Exp Mol Med.  2006 Apr;38(2):134-143.

Downstream molecular events in the altered profiles of lysophosphatidic acid-induced cAMP in senescent human diploid fibroblasts

• Jang IS
• Rhim JH
• Park SC
• Yeo EJ

Abstract

Lysophosphatidic acid (LPA) is a phospholipid growth factor that acts through G-protein-coupled receptors. Previously, we demonstrated an altered profile of LPA-dependent cAMP content during the aging process of human diploid fibroblasts (HDFs). In attempts to define the molecular events associated with the age-dependent changes in cAMP profiles, we determined the protein kinase A (PKA) activity, phosphorylation of cAMP-response element binding protein (CREB), and the protein expression of CRE-regulatory genes, c-fos and COX-2 in young and senescent HDFs. We observed in senescent cells, an increase in mRNA levels of the catalytic subunit a of PKA and of the major regulatory subunit Ia. Senescence-associated increase of cAMP after LPA treatment correlated well with increased CREB phosphorylation accompanying activation of PKA in senescent cells. In senescent cells, after LPA treatment, the expression of c-fos and COX-2 decreased initially, followed by an increase. In young HDFs, CREB phosphorylation decreased following LPA treatment, and both c-fos and COX-2 protein levels increased rapidly. CRE-luciferase assay revealed higher basal CRE-dependent gene expression in young HDFs compared to senescent HDFs. However, LPA-dependent slope of luciferase increased more rapidly in senescent cells than in young cells, presumably due to an increase of LPA-induced CREB phosphorylation. CRE-dependent luciferase activation was abrogated in the presence of inhibitors of PKC, MEK1, p38MAPK, and PKA, in both young and senescent HDFs. We conclude that these kinase are coactivators of the expression of CRE-responsive genes in LPA-induced HDFs and that their changed activities during the aging process contribute to the final expression level of CRE-responsive genes.

Keyword

aging; cyclic AMP response element-binding protein; cyclic AMP-dependent protein kinases; cyclooxygenase 2; lysophosphatidic acid; proto-oncogene proteins c-fos

MeSH Terms

Time Factors
Protein Kinase Inhibitors/pharmacology
Phosphorylation
Male
Lysophospholipids/*pharmacology
Luciferases/genetics/metabolism
Humans
Gene Expression/drug effects
Fibroblasts/cytology/*drug effects/metabolism
Diploidy
Cyclic AMP-Dependent Protein Kinases/genetics/metabolism
Cyclic AMP Response Element-Binding Protein/metabolism
Cyclic AMP/*metabolism
Cells, Cultured
Cell Aging/physiology
Catalytic Domain/genetics