Abstract
Sumoylation is an essential post-translational regulator of plant development and the response to environmental stimuli. SUMO conjugation occurs via an E1-E2-E3 cascade, and can be removed by SUMO proteases (ULPs). ULPs are numerous and likely to function as sources of specificity within the pathway, yet most ULPs remain functionally unresolved. In this report we used loss-of-function reverse genetics and transcriptomics to functionally characterize Arabidopsis thaliana ULP1c and ULP1d SUMO proteases. GUS reporter assays implicated ULP1c/d in various developmental stages, and subsequent defects in growth and germination were uncovered using loss-of-function mutants. Microarray analysis evidenced not only a deregulation of genes involved in development, but also in genes controlled by various drought-associated transcriptional regulators. We demonstrated that ulp1c ulp1d displayed diminished in vitro root growth under low water potential and higher stomatal aperture, yet leaf transpirational water loss and whole drought tolerance were not significantly altered. Generation of a triple siz1 ulp1c ulp1d mutant suggests that ULP1c/d and the SUMO E3 ligase SIZ1 may display separate functions in development yet operate epistatically in response to water deficit. We provide experimental evidence that Arabidopsis ULP1c and ULP1d proteases act redundantly as positive regulators of growth, and operate mainly as isopeptidases downstream of SIZ1 in the control of water deficit responses.
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Acknowledgments
This work was supported by FEDER through the Operational Competitiveness Program—COMPETE—and by national funds through the Foundation for Science and Technology—FCT—within the scope of project “SUMOdulator” [Refs. FCOMP-01-0124-FEDER-028459 and PTDC/BIA-PLA/3850/2012]. PHC was supported by FCT [grant ref. SFRH/BD/44484/2008 and PTDC/BIA-PLA/3850/2012]. HA was supported by the “Genomics and Evolutionary Biology” project, co-financed by North Portugal Regional Operational Programme 2007/2013 (ON.2—O Novo Norte), under the National Strategic Reference Framework (NSRF), through the European Regional Development Fund (ERDF), and by FEDER (COMPETE) and FCT, for Rede de Investigação em Biodiversidade e Biologia Evolutiva [UID/BIA/50027/2013 and POCI-01-0145-FEDER-006821].
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P.H.C. and D.C. performed most of the experiments; S.F., N.V. and A.P.M. performed or helped perform specific experiments; S.H. performed microarray experimentation; H.A., E.R.B., R.M.T., J. R.-A. and M.A.B. supervised the experiments. H.A., P.H.C. and D.C. conceived the project and analyzed the data. H.A. and P.H.C. wrote the article with contributions from all the authors.
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Pedro Humberto Castro and Daniel Couto have contributed equally to this work.
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Castro, P.H., Couto, D., Freitas, S. et al. SUMO proteases ULP1c and ULP1d are required for development and osmotic stress responses in Arabidopsis thaliana . Plant Mol Biol 92, 143–159 (2016). https://doi.org/10.1007/s11103-016-0500-9
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DOI: https://doi.org/10.1007/s11103-016-0500-9