UBP12 and UBP13 negatively regulate the activity of the ubiquitin-dependent peptidases DA1, DAR1 and DAR2.

Hannes Vanhaeren,Valerie De Vleeschhauwer,Liesbeth De Milde,Dirk Inzé,Kris Gevaert,Annelore Natran,Geert De Jaeger,Mattias Vermeersch,Dominique Eeckhout,Ying Chen,Geert Persiau

doi:10.7554/elife.52276

Hannes Vanhaeren, Valerie De Vleeschhauwer + Show 9 more

Open Access

https://doi.org/10.7554/elife.52276

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Abstract

Protein ubiquitination is a very diverse post-translational modification leading to protein degradation or delocalization, or altering protein activity. In Arabidopsis thaliana, two E3 ligases, BIG BROTHER (BB) and DA2, activate the latent peptidases DA1, DAR1 and DAR2 by mono-ubiquitination at multiple sites. Subsequently, these activated peptidases destabilize various positive growth regulators. Here, we show that two ubiquitin-specific proteases, UBP12 and UBP13, deubiquitinate DA1, DAR1 and DAR2, hence reducing their peptidase activity. Overexpression of UBP12 or UBP13 strongly decreased leaf size and cell area, and resulted in lower ploidy levels. Mutants in which UBP12 and UBP13 were downregulated produced smaller leaves that contained fewer and smaller cells. Remarkably, neither UBP12 nor UBP13 were found to be cleavage substrates of the activated DA1. Our results therefore suggest that UBP12 and UBP13 work upstream of DA1, DAR1 and DAR2 to restrict their protease activity and hence fine-tune plant growth and development.

Highlights

Ubiquitination plays a prominent role in the signaling cascades of many plant hormones (Santner and Estelle, 2010), such as auxins (Salehin et al, 2015), jasmonates (Nagels Durand et al, 2016), gibberellins (Wang and Deng, 2011), and strigolactones (Marzec, 2016), and in many plant developmental processes and responses to stress (Shu and Yang, 2017)
We identified two ubiquitin-specific proteases, UBIQUITIN-SPECIFIC PROTEASE 12 (UBP12) an UBP13, that interact with DA1, DAR1 and DAR2 in vitro and in vivo
Our experiments demonstrate that these ubiquitin binding proteins (UBPs) work antagonistically to BIG BROTHER (BB) and DA2

Summary

Introduction

Ubiquitination plays a prominent role in the signaling cascades of many plant hormones (Santner and Estelle, 2010), such as auxins (Salehin et al, 2015), jasmonates (Nagels Durand et al, 2016), gibberellins (Wang and Deng, 2011), and strigolactones (Marzec, 2016), and in many plant developmental processes and responses to stress (Shu and Yang, 2017). UBP12 and UBP13 are the largest UBPs and contain a unique meprin and TRAF homology (MATH) domain They were first reported to be functional deubiquitinating enzymes that negatively regulate plant immunity (Ewan et al, 2011). Plant growth is strongly enhanced in the double mutant da1ko_dar, comparable to da mutants, which carry a point mutation (DA1R358K) (Li et al, 2008) The latter mutation has a dominant-negative action towards DA1 and DAR1 (Li et al, 2008) and causes a reduction in peptidase activity (Dong et al, 2017). Our data provide evidence for a pivotal role of UBP12 and UBP13 in restricting the protease activity of DA1, DAR1 and DAR2 during plant growth and development

Results

25 IP: α-GST IB: α-GST

Discussion

A Normal levels of UBP12 and UBP13

Materials and methods

Funding Funder Bijzonder Onderzoeksfonds