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Quantitative Proteomics to Profile Post‐translational Modifications During M Phase: Interplay Between O‐GlcNAcylation and Phosphorylation
Author(s) -
Whelan Stephen A.,
Tan Ee Phie,
Costello Catherine E.,
McComb Mark E.,
Slawson Chad
Publication year - 2013
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.27.1_supplement.555.4
Subject(s) - stable isotope labeling by amino acids in cell culture , phosphorylation , threonine , serine , mitosis , biochemistry , proteomics , protein phosphorylation , posttranslational modification , microbiology and biotechnology , biology , function (biology) , chemistry , enzyme , protein kinase a , gene
During mitosis, proteins undergo a variety of post‐translational modifications (PTM) in order to alter function and localization. Our objective is to determine the interplay between O‐GlcNAcylation and phosphorylation on proteins during mitotic progression. O‐GlcNAc is the attachment of a single N‐acetylglucosamine moiety to serine and threonine residues in nuclear and cytoplasmic proteins. O‐GlcNAc transferase (OGT) adds the modification and O‐GlcNAcase (OGA) removes the modification. O‐GlcNAc is similar to phosphorylation as both modifications are responsive to the extracellular environment, nutrients, and growth hormones. Using SILAC (Stable Isotope Labeling of Amino Acids in Cell culture) labeling, we are able to quantitate differences in cells with either an OGT or OGA gain of function. Combined samples were digested and fractionated to enrich for O‐GlcNAcylated or phosphorylated peptides. We identified numerous proteins whose expression or phosphorylation status was altered by aberrant O‐GlcNAcylation. These results demonstrate the interconnectedness of these two modifications in regulating mitotic progression. Furthermore, these data suggest that diseases that disrupt one PTM will also alter the actions of the other PTM.