SH2-B is a positive regulator of nerve growth factor-mediated activation of the Akt/Forkhead pathway in PC12 cells.
To gain insight into the mechanism by which the adapter protein SH2-B promotes nerve growth factor (NGF)-mediated neuronal differentiation and survival, the effect of SH2-B on the serine/threonine kinase Akt/protein kinase B and downstream effector proteins was examined. PC12 cells stably overexpressing SH2-Bbeta, which exhibit enhanced NGF-induced neuronal differentiation compared with control cells, showed enhanced and prolonged NGF-induced phosphorylation of Akt on Ser473 and Akt enzymatic activity. Surprisingly, NGF-induced phosphorylation of Akt on Ser473 and Akt activity were not altered in cells overexpressing SH2-Bbeta(R555E) with a defective SH2 domain, despite the ability of the overexpressed SH2-Bbeta(R555E) to block NGF-induced differentiation. Consistent with SH2-Bbeta enhancing the activity of Akt, cells overexpressing SH2-Bbeta but not SH2-Bbeta(R555E) exhibited increased and/or prolonged phosphorylation of the pro-apoptotic Akt effector proteins, glycogen synthase kinase-3, and forkhead transcription factors, FKHRL1/FOXO3 and FKHR/FOXO1. Immunolocalization studies indicated that, although ectopically expressed FKHR was primarily concentrated in the cytoplasm of control cells and cells transiently overexpressing SH2-Bbeta, it was concentrated in the nucleus of cells transiently overexpressing SH2-Bbeta(R555E). Similarly, SH2-Bbeta stimulated the accumulation of FKHR in the cytoplasm of 293T and COS-7 cells, whereas SH2-Bbeta(R555E) enhanced its accumulation in the nucleus. In PC12 cells stably expressing forms of SH2-Bbeta, SH2-Bbeta mimicked the ability of NGF to promote redistribution of FKHR to the cytoplasm whereas SH2-Bbeta(R555E) blocked this effect of NGF. Taken together, these data indicate that SH2-B is a positive regulator of NGF-mediated activation of the Akt/Forkhead pathway.
Wang
Xiaohong
X
Department of Molecular and Integrative Physiology, University of Michigan Medical School, Ann Arbor, Michigan 48109-0622, USA.
Chen
Linyi
L
Maures
Travis J
TJ
Herrington
James
J
Carter-Su
Christin
C
D048868:Adaptor Proteins, Signal Transducing; D019943:Amino Acid Substitution; D000818:Animals; D002352:Carrier Proteins / Q000235:genetics / Q000502:physiology*; D002454:Cell Differentiation / Q000187:drug effects; D002460:Cell Line; D017931:DNA Primers; D051858:Forkhead Transcription Factors; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D007700:Kinetics; D016297:Mutagenesis, Site-Directed; D020932:Nerve Growth Factor / Q000494:pharmacology*; D009687:Nuclear Proteins / Q000378:metabolism*; D016716:PC12 Cells; D010766:Phosphorylation; D010768:Phosphoserine / Q000378:metabolism; D020033:Protein Isoforms / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000187:drug effects; D014157:Transcription Factors / Q000378:metabolism*; D018909:src Homology Domains
D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; D017931:DNA Primers; D051858:Forkhead Transcription Factors; D047908:Intracellular Signaling Peptides and Proteins; D009687:Nuclear Proteins; D020033:Protein Isoforms; D011518:Proto-Oncogene Proteins; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; C518910:SH2B1 protein, human; C492010:SH2B1 protein, rat; D014157:Transcription Factors; D010768:Phosphoserine; D020932:Nerve Growth Factor; C494918:AKT1 protein, human; C494920:Akt1 protein, rat; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt