Ubiquitome profiling reveals metabolism related cell senescence and protein translation modified by UPL3

ABSTRACT: The HECT domain-containing UPL3 ligase plays critical roles in plant development and stress protection, but its mechanism of action remains unclear. To identify its targets in Arabidopsis thaliana, we conducted proteomic analyses of enriched ubiquitinated proteins in upl3 mutants via label-free mass spectrometry. In 6-week-old plants, a landscape of UPL3-dependent ubiquitinated proteins is constructed based on correlated datasets of ubiquitome, proteome, and transcriptome. Preferential ubiquitination of proteins related to carbon fixation and cell senescence represented the largest set of enriched ubiquitinated proteins that occurred in the upl3 plant, while a small set of reduced ubiquitinated proteins caused by the upl3 mutation were linked to cysteine/methionine synthesis and protein translational processes. Furthermore, the upl3 mutation led to an increase in ubiquitination of some of its targets including chromatin remodeling ATPase (BRM), histone proteins, and carbon metabolic enzymes, but decreased the ubiquitination of Hexokinase 1 (HXK1), Phosphoenolpyruvate carboxylase 2 (PPC2), ribosomal protein S5, and H1/H5 domain proteins. Notably, ubiquitin hydrolase 12 (UBP12), BRM, HXK1, and PPC2 were among the UPL3-interacting partners identified by both GFP-nanotrap-Mass-Spectroscopy analyses and yeast two-hybrid assay. Characterization of upl3, and brm-em, ubp12, ubp12ubp13 double mutant plants suggested that UPL3 fine-tuned carbon metabolism during cellular senescence by interacting, in the nucleus, with dually modified targets specific to histone variants, cell senescence, and the carbon metabolic pathway. Our results highlight an important function of UPL3 as a hub of regulated degradation.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Arabidopsis Thaliana (mouse-ear Cress)

TISSUE(S): Plant Cell, Leaf

SUBMITTER: Wei Lan  

LAB HEAD: Ying Miao

PROVIDER: PXD027037 | Pride | 2022-10-14

REPOSITORIES: Pride