dataset, and present additional datasets comparing two mouse strains (CD-1 and C57Bl/6) and two developmental time points (E12.5 and E14.5). Targeted metabolomics further validated changes upon MIA. We show a significant elevation of glucocorticoids and kynurenine pathway related metabolites. Both pathways are relevant for suppressing inflammation or could be informative as disease biomarkers. Our resource should inform future mechanistic studies regarding the etiology of MIA neuropathology and roles and contributions of eCSF metabolites to brain development.