Substitution of two insulin receptor carboxy-terminal tyrosines with phenylalanine impairs the expression of MAP kinase phosphatase-1 (MKP-1) mRNA.
Cells expressing mutant insulin receptors (Y/F2), in which tyrosines 1316 and 1322 have been replaced with phenylalanine, exhibit enhanced insulin-induced MAP kinase activity and DNA synthesis in comparison with cells expressing wild type insulin receptors (Hirc B). To elucidate the mechanism of enhanced responsiveness, the expression of MAP kinase phosphatase-1 (MKP-1), a negative regulator of MAP kinase activity, was measured in Hirc B and Y/F2 cells incubated in the absence and presence of insulin for various periods of time, and over increasing concentrations of the ligand. Treatment of both cell lines with insulin induced a time and concentration-dependent relative increase in MKP-1 mRNA expression. However, in Y/F2 cells both basal and insulin-stimulated MKP-1 mRNA levels were more than 60% lower than that observed in cells transfected with the wildtype receptors. Cyclic AMP analog (8-Br-cAMP)/inducer (Forskoline) increased MKP-1 mRNA levels in both cell lines, and to a lesser extent in Y/F2 cells. In contrast to insulin the relative increase in MKP-1 mRNA expression induced by 8-Br-cAMP or forskoline was similar in Y/F2 and Hirc B cells. The overexpression of MKP-1 in Y/F2 cells inhibited insulin stimulated DNA synthesis. Transfection of wild type insulin receptors into Y/F2 cells increased basal levels of MKP-1. These results suggest that insulin receptor tyrosine residues 13/16 and 1322 play an important role in the regulation of MKP-1 expression both under basal and insulin stimulated conditions, and are not necessary for the induction of MKP-1 mRNA by cAMP. Furthermore, the enhanced insulin induced mitogenic signaling seen in Y/F2 cells is, at least in part, due to impaired MKP-1 expression.
Kusari
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AB
The Department of Physiology, Tulane University School of Medicine, New Orleans, LA, USA.
Byon
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JC
Kusari
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J
D015124:8-Bromo Cyclic Adenosine Monophosphate / Q000494:pharmacology; D019943:Amino Acid Substitution; D000818:Animals; D018797:Cell Cycle Proteins; D002460:Cell Line; D005576:Colforsin / Q000494:pharmacology; D016895:Culture Media, Serum-Free; D004247:DNA / Q000096:biosynthesis; D004305:Dose-Response Relationship, Drug; D054638:Dual Specificity Phosphatase 1; D006801:Humans; D017874:Immediate-Early Proteins / Q000235:genetics* / Q000378:metabolism*; D007328:Insulin / Q000494:pharmacology; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D010749:Phosphoprotein Phosphatases; D054645:Protein Phosphatase 1; D017027:Protein Tyrosine Phosphatases / Q000235:genetics* / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D011972:Receptor, Insulin / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D015398:Signal Transduction; D013936:Thymidine / Q000378:metabolism; D013997:Time Factors; D014162:Transfection
D018797:Cell Cycle Proteins; D016895:Culture Media, Serum-Free; D017874:Immediate-Early Proteins; D007328:Insulin; D012333:RNA, Messenger; D005576:Colforsin; D015124:8-Bromo Cyclic Adenosine Monophosphate; D004247:DNA; D011972:Receptor, Insulin; D020928:Mitogen-Activated Protein Kinases; D010749:Phosphoprotein Phosphatases; D054645:Protein Phosphatase 1; C517019:DUSP1 protein, human; D054638:Dual Specificity Phosphatase 1; C517021:Dusp1 protein, rat; D017027:Protein Tyrosine Phosphatases; D013936:Thymidine