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

The KEGG pathways "regulation of actin cytoskeleton" and "focal adhesions" were also identified during the bioinformatics analysis. Previous studies have implicated GH in regulation of the actin cytoskeleton. GH has been shown to promote membrane ruffling of 3T3-F442A cells (54) and CHO cells expressing GH receptors (53) . It has also been shown to increase the motility of 293T cells stably expressing GH receptor (56) and the directed migration of monocytes (52, 55) and RAW264.7 macrophages (Su, H.-W., and C. Carter-Su, manuscript in preparation). During cell migration, focal complexes assemble at the leading edge and then disassemble as a cell moves. Some focal complexes mature and recruit new proteins; these focal adhesions anchor cells to the matrix and are important for cell movement. Multiple focal adhesion proteins have been shown to undergo GHdependent changes in tyrosyl phosphorylation (18, 81, 82) . Of the seven proteins containing GH-enhanced phosphosites identified by KEGG analysis as "focal adhesion" proteins, four (filamin, zyxin, parvin, Ppp1r12a/regulatory subunit of myosin light chain phosphatase) had not been identified previously as GHregulated proteins. Among the five GH-up-regulated phosphoproteins in the KEGG category "regulation of the actin cytoskeleton," three (NHE1, myosin heavy chain 9, and Ppp1r12a) had not been identified previously as GH-regulated phosphoproteins.