Research Resource: Identification of Novel Growth Hormone-Regulated Phosphorylation Sites by Quantitative Phosphoproteomics

Ray, Bridgette N.; Kweon, Hye Kyong; Argetsinger, Lawrence S.; Fingar, Diane C.; Andrews, Philip C.; Carter-Su, Christin

Of the 132 unique localized phosphosites showing GH-dependent increases in phosphorylation, only six were previously documented as GH responsive. These were the activating Tyr in the transcription factors Stat5a (Tyr694) 1 and Stat5b (Tyr699) (only detected in GHstimulated cells); the activating Thr and Tyr in the serine/ threonine kinase Erk2 (Thr183/Tyr185) (ϩGH/ϪGH ϭ 10.9); the activating Tyr in Erk1 (Tyr205) (ϩGH/ϪGH ϭ 2.4); and the regulatory Ser939 in tuberous sclerosis protein 2 or tuberin (TSC2) (ϩGH/ϪGH ϭ 1.9). TSC2 is a GTPase-activating protein, which inhibits Ras homolog enriched in brain (Rheb), and thereby inhibits mammalian target of rapamycin (mTOR) complex 1 (mTORC1) ( Fig. 3 ; reviewed in Ref. 48 ). mTORC1 activity is required for rapid GH-stimulated protein synthesis (14) . Phosphorylation of TSC2 Ser939 suppresses TSC2 function, resulting in increased mTORC1 signaling (49, 50) . The remaining 126 phosphosites have not been identified previously as GH-regulated sites of phosphorylation. Of the newly identified GH-regulated sites, only 13 have been identified in the context of other stimuli and their function analyzed. The function and mechanism of phosphorylation is unknown for the remaining 113 sites.