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Am J Physiol. 1997 Oct;273(4 Pt 2):H1920-6.

c-Ha-rasEJ transfection in vascular smooth muscle cells circumvents PKC requirement during mitogenic signaling.

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1
Department of Physiology and Pharmacology, College of Veterinary Medicine, Texas A & M University, College Station 77843-4466, USA.

Abstract

In view of the prominent role of protein kinase C (PKC) in the regulation of vascular smooth muscle cell (VSMC) growth and differentiation, the present studies were conducted to assess the impact of c-Ha-rasEJ transfection on PKC-dependent growth programming. PKC activity was elevated in the cytosolic and particulate compartments of c-Ha-rasEJ VSMC, relative to naive or pSV2neo vector controls. Constitutive and 12-O-tetradecanoyl phorbol 13-acetate (TPA)-inducible binding to a TPA-responsive element (TRE) was also enhanced in c-Ha-rasEJ VSMC. Fetal bovine serum (FBS) did not increase TRE-binding activity in serum-starved c-Ha-rasEJ VSMC but increased TRE-binding activity in pSV2neo VSMC. FBS-mediated TRE-binding activity was dramatically decreased in serum-starved pSV2neo VSMC pretreated with 100 ng/ml TPA for 24 h to downregulate PKC activity. c-Ha-rasEJ VSMC exhibited a marked proliferative advantage over controls under both restrictive and growth-permissive serum conditions. PKC downregulation did not influence the mitogenic response to serum in c-Ha-rasEJ VSMC but ablated [3H]thymidine incorporation into DNA in naive or pSV2neo vector counterparts. Western blot analysis demonstrated increased expression of extracellular signal-regulated kinase 2 (ERK2), but not ERK1, in c-Ha-rasEJ VSMC, relative to pSV2neo control. Immunoblots of serum-starved and PKC-depleted c-Ha-rasEJ VSMC demonstrated a dramatic increase in the phosphorylated form of ERK2, relative to pSV2neo controls. These data suggest that oncogenic c-Ha-rasEJ circumvents a requirement for a TPA-responsive PKC isoform(s) during mitogenic stimulation of VSMC.

PMID:
9362261
[Indexed for MEDLINE]
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