title: Insulin-induced egr-1 expression in Chinese hamster ovary cells is insulin receptor and insulin receptor substrate-1 phosphorylation-independent. Evidence of an alternative signal transduction pathway.
abstract: Insulin's effects primarily are initiated by insulin binding to its plasma membrane receptor and the sequential tyrosine phosphorylation of the insulin receptor and intracellular substrates, such as insulin receptor substrate-1 (IRS-1). However, studies suggest some insulin effects, including those at the nucleus, may not be regulated by this pathway. The present study compared the levels of insulin binding, insulin receptor and IRS-1 tyrosine phosphorylation, and phosphatidylinositol 3'-kinase activity to immediate early gene c-fos and egr-1 mRNA expression in Chinese hamster ovary (CHO) cells expressing only neomycin-resistant plasmid (CHONEO), overexpressing wild type human insulin receptor (CHOHIRc) or ATP binding site-mutated insulin receptors (CHOA1018K). Insulin binding in CHONEO cells was markedly lower than that in other cell types. 10 nM insulin significantly increased tyrosine phosphorylation of insulin receptor and IRS-1 in CHOHIRc cells. Phosphorylation of insulin receptor and IRS-1 in CHONEO and CHOA1018K cells was not detected in the presence or absence of insulin. Similarly, insulin increased phosphatidylinositol 3-kinase activity only in CHOHIRc cells. As determined by Northern blot, nuclear run-on analysis, and in situ hybridization, insulin induced c-fos mRNA expression, through transcription, in CHOHIRc cells but not in CHONEO and CHOA1018K cells, consistent with previous reports. In contrast, all three cell types showed a similar insulin dose-dependent increase of egr-1 mRNA expression through transcription. These data indicated that insulin-induced egr-1 mRNA expression did not correlate with the levels of insulin binding to insulin receptor or phosphorylation of insulin receptor and IRS-1. These results suggest that different mechanisms are involved in induction of c-fos and egr-1 mRNA expression by insulin, the former by the more classic insulin receptor tyrosine kinase pathway and the latter by a yet to be determined alternative signal transduction pathway.
authors.affiliation: Department of Pathology and Laboratory Medicine, University of Pennsylvania School of Medicine, Philadelphia 19104, USA.
authors.forename: S
authors.initials: S
authors.lastname: Harada
authors.forename: R M
authors.initials: RM
authors.lastname: Smith
authors.forename: J A
authors.initials: JA
authors.lastname: Smith
authors.forename: N
authors.initials: N
authors.lastname: Shah
authors.forename: D Q
authors.initials: DQ
authors.lastname: Hu
authors.forename: L
authors.initials: L
authors.lastname: Jarett
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001665:Binding Sites; D016466:CHO Cells; D002467:Cell Nucleus / Q000378:metabolism / Q000648:ultrastructure; D006224:Cricetinae; D003412:Cricetulus; D004268:DNA-Binding Proteins / Q000096:biosynthesis* / Q000235:genetics; D051766:Early Growth Response Protein 1; D015870:Gene Expression / Q000187:drug effects*; D017781:Genes, Immediate-Early; D006801:Humans; D017874:Immediate-Early Proteins; D017403:In Situ Hybridization; D007328:Insulin / Q000378:metabolism / Q000494:pharmacology*; D053590:Interleukin 1 Receptor Antagonist Protein; D007700:Kinetics; D008854:Microscopy, Electron; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation; D012333:RNA, Messenger / Q000096:biosynthesis; D011972:Receptor, Insulin / Q000378:metabolism*; D012795:Sialoglycoproteins / Q000378:metabolism*; D015398:Signal Transduction; D014157:Transcription Factors / Q000096:biosynthesis* / Q000235:genetics; D014158:Transcription, Genetic; D016335:Zinc Fingers
chemical_list: D004268:DNA-Binding Proteins; C497325:EGR1 protein, human; D051766:Early Growth Response Protein 1; C507763:IL1RN protein, human; D017874:Immediate-Early Proteins; D007328:Insulin; D053590:Interleukin 1 Receptor Antagonist Protein; D012333:RNA, Messenger; D012795:Sialoglycoproteins; D014157:Transcription Factors; D000255:Adenosine Triphosphate; D011972:Receptor, Insulin