Control of Amino Acid Homeostasis by a Ubiquitin Ligase-Coactivator Protein Complex

Abstract

Edited by James N. SiedowIntercellular amino acid transport is essential for the growth of all multicellular organisms, and its dysregulation is implicated in developmental disorders. By an unknown mechanism, amino acid efflux is stimulated in plants by overexpression of a membrane-localized protein (GLUTAMINE DUMPER 1 (GDU1)) that requires a ubiquitin ligase (LOSS OF GDU 2 (LOG2). Here we further explore the physiological consequences of the interaction between these two proteins. LOG2 ubiquitin ligase activity is necessary for GDU1-dependent tolerance to exogenous amino acids, and LOG2 self-ubiquitination was markedly stimulated by the GDU1 cytosolic domain, suggesting that GDU1 functions as an adaptor or coactivator of amino acid exporter(s). However, other consequences more typical of a ligase-substrate relationship are observed: disruption of the LOG2 gene increased the in vivo half-life of GDU1, mass spectrometry confirmed that LOG2 ubiquitinates GDU1 at cytosolic lysines, and GDU1 protein levels decreased upon coexpression with active, but not enzymatically inactive LOG2. Altogether these data indicate LOG2 negatively regulates GDU1 protein accumulation by a mechanism dependent upon cytosolic GDU1 lysines. Although GDU1-lysine substituted protein exhibited diminished in vivo ubiquitination, overexpression of GDU1 lysine mutants still conferred amino acid tolerance in a LOG2-dependent manner, consistent with GDU1 being both a substrate and facilitator of LOG2 function. From these data, we offer a model in which GDU1 activates LOG2 to stimulate amino acid export, a process that could be negatively regulated by GDU1 ubiquitination and LOG2 self-ubiquitination.

The UBC27–AIRP3 ubiquitination complex modulates ABA signaling by promoting the degradation of ABI1 in Arabidopsis

“…AIRP3 was identified as an interacting protein of UBC27. Previously, AIRP3 was shown to modulate high-salt and drought stress responses through ABA signaling by promoting the degradation of RD21 (37) and to modulate amino acid export by working as the coactivator of GLUTAMINE DUMPER1 (43,44). Here, we demonstrated that AIRP3 also physically interacts with UBC27 and ABI1 (Figs.…”

Identification and biochemical characterization of the fructokinase gene family in Arabidopsis thaliana

“…Agrobacteria strain AGL1 (gift from Charles Gasser, Department of Molecular and Cellular Biology, UC-Davis [ 48 ]) carrying vectors to express C-terminally YFP-tagged FRKs were co-infiltrated with AGL Agrobacteria carrying P19 (gift from Richard Michelmore, Plant Sciences Department, UC-Davis [ 49 ]) into Nicotiana benthamiana leaves. After 48 h, a small leaf disc close to the site of infiltration was excised and then prepared and mounted similarly to what was shown in Littlejohn and Love [ 50 ].…”

Cystathionine γ-lyase promotes estrogen-stimulated uterine artery blood flow via glutathione homeostasis

“…The mechanisms by which CSE promotes vasodilation at the maternal-fetal interface likely involve eNOS. Protein cysteine residues are susceptible to oxidation [ 62 , 63 ], and preeclamptic placentae feature increased eNOS oxidation and decreased NO synthesis [ 15 ]. This suggests that uterine artery CSE deficiency could promote preeclampsia via higher oxidative stress and eNOS inactivation.…”