Project description:We performed diGLY enrichment followed by label-free proteomics in C. elegans to define and quantify changes in the ubiquitinated (Ub)-proteome during the aging process. More specifically, we compared wild-type worms at the first day of adulthood with young (day 5), mid-age (day 10) and aged adults (day 15). Moreover, we assessed the Ub-modified proteome of age-matched long-lived genetic models of dietary restriction (eat-2(ad1116)) and reduced reduced insulin/IGF-1 signaling (daf-2(e1370)).
Project description:We performed label-free proteomics in C. elegans to define and quantify changes in the ubiquitinated (Ub)-proteome during the aging process. More specifically, we compared wild-type worms at the first day of adulthood with young (day 5), mid-age (day 10) and aged adults (day 15). Moreover, we assessed protein levels of age-matched long-lived genetic models of dietary restriction (eat-2(ad1116)) and reduced insulin/IGF-1 signaling (daf-2(e1370)).
Project description:We analyzed genome-wide modification of chromatin by ubiquitination in human cells and whether this mark changes through the cell cycle using modified ChIP-seq technique (ChAP-seq). Chromatin affinity purification was based on a standard ChIP method with modification of a two-step affinity purification.
Project description:This project was aimed to identify lipid droplet (LD) membrane proteins that are ubiquitylated. We isolated LDs from the livers of control and alcohol-fed rats and extracted LD membrane proteins.
Project description:We performed immunoprecipitation experiments in young worms using an antibody against Lys48-linked polyUb followed by single shot label-free proteomics.
Project description:We performed immunoprecipitation experiments in young worms using an antibody against Lys63-linked polyUb followed by single shot label-free proteomics.
Project description:DUSP22 (also named JKAP) is a dual-specificity phosphatase that inhibits T cell activation. Here we identified the E3 ubiquitin ligase UBR2 as an upstream activator of Lck during T-cell activation. JKAP dephosphorylated UBR2 at two residues, leading to ubiquitin-mediated UBR2 degradation. The SCF (SKP1-CUL1-βTrCP) complex induced UBR2 Lys48-linked ubiquitination at three lysine residues. Moreover, single-cell RNA sequencing analysis and UBR2 knockout showed that UBR2 increased proinflammatory cytokines. Remarkably, UBR2 induced Lys63-linked ubiquitination of Lck at two lysine residues and subsequent Lck Tyr394phosphorylation/activation in TCR signaling. Conversely, TCR-induced Lck activation and JKAP knockout-enhanced inflammatory phenotypes were attenuated by UBR2 knockout. Consistently, the UBR2-Lck interaction and Lck Lys63-linked ubiquitination were induced in peripheral blood T cells of human SLE patients. Collectively, UBR2 protein stability and UBR2-induced Lck ubiquitination/activation are inhibited by JKAP, leading to attenuation of T-cell activation and T-cell-mediated inflammation.