Premium
Proteomic profiling of ubiquitinated proteins in human urinary exosomes (1109.1)
Author(s) -
Huebner Alyssa,
Cheng Lei,
Fenton Robert,
Pisitkun Trairak
Publication year - 2014
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.28.1_supplement.1109.1
Subject(s) - microvesicles , ubiquitin , exosome , microbiology and biotechnology , chemistry , membrane protein , biology , biochemistry , membrane , microrna , gene
Urinary exosomes are small (20‐100 nm) membrane vesicles that contain various molecular constituents of every epithelial cell type facing the urinary space. Based on our previous proteomic studies of urinary exosomes, ubiquitin was identified throughout the molecular weight range of the analysis, suggesting the presence of multiple ubiquitinated proteins in urinary exosomes. Ubiquitination is the classical signal that marks plasma membrane proteins for incorporation into multivesicular bodies where exosomes are originated as internal vesicles. We postulated that ubiquitinated plasma membrane proteins are present in urinary exosomes, potentially providing non‐invasive clues for ubiquitin‐dependent signaling processes in renal epithelial cells. Immunoelectron microscopy revealed ubiquitin labeling on many, mostly inside, urinary exosomes. LC‐MS/MS analysis of urinary exosomes with and without enrichment by di‐glycine‐lysine antibody identified 747 ubiquitinated peptides corresponding to 593 proteins, out of 5007 total proteins. Ubiquitinated proteins were mainly associated with plasma membrane including transporters, channels, and receptors and comprising a number of novel ubiquitinated sites. All but one polyubiquitin chain linkages were identified, with K63‐linked chains being the most frequently observed suggesting non‐proteolytic roles of urinary exosomal ubiquitination. Grant Funding Source : Supported by the Lundbeck Foundation, AU Research Foundation, Danish Medical Research Foundation