Catalytic activation of extracellular signal-regulated kinases induces cyclin D1 expression in primary tracheal myocytes.
We have demonstrated that extracellular signal-regulated kinases (ERKs) and cyclin D1 are required for bovine tracheal myocyte DNA synthesis. We hypothesized that catalytic activation by ERKs may regulate cyclin D1 expression in these cells. To test this hypothesis, we examined the effects of two inhibitors of ERKs and two reagents that increase the level of activated ERKs on cyclin D1 protein abundance and promoter activity. ERK activity was inhibited either by PD98059, a synthetic inhibitor of mitogen-activated protein kinase (MAPK)/ERK kinase (MEK), the upstream signaling intermediate required and sufficient for ERK activation, or by transient transfection with a dominant-negative mutant of MEK1 (MEK-2A). The level of activated ERKs was increased by transient transfection with either a constitutively active form of MEK1 (MEK-2E) or wild-type ERK2 (MAPKwt). Cyclin D1 expression was assessed either by immunoblot or cotransfection with the full-length cyclin D1 promoter subcloned into a luciferase reporter. We found that pretreatment of bovine tracheal myocytes with PD98059 significantly attenuated platelet- derived growth factor (PDGF)-induced cyclin D1 protein abundance. Furthermore, transfection with MEK-2A reduced PDGF-induced cyclin D1 promoter activity. Finally, transfection with either MEK-2E or MAPKwt induced cyclin D1 promoter activity in the absence of growth factor treatment. We conclude that catalytic activation of ERKs regulates cyclin D1 expression in airway smooth-muscle cells.
Ramakrishnan
M
M
Department of Pediatrics, University of Chicago, Chicago, Illinois 60637-1470, USA.
Musa
N L
NL
Li
J
J
Liu
P T
PT
Pestell
R G
RG
Hershenson
M B
MB
D000818:Animals; D015153:Blotting, Western; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism* / Q000502:physiology; D002417:Cattle; D002478:Cells, Cultured; D019938:Cyclin D1 / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000378:metabolism; D005419:Flavonoids / Q000494:pharmacology; D015870:Gene Expression; D048369:MAP Kinase Kinase 1; D019950:Mitogen-Activated Protein Kinase 1; D020929:Mitogen-Activated Protein Kinase Kinases; D009130:Muscle, Smooth / Q000187:drug effects / Q000378:metabolism*; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism / Q000502:physiology; D011505:Protein-Tyrosine Kinases / Q000378:metabolism / Q000502:physiology; D014132:Trachea / Q000187:drug effects / Q000378:metabolism*; D015533:Transcriptional Activation; D014162:Transfection
D004791:Enzyme Inhibitors; D005419:Flavonoids; D010982:Platelet-Derived Growth Factor; D019938:Cyclin D1; D011505:Protein-Tyrosine Kinases; D017346:Protein Serine-Threonine Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048369:MAP Kinase Kinase 1; D020929:Mitogen-Activated Protein Kinase Kinases; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one