A MYB/ZML complex regulates wound-induced lignin genes in maize

Lignin is an essential polymer in vascular plants that plays key structural roles in vessels and fibers. Lignification is induced by external inputs such as wounding, but the molecular mechanisms that link this stress to lignification remain largely unknown. In this work, we provide evidence that three maize (Zea mays) lignin repressors, MYB11, MYB31, and MYB42, participate in wound-induced lignification by interacting with ZML2, a protein belonging to the TIFY family. We determined that the three R2R3-MYB factors and ZML2 bind in vivo to AC-rich and GAT(A/C) cis-elements, respectively, present in a set of lignin genes. In particular, we show that MYB11 and ZML2 bind simultaneously to the AC-rich and GAT(A/C) cis-elements present in the promoter of the caffeic acid O-methyl transferase (comt) gene. We show that, like the R2R3-MYB factors, ZML2 also acts as a transcriptional repressor. We found that upon wounding and methyl jasmonate treatments, MYB11 and ZML2 proteins are degraded and comt transcription is induced. Based on these results, we propose a molecular regulatory mechanism involving a MYB/ZML complex in which wound-induced lignification can be achieved by the derepression of a set of lignin genes.Research in D.C.-R.'s laboratory was supported by a grant from the Spanish Ministry of Science and Education (AGL2011-30545-C02-01), the “Xarxa de Referència de Biotecnologia” (XarBa) from the Autonomous Government of Catalonia, the CONSOLIDER-INGENIO program (CSD2007-00036) from the Spanish Ministry of Science and Innovation, and the SGR programs (SGR2009-GRC703). Research in M.P.'s laboratory was supported by two grants from the Spanish Ministry of Science and Education (BIO2009-13044-C02-01 and BIO2012-31860), the framework of the XarBa, and the SGR programs (SGR2009-GRC626) from the Autonomous Government of Catalonia. Research in R.S.'s laboratory was supported by grants from the Ministry of Science and Innovation to R.S. (BIO2013-44407). M.P. and R.S. received financial support from the CONSOLIDER-INGENIO program (CSD2007-00057-B) from the Spanish Ministerio de Ciencia e Innovación. Research in the W.S. laboratory is supported by grants from the Ministry of Science and Technology and Academia Sinica. Research in phenylpropanoid gene regulation in the laboratories of E.G. and J.G. was supported by a grant from the National Science Foundation (IOS-1125620). I.-C.V.-B. was supported by a Spanish FPI Fellowship (BES-2007-17316). J.E.S.-H. was supported by the Department of Innovation, Universities and Enterprise of the Generalitatde Catalunya, the European Social Fund FI Fellowship (AGAUR: FI-2006, Resolució EDU/3600/2006; FI-2008, Resolució IUE/2658/2007 and BE-DGR2010), and CRAG.Peer reviewe

Elucidating compositional factors of maize cell walls contributing to stalk strength and lodging resistance

Lodging is one of the causes of maize (Zea mays L.) production losses worldwide and, at least, the resistance to stalk lodging has been positively correlated with stalk strength. In order to elucidate the putative relationship between cell wall, stalk strength and lodging resistance, twelve maize inbreds varying in rind penetration strength and lodging resistance were characterized for cell wall composition and structure. Stepwise multiple regression indicates that H lignin subunits confer a greater rind penetration strength. Besides, the predictive model for lodging showed that a high ferulic acid content increases the resistance to lodging, whereas those of diferulates decrease it. These outcomes highlight that the strength and lodging susceptibility of maize stems may be conditioned by structural features of cell wall rather than by the net amount of cellulose, hemicelluloses and lignin. The results presented here provide biotechnological targets in breeding programs aimed at improving lodging in maize.This work was funded by Projects AGL2014−58126-R and RTC-2016−5816-2 from the Spanish Ministry of Science, Innovation and Universities. This work was also supported by the CERCA Program and the SGR program (SGR-710) from the Generalitat de Catalunya. We acknowledge financial support from the Spanish Ministry of Economy and Competitiveness, through the “Severo Ochoa Program for Centres of Excellence in R&D” 2016–2019 (SEV‐2015‐0533)”. Alba Manga-Robles’s contract was granted by the Junta de Castilla y León and the Fondo Social Europeo through “Sistema Nacional de Garantía Juvenil” Program, Universidad de León and Junta de Castilla y León predoctoral Programs. Rogelio Santiago acknowledges a postdoctoral contract “Ramón y Cajal” financed by the Ministry of Economy and Competitiveness of Spain (RYC-2012-10603).Peer reviewe

CRISPR/CAS9-mediated genome editing of CmNAC-Nor delays fruit ripening in melon / Development of genetic and genomic tools for a Papaver somniferum breeding program

Trabajo presentado al Seminario del CRAG (Internal Seminar), celebrado online el 25 de junio de 2021.Peer reviewe

Towards the identification of rusticity-related QTLs in Papaver somniferum

Trabajo presentado al Internal Seminar of the Centre de Recerca Agrigenómica (CRAG), celebrado el 12 de julio de 2019

Isolation and characterisation of a family of laccases in maize

Plant laccases are enzymes that have been proposed to participate in the last step of lignin biosynthesis. The polymerisation event remains still much unknown, implicating other enzymes such as peroxidases. To gain more insight in how this polymerisation process takes place in maize, we isolated by differential screening of an elongation maize root cDNA library four cDNA clones encoding a family of laccases. Three of them (ZmLac2, ZmLac4, ZmLac5) were basic enzymes, while one of them (ZmLac3) was an acidic enzyme. Southern analysis indicates that laccases belong to a multigene family in maize. Phylogenetic analysis reveals that ZmLac2, ZmLac4, and ZmLac5 are closely related enzymes, whereas ZmLac3 is a slightly different enzyme. The pattern of mRNA accumulation of ZmLac2, ZmLac3, ZmLac4, and ZmLac5 genes correlates with the maize regions undergoing lignification. Moreover, ZmLac3 is induced by wounding, whereas ZmLac2 and ZmLac5 are repressed and ZmLac4 gene expression is not affected. Taken together, our results suggest that the acidic ZmLac3 enzyme could be involved in the polymerisation of phenolic compounds in maize. Instead, and in agreement with the idea that laccases are enzymes involved in a wide range of physiological processes, results obtained with ZmLac2, ZmLac4, and ZmLac5 lead us to exclude a direct role of these laccases in lignin polymerisation.This work has been funded by the Spanish “Ministerio de Ciencia y Tecnología” (BIO2001-1140). We are indebted to the sequencing team of IBMB-CSIC as well as Dr. Castresana (IBMB-CSIC) for his advices and comments on phylogenetic analyses performed in this work. D-C.R was initially financed by the European COPOL Project QLRT-1999-31493 and later on by the Spanish “Ministerio de Educacion y Ciencia” (“Ramon y Cajal” Program). S-F was financed by a post-doctoral grant of the “Generalitat de Catalunya” (2004-CRED-10005). We are indebted to Dr. Burgess for the English correction of this manuscript. This work was carried out within the framework of the Centre de Referència de Biotecnologia (CeRBA) from the Generalitat de Catalunya.Peer reviewe

Diverging cell wall strategies for drought adaptation in two maize inbreds with contrasting lodging resistance

The plant cell wall is a plastic structure of variable composition that constitutes the first line of defence against environmental challenges. Lodging and drought are two stressful conditions that severely impact maize yield. In a previous work, we characterised the cell walls of two maize inbreds, EA2024 (susceptible) and B73 (resistant) to stalk lodging. Here, we show that drought induces distinct phenotypical, physiological, cell wall, and transcriptional changes in the two inbreds, with B73 exhibiting lower tolerance to this stress than EA2024. In control conditions, EA2024 stalks had higher levels of cellulose, uronic acids and p‐coumarate than B73. However, upon drought EA2024 displayed increased levels of arabinose‐enriched polymers, such as pectin‐arabinans and arabinogalactan proteins, and a decreased lignin content. By contrast, B73 displayed a deeper rearrangement of cell walls upon drought, including modifications in lignin composition (increased S subunits and S/G ratio; decreased H subunits) and an increase of uronic acids. Drought induced more substantial changes in gene expression in B73 compared to EA2024, particularly in cell wall‐related genes, that were modulated in an inbred‐specific manner. Transcription factor enrichment assays unveiled inbred‐specific regulatory networks coordinating cell wall genes expression. Altogether, these findings reveal that B73 and EA2024 inbreds, with opposite stalk‐lodging phenotypes, undertake different cell wall modification strategies in response to drought. We propose that the specific cell wall composition conferring lodging resistance to B73, compromises its cell wall plasticity, and renders this inbred more susceptible to drought.This work was supported by the Grants AGL2014-58126-R; RTC-2016-5816-2 and PID2022-142786NB-I00 funded by MCIN/AEI/10.13039/501100011033 and ERDF “A way to make Europe” and received financial support from the CONSOLIDER-INGENIO programme (CSD2007-00036) from the Spanish Ministerio de Ciencia e Innovación. S.C. was financed with a PhD contract (PRE2019-089329) funded by MCIN/AEI/10.13039/501100011033 and by “ESF Investing in your future”, and by SEV-2015-0533-19-1 and CEX2019-000902-S funded by MCIN/AEI/10.13039/501100011033. A.M.-R. was financed with a PhD contract from the Consejería de Educación de Castilla y León and the Fondo Social Europeo (ORDEN EDU/601/2020, July 7th). This work was also supported by the SGR programmes (2017SGR710 and 2021-SGR-01131) from the Secretaria d'Universitats i Recerca del Departament d'Empresa i Coneixement de la Generalitat de Catalunya and by the CERCA Programme/Generalitat de Catalunya. This work was financially supported by RYC2021-033414-l to R.U funded by MCIN/AEI/10.13039/501100011033 by the “European Union Next Generation EU/PRTR”. Finally, we acknowledge financial support from the Grants SEV-2015-0533-19-1 and CEX2019-000902-S funded by MCIN/AEI/10.13039/501100011033.info:eu-repo/semantics/publishedVersio

AtMYB7, a new player in the regulation of UV-sunscreens in Arabidopsis thaliana

The phenylpropanoid metabolic pathway provides a wide variety of essential compounds for plants. Together with sinapate esters, in Brassicaceae species, flavonoids play an important role in protecting plants against UV irradiation. In this work we have characterized Arabidopsis thaliana AtMYB7, the closest homolog of AtMYB4 and AtMYB32, described as repressors of different branches of phenylpropanoid metabolism. The characterization of atmyb7 plants revealed an induction of several genes involved in flavonol biosynthesis and an increased amount of these compounds. In addition, AtMYB7 gene expression is repressed by AtMYB4. As a consequence, the atmyb4 mutant plants present a reduction of flavonol contents, indicating once more that AtMYB7 represses flavonol biosynthesis. Our results also show that AtMYB7 gene expression is induced by salt stress. Induction assays indicated that AtMYB7 represses several genes of the flavonoid pathway, DFR and UGT being early targets of this transcription factor. The results obtained indicate that AtMYB7 is a repressor of flavonol biosynthesis and also led us to propose AtMYB4 and AtMYB7 as part of the regulatory mechanism controlling the balance of the main A. thaliana UV-sunscreens.This work was supported by the Spanish ‘Ministerio de Economía y Competitividad’ [AGL2011-30545-C02-01 to D.C.-R.]; the Spanish Ministerio de Ciencia e Innovación [CONSOLIDER-INGENIO program (CSD2007-00036)]; Autonomous Government of Catalonia [grant AGAUR (2008FI-B 00399) to J.E.S.-H.]; CRAG [a contract to J.E.S.-H.]. This work was carried out within the framework of the ‘Xarxa de Referència de Biotecnologia’ (XarBa) from the Autonomous Government of Catalonia.Peer reviewe

Down-regulation of the maize and Arabidopsis thaliana caffeic acid O-methyl-transferase genes by two new maize R2R3-MYB transcription factors

The maize (Zea mays L.) caffeic acid O-methyl-transferase (COMT) is a key enzyme in the biosynthesis of lignin. In this work we have characterized the involvement of COMT in the lignification process through the study of the molecular mechanisms involved in its regulation. The examination of the maize COMT gene promoter revealed a putative ACIII box, typically recognized by R2R3-MYB transcription factors. We used the sequence of known R2R3-MYB factors to isolate five maize R2R3-MYB factors (ZmMYB2, ZmMYB8, ZmMYB31, ZmMYB39, and ZmMYB42) and study their possible roles as regulators of the maize COMT gene. The factors ZmMYB8, ZmMY31, and ZmMYB42 belong to the subgroup 4 of the R2R3-MYB family along with other factors associated with lignin biosynthesis repression. In addition, the induction pattern of ZmMYB31 and ZmMYB42 gene expression on wounding is that expected for repressors of the maize COMT gene. Arabidopsis thaliana plants over-expressing ZmMYB31 and ZmMYB42 down-regulate both the A. thaliana and the maize COMT genes. Furthermore, the over-expression of ZmMYB31 and ZmMYB42 also affect the expression of other genes of the lignin pathway and produces a decrease in lignin content of the transgenic plants.This work was funded by the Spanish “Ministerio de Ciencia y Tecnología” (BIO2001-1140). S-F was financed by a post-doctoral grant from the Autonomous Government of Catalonia (2003PIV-A-00033). F.M-S was financed by the European Project INCO-II (ICA4-CT-2000-30017) and by the “Consorci CSIC-IRTA” laboratory. T-M was financed by the European Project MapMaize (PL 962312). D-C.R was financed by the Spanish “Ministerio de Ciencia y Tecnología” (“Ramón y Cajal” Program). This work was carried out within the framework of the “Centre de Referència de Biotecnologia” (CERBA) from the Autonomous Government of Catalonia.Peer reviewe