Proteomic analyses reveal divergent ubiquitylation site patterns in murine tissues.
Posttranslational modifications of proteins increase the complexity of the cellular proteome and enable rapid regulation of protein functions in response to environmental changes. Protein ubiquitylation is a central regulatory posttranslational modification that controls numerous biological processes including proteasomal degradation of proteins, DNA damage repair and innate immune responses. Here we combine high-resolution mass spectrometry with single-step immunoenrichment of di-glycine modified peptides for mapping of endogenous putative ubiquitylation sites in murine tissues. We identify more than 20,000 unique ubiquitylation sites on proteins involved in diverse biological processes. Our data reveals that ubiquitylation regulates core signaling pathways common for each of the studied tissues. In addition, we discover that ubiquitylation regulates tissue-specific signaling networks. Many tissue-specific ubiquitylation sites were obtained from brain highlighting the complexity and unique physiology of this organ. We further demonstrate that different di-glycine-lysine-specific monoclonal antibodies exhibit sequence preferences, and that their complementary use increases the depth of ubiquitylation site analysis, thereby providing a more unbiased view of protein ubiquitylation.
Department of Proteomics, The NNF Center for Protein Research, Faculty of Health Sciences, University of Copenhagen, Blegdamsvej 3B, DK-2200 Copenhagen, Denmark.
Sebastian A
SA
Wagner
Petra
P
Beli
Brian T
BT
Weinert
Christian
C
Schölz
Christian D
CD
Kelstrup
Clifford
C
Young
Michael L
ML
Nielsen
Jesper V
JV
Olsen
Cord
C
Brakebusch
Chunaram
C
Choudhary
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D004151:Dipeptides / Q000276:immunology; D006033:Glycylglycine / Q000737:chemistry; D013058:Mass Spectrometry; D051379:Mice; D008810:Mice, Inbred C57BL; D020543:Proteome / Q000378:metabolism*; D040901:Proteomics / Q000379:methods; D015398:Signal Transduction; D025801:Ubiquitin / Q000378:metabolism*; D054875:Ubiquitination
D000911:Antibodies, Monoclonal; D004151:Dipeptides; D020543:Proteome; D025801:Ubiquitin; D006033:Glycylglycine; C035809:glycyllysine