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bioRxiv · 10.1101/2023.03.15.532802

NAC1 directs CEP1-CEP3 peptidase expression and decreases cell wall extensins linked to root hair growth in Arabidopsis

Rodriguez-Garcia, D. R.·Marzol, E.·Rondon Guerrero, Y. d. C.·Ferrero, L. V.·Rossi, A. H.·Miglietta, E. A.·Aptekmann, A. A.·Martinez Pacheco, J.·Carignani, M.·Berdion Gabarain, V.·Lopez, L. E.·Diaz Dominguez, G.·Sanchez-Serrano, J. J.·Xu, L.·Nadra, A. D.·Ariel, F. D.·Rojo, E.·Estevez, J.
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Abstract

Plant genomes encode a unique group of papain-type Cysteine EndoPeptidases (CysEPs) containing a KDEL endoplasmic reticulum (ER) retention signal (KDEL-CysEPs or CEPs). CEPs process the cell-wall scaffolding EXTENSIN proteins (EXTs), which regulate de novo cell wall formation and cell expansion. Since CEPs are able to cleave EXTs and EXT-related proteins, acting as cell wall-weakening agents, they may play a role in cell elongation. Arabidopsis thaliana genome encodes three CEPs (AtCPE1-AtCEP3). Here we report that the three Arabidopsis CEPs, AtCEP1-AtCEP3, are highly expressed in root-hair cell files. Single mutants have no evident abnormal root-hair phenotype, but atcep1-3 atcep3-2 and atcep1-3 atcep2-2 double mutants have longer root hairs (RHs) than wild type (Wt) plants, suggesting that expression of AtCEPs in root trichoblasts restrains polar elongation of the RH. We provide evidence that the transcription factor NAC1 activates AtCEPs expression in roots to limit RH growth. Chromatin immunoprecipitation indicates that NAC1 binds the promoter of AtCEP1, AtCEP2, and to a lower extent to AtCEP3 and may directly regulate their expression. Indeed, inducible NAC1 overexpression increases AtCEP1 and AtCEP2 transcript levels in roots and leads to reduced RH growth while the loss of function nac1-2 mutation reduces AtCEP1-AtCEP3 gene expression and enhances RH growth. Likewise, expression of a dominant chimeric NAC1-SRDX repressor construct leads to increased RH length. Finally, we show that RH cell walls in the atcep1-1 atcep3-2 double mutant have reduced levels of EXT deposition, suggesting that the defects in RH elongation are linked to alterations in EXT processing and accumulation. Taken together, our results support the involvement of AtCEPs in controlling RH polar growth through EXT-processing and insolubilization at the cell wall.

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Rodriguez-Garcia, D. R., Marzol, E., Rondon Guerrero, Y. d. C., Ferrero, L. V., Rossi, A. H., Miglietta, E. A., Aptekmann, A. A., Martinez Pacheco, J., Carignani, M., Berdion Gabarain, V., Lopez, L. E., Diaz Dominguez, G., Sanchez-Serrano, J. J., Xu, L., Nadra, A. D., Ariel, F. D., Rojo, E., Estevez, J.. 2023-03-17. NAC1 directs CEP1-CEP3 peptidase expression and decreases cell wall extensins linked to root hair growth in Arabidopsis. https://doi.org/10.1101/2023.03.15.532802

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