Epidermal Growth Factor Receptor–Mediated Proliferation of Enterocytes Requires p21waf1/cip1 Expression

Department of Surgery, University of Cincinnati College of Medicine, Ohio, USA.
Gastroenterology (Impact Factor: 16.72). 07/2006; 131(1):153-64. DOI: 10.1053/j.gastro.2006.05.007
Source: PubMed


Epidermal growth factor receptor (EGFR)-mediated increase in enterocyte proliferation following massive resection is a major mechanism by which the small intestine adapts to the loss of its mucosal surface area. In addition, expression of the cyclin-dependent kinase inhibitor p21(waf1/cip1) is required for resection-induced enterocyte proliferation. This study sought to establish a mechanistic link between EGFR-mediated intestinal epithelial cell proliferation and p21(waf1/cip1) expression.
EGF was used to stimulate IEC-6 and HCA-7 cells. P21(waf1/cip1) messenger RNA (mRNA) and protein expression were measured by real-time polymerase chain reaction and Western blot, respectively. P21(waf1/cip1) promoter studies were performed using p21(waf1/cip1) promoter-driven luciferase assay. Pharmacologic inhibitors of PI3-kinase and mitogen activated protein kinase (MAPK) were used to block these pathways downstream of the activated EGFR. Constitutively active Ras, Raf, or MEK-1 constructs were transfected into cells for overexpression studies. Cell proliferation was measured by bromodeoxyuridine incorporation following p21(waf1/cip1) silencing with RNAi. Finally, Cyclin D(1)/Cdk interaction was evaluated by immunoprecipitation.
EGFR activation in intestinal epithelial cells induced the expression of p21(waf1/cip1) mRNA and protein This event was transcriptionally regulated via a 50-bp segment of the p21(waf1/cip1) promoter as a result of MAPK activation. Exogenous EGF failed to induce proliferation in p21(waf1/cip1)-silenced cells and adaptive proliferation after intestinal resection in p21(waf1/cip1)-null mice. Functionally, p21(waf1/cip1) up-regulation was required for stabilizing Cyclin D/Cdk 4 complexes and intestinal cell proliferation.
EGFR-mediated induction of enterocyte proliferation requires MAPK-dependent increase in p21(waf1/cip1) expression in intestinal epithelial cells. These studies elucidate an important mechanism for resection-induced enterocyte proliferation during intestinal adaptation.

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