# The maize low-lignin mutant F2bm3 shows pleiotropic effects on photosynthetic and cell wall metabolisms in response to chilling

Catalina Duran Garzon, Michelle Lequart, Léo Bellenger, Alejandro Lovo, Françoise Fournet, Solène Bassard, Hélène Sellier-Richard, Paulo Marcelo, Karine Pageau, Catherine Giauffret, et al.

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Catalina Duran Garzon, Michelle Lequart, Léo Bellenger, Alejandro Lovo, Françoise Fournet, et al.. The maize low-lignin mutant F2bm3 shows pleiotropic effects on photosynthetic and cell wall metabolisms in response to chilling. cell wall meeting, 2019, Cambridge, United Kingdom. hal-04454441

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# The maize low-lignin mutant F2bm3 shows pleiotropic effects on photosynthetic and cell wall metabolisms in response to chilling

Catalina Duran Garzon¹, Michelle Lequart¹, Léo Bellenger¹, Alejandro Lovo⁵, Françoise Fournet¹, Solène Bassard¹, Hélène Sellier-Richard², Paulo Marcelo³, Karine Pageau¹, Catherine Giauffret⁴, Wilfred Vermerris⁵, Jean-Marc Domon¹, Catherine Rayon¹

¹EA3900-BIOPI, Biologie des Plantes et Innovation, Université de Picardie Jules Verne, 80039, Amiens, France ; ²Unité Expérimentale Grandes Cultures Innovation et Environnement, INRA-Estrées-Mons, 80203 Péronne, France ; ³Plateforme d'Ingénierie Cellulaire & Analyses des Protéines ICAP Université de Picardie Jules Verne, 80039 Amiens, France ; ⁴UR AgrolImpact, INRA, Estrées-Mons, 80203 Péronne, France ; ⁵Department of Microbiology & Cell Science, University of Florida, Gainesville, FL 32610, USA.

## Introduction

Maize (Zea mays L.) is one of the major crops in the world and is highly sensitive to low temperature due to its tropical origin. Chilling may particularly affect maize during early seedling growth by altering physiological processes including photosynthesis and cell wall properties, leading to biomass reduction. Changes in photosynthetic and cell wall metabolisms were investigated during a long chilling exposure in a low-lignin maize mutant, *brown midrib3* (*bm3*), which contains a null-mutation in the gene encoding caffeic acid O-methyltransferase (*COMT*). Wild-type F2 plants and the near-isogenic *F2bm3* mutant were grown during 60 days with a day/night temperature regime of 15°C/11°C in a greenhouse. Photosynthetic pigments, cell wall sugars, lignin, cell wall bound hydroxycinnamic acids (HCA) contents were determined when the 4th leaf had fully emerged.
Does *COMT* mutation compensate or increase deleterious chilling effects on growth or development ?

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comt WT

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Fornalé et al., 2017

## 1 Experimental growth conditions

Greenhouse

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## 4 Chlorophyll and zeaxanthin pigments are more abundant in F2bm3 than in F2 under chilling exposure

<table><caption>Some photosynthetic pigments content in F2 and F2bm3 leaves</caption><thead><tr><th rowspan="3">Pigment</th><th colspan="2">25°C/22°C</th><th colspan="2">15°C/11°C</th></tr><tr><th colspan="2">F2</th><th colspan="2">F2bm3</th></tr><tr><th>SD</th><th>SD</th><th>SD</th><th>SD</th></tr></thead><tbody><tr><td>Chl a (µmol.g<sup>-1</sup>FW)</td><td>2.77 ± 0.19 b</td><td>2.47 ± 0.13 b</td><td>6.23 ± 0.12 a</td><td>5.95 ± 0.16 a</td></tr><tr><td>Chl b (µmol.g<sup>-1</sup>FW)</td><td>0.78 ± 0.11 b</td><td>0.74 ± 0.12 b</td><td>0.93 ± 0.05 a</td><td>0.67 ± 0.10 b</td></tr><tr><td>Chl a+b (µmol.g<sup>-1</sup>FW)</td><td>3.55 ± 0.30 b</td><td>3.21 ± 0.13 b</td><td>7.16 ± 0.14 a</td><td>6.62 ± 0.22 a</td></tr><tr><td>Chl a/b</td><td>3.58 ± 0.29 c</td><td>3.41 ± 0.73 c</td><td>6.72 ± 0.38 b</td><td>9.04 ± 1.19 a</td></tr><tr><td>Zeaxanthin (µmol.g<sup>-1</sup>FW)</td><td>0.003 ± 0.000 c</td><td>0.015 ± 0.002 b</td><td>0.002 ± 0.001 c</td><td>0.070 ± 0.001 a</td></tr><tr><td>Carotenoid /Chl a+b</td><td>0.56 ± 0.05 a</td><td>0.60 ± 0.10 a</td><td>0.25 ± 0.02 b</td><td>0.24 ± 0.02 b</td></tr></tbody></table>

Data are means ±SD (n=16). Differences were statistically analyzed using Student's t-test. Different letters indicate significant differences between groups (P < 0.05).

## 6 Chilling increases hydroxycinnamic acids in F2bm3

<table><caption>Cell wall hydroxycinnamates content in F2 and F2bm3 leaves</caption><thead><tr><th rowspan="3">Genotype</th><th colspan="2">25°C/22°C</th><th colspan="2">15°C/11°C</th></tr><tr><th colspan="2">F2</th><th colspan="2">F2bm3</th></tr><tr><th>SD</th><th>SD</th><th>SD</th><th>SD</th></tr></thead><tbody><tr><td>FA (Ester) (mg.g<sup>-1</sup> DW)</td><td>0.13 ± 0.07 b</td><td>0.36 ± 0.09 a</td><td>0.24 ± 0.13 b</td><td>0.41 ± 0.05 a</td></tr><tr><td>FA ester /Ara</td><td>9.63 ± 5.86 a</td><td>12.89 ± 1.47 a</td><td>15.18 ± 8.53 a</td><td>14.93 ± 0.84 a</td></tr><tr><td>FA (Ether) (mg.g<sup>-1</sup> DW)</td><td>0.14 ± 0.05 b</td><td>0.13 ± 0.04 b</td><td>0.18 ± 0.02 b</td><td>0.28 ± 0.08 a</td></tr><tr><td>TOTAL FA (mg.g<sup>-1</sup> DW)</td><td>0.27 ± 0.12 b</td><td>0.49 ± 0.08 b</td><td>0.41 ± 0.13 b</td><td>0.70 ± 0.10 a</td></tr><tr><td>pCA (Ester) (mg.g<sup>-1</sup> DW)</td><td>0.66 ± 0.09 a</td><td>0.36 ± 0.03 b</td><td>0.34 ± 0.06 b</td><td>0.40 ± 0.08 b</td></tr><tr><td>pCA (Ether) (mg.g<sup>-1</sup> DW)</td><td>0.48 ± 0.06 a</td><td>0.25 ± 0.15 b</td><td>0.26 ± 0.03 b</td><td>0.23 ± 0.06 b</td></tr><tr><td>TOTAL pCA (mg.g<sup>-1</sup> DW)</td><td>1.14 ± 0.11 a</td><td>0.60 ± 0.37 b</td><td>0.60 ± 0.08 b</td><td>0.63 ± 0.13 b</td></tr><tr><td>TOTAL FA + pCA (mg.g<sup>-1</sup> DW)</td><td>1.43 ± 0.21 a</td><td>1.09 ± 0.45 b</td><td>1.02 ± 0.21 b</td><td>1.32 ± 0.23 a</td></tr><tr><td>pCA/FA</td><td>4.88 ± 2.45 a</td><td>1.18 ± 0.59 b</td><td>1.60 ± 0.65 b</td><td>0.92 ± 0.28 b</td></tr></tbody></table>

Data are means ±SD (n=16). Differences were statistically analyzed using Student's t-test. Different letters indicate significant differences between groups (P < 0.05). Ferulic acid (FA), p-coumaric acid (pCA), arabinose (Ara).

## 7 Chilling does not change lignin content but H-residues increases while a reduction in S- and G-units is observed in F2bm3

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### Lignin content using acetylbromide method

Data are means ±SD (n=16). Differences were statistically analyzed using Student's t-test. Letters denote differences at P<0.05, *a* between F2 and F2bm3 with a day/night temperature regime of 25°C/22°C, *b* between F2 and F2bm3 with a day/night temperature regime of 15°C/11°C, *c* and *d* between temperatures for each genotype. Black columns represents the experiment under ambient temperature condition and grey columns under chilling exposure.

<table><caption>Identification of H, S and G phenylpropanoid units (%) by thioacydolysis in F2 and F2bm3 leaves.</caption><thead><tr><th rowspan="3">Genotype</th><th colspan="2">25°C/22°C</th><th colspan="2">15°C/11°C</th></tr><tr><th colspan="2">F2</th><th colspan="2">F2bm3</th></tr><tr><th>SD</th><th>SD</th><th>SD</th><th>SD</th></tr></thead><tbody><tr><td>H</td><td>7.00 ± 0.06</td><td>2.00 ± 0.03</td><td></td><td></td></tr><tr><td>G</td><td>64.00 ± 0.12</td><td>59.00 ± 0.02</td><td></td><td></td></tr><tr><td>S</td><td>29.00 ± 0.06</td><td>39.00 ± 0.04</td><td></td><td></td></tr></tbody></table>

<table><thead><tr><th rowspan="3">Genotype</th><th colspan="2">25°C/22°C</th><th colspan="2">15°C/11°C</th></tr><tr><th colspan="2">F2bm3</th><th colspan="2">F2bm3</th></tr><tr><th>SD</th><th>SD</th><th>SD</th><th>SD</th></tr></thead><tbody><tr><td>H</td><td>0.00 ± 0.00</td><td>14.00 ± 0.03</td><td></td><td></td></tr><tr><td>G</td><td>82.00 ± 0.13</td><td>73.00 ± 0.07</td><td></td><td></td></tr><tr><td>S</td><td>18.00 ± 0.13</td><td>13.00 ± 0.04</td><td></td><td></td></tr></tbody></table>

**Acknowledgments:** This work was financed by the «Conseil Régional Hauts de France» and «FEDER», in support of the REG15019-COOLBIOM project (2015–2018). We thank Dr Matthieu Reymond (IJPB, INRA Versailles, France) for providing the seeds.

Farnale et al (2017) Plant and Cell Physiology, Volume 58, Issue 2, February 2017, Pages 240–255, https://doi.org/10.1093/pcp/pcw198

* The increase in arabinose substitution on the xylan backbone and the abundance of HCA in the F2bm3 mutant could reinforce the cell wall and maintain the hydration status of the cell wall during chilling.

* The reduction in G-residues in F2bm3 may be due to a greater demand in ferulic acids as photoprotectant under chilling.

* Therefore, the F2bm3 displays photosynthesis and cell wall traits that might be useful to cope with early sowing stress.

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A: L-Arabinose; GAX: Glucuronoarabinoxylan; FA: Ferulic acid; pCA: p-Coumaric acid
*: lignin is generally considered as a mixture of all three units but to show the variations of lignin units observed between genotypes, we separated them.

## Conclusion