Ubiquitome profiling reveals a regulatory pattern of UPL3 with UBP12 on metabolic-leaf senescence

Abstract

The HECT-type UPL3 ligase plays critical roles in plant development and stress protection, but understanding of its regulation remains limited. Here, the multi-omics analyses of ubiquitinated proteins in upl3 mutants were performed. A landscape of UPL3- dependent ubiquitinated proteins is constructed: Preferential ubiquitination of proteins related to carbon fixation represented the largest set of proteins with increased ubiquitination in the upl3 plant, including most of carbohydrate metabolic enzymes, BRM, and variant histone, whereas a small set of proteins with reduced ubiquitination caused by the upl3 mutation were linked to cysteine/methionine synthesis, as well as hexokinase 1 (HXK1) and phosphoenolpyruvate carboxylase 2 (PPC2). Notably, ubiquitin hydrolase 12 (UBP12), BRM, HXK1, and PPC2 were identified as the UPL3-interacting partners in vivo and in vitro. Characterization of brm, upl3, ppc2, gin2, and ubp12 mutant plants and proteomic and transcriptomic analysis suggested that UPL3 fine-tunes carbohydrate metabolism, mediating cellular senescence by interacting with UBP12, BRM, HXK1, and PPC2. Our results highlight a regulatory pattern of UPL3 with UBP12 as a hub of regulator on proteolysis-independent regulation and proteolysisdependent degradation.