| Title | Measuring Gli2 Phosphorylation by Selected Reaction Monitoring Mass Spectrometry. |
| Publication Type | Journal Article |
| Year of Publication | 2015 |
| Authors | Ahrends R, Niewiadomski P, Teruel MN, Rohatgi R |
| Journal | Methods Mol Biol |
| Volume | 1322 |
| Pagination | 105-23 |
| Date Published | 2015 |
| ISSN | 1940-6029 |
| Keywords | Animals, Immunoprecipitation, Kruppel-Like Transcription Factors, Mass Spectrometry, Mice, NIH 3T3 Cells, Peptide Fragments, Phosphorylation, Proteolysis, Zinc Finger Protein Gli2 |
| Abstract | Phosphorylation is an important mechanism by which Gli proteins are regulated. When the Hedgehog (Hh) pathway is activated, multiple serine and threonine residues of Gli2 are dephosphorylated, while at least one residue undergoes phosphorylation. These changes in phosphorylation have functional relevance for the transcriptional activity of Gli proteins, as shown by in vitro and in vivo assays on Gli mutants lacking the phosphorylated residues. Here, we describe a method of quantitatively monitoring the phosphorylation of Gli proteins by triple quadrupole mass spectrometry of Gli2 immunoprecipitated from cell lysates. This method is broadly applicable to the monitoring of phosphorylation changes of immunoprecipitated Gli proteins when the putative phosphosites are known. |
| DOI | 10.1007/978-1-4939-2772-2_10 |
| Custom 1 | |
| Alternate Journal | Methods Mol. Biol. |
| PubMed ID | 26179043 |
| PubMed Central ID | PMC4699800 |
| Grant List | R21NS074091 / NS / NINDS NIH HHS / United States 1R01DK10174301 / DK / NIDDK NIH HHS / United States R01 DK106241 / DK / NIDDK NIH HHS / United States R01 GM112988 / GM / NIGMS NIH HHS / United States R01 DK101743 / DK / NIDDK NIH HHS / United States R01 GM106078 / GM / NIGMS NIH HHS / United States R21 NS074091 / NS / NINDS NIH HHS / United States P50 GM107615 / GM / NIGMS NIH HHS / United States P50GM107615 / GM / NIGMS NIH HHS / United States DP2 GM105448 / GM / NIGMS NIH HHS / United States |