Proximal proteome and ubiquitination landscape of E3 Ubiquitin Ligase Mindbomb 1

ABSTRACT: To characterize the proximal proteome of Mindbomb 1 (MIB1) during adenovirus (AdV) infection ?we performed proximity labeling. MIB1 KO cell lines generated in the Hap1 cell line were stably reconstituted with either WT MIB1-APEX2, the MIB1 ubiquitination-defective mutant C985S-APEX2 or APEX2-mKate. Cell lines were infected with ?replication-competent (rep-comp) AdV5-GFP followed by pulse-labeling with H202 at 1.25 ?hours post infection (hpi), streptavidin isolation of biotinylated proteins and LC-MS/MS analysis of the proximal proteomes. To determine possible u?biquit?ination targets of MIB1 during AdV infection, Tandem-Ubiquitin-Binding-Entities (TUBEs) were used to isolate endogenous ubiquitinated proteins. MIB1 KO Hap1 cells, and KO cells reconstituted with either WT MIB1 or the ubiquitination-dead mutant C985S MIB1 were infected with rep-comp AdV5-GFP and collected at 1 hpi. Lysates were incubated with TUBE Dynabeads, and the isolated proteins were submitted for LC-MS/M

INSTRUMENT(S): Q Exactive HF

ORGANISM(S): Homo Sapiens (human)

SUBMITTER: Soren Heissel  

LAB HEAD: Charles M. Rice

PROVIDER: PXD020542 | Pride | 2021-09-09

REPOSITORIES: Pride