Role of protein kinase C-dependent A-kinase anchoring proteins in lysophosphatidic acid-induced cAMP signaling in human diploid fibroblasts.
Previously, we reported that lysophosphatidic acid (LPA)-induced adenosine 3',5'-cyclic monophosphate (cAMP) production by human diploid fibroblasts depends on the age of the fibroblasts. In this study, we examined the role of A-kinase anchoring proteins (AKAP) in the regulation of LPA-stimulated cAMP production in senescent fibroblasts. We found that levels of protein kinase C (PKC)-dependent AKAPs, such as Gravin and AKAP79, were elevated in senescent cells. Co-immunoprecipitation experiments revealed that Gravin and AKAP79 do not associate with adenylyl cyclase type 2 (AC2) but bind to AC4/6, which interacts with calcium-dependent PKCs alpha/beta both in young and senescent fibroblasts. When the expression of Gravin and AKAP79 was blocked by small interference RNA transfection, the basal level of cAMP was greatly reduced and the cAMP status after LPA treatment was also reversed. Protein kinase A showed a similar pattern in terms of its basal activity and LPA-dependent modulation. These data suggest that Gravin and to a lesser extent, AKAP79, may play important roles in maintaining the basal AC activity and in coupling the AC systems to inhibitory signals such as Gialpha in young cells, and to stimulatory signals such as PKCs in senescent cells. This study also demonstrates that Gravin is especially important for the long-term activation of PKC by LPA in senescent cells. We conclude that LPA-dependent increased level of cAMP in senescent human diploid fibroblasts is associated with increases in Gravin levels resulting in its increased binding with and activation of calcium-dependent PKC alpha/beta and AC4/6.
Rhim
Ji-Heon
JH
Department of Biochemistry and Molecular Biology, Aging and Apoptosis Research Center, Seoul National University College of Medicine, Seoul 110-799, South Korea.
Jang
Ik-Soon
IS
Yeo
Eui-Ju
EJ
Song
Kye-Yong
KY
Park
Sang Chul
SC
D054758:A Kinase Anchor Proteins; D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism*; D000262:Adenylyl Cyclases / Q000378:metabolism; D000818:Animals; D018797:Cell Cycle Proteins / Q000378:metabolism*; D002478:Cells, Cultured; D016922:Cellular Senescence / Q000187:drug effects / Q000502:physiology*; D000242:Cyclic AMP / Q000096:biosynthesis*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D004171:Diploidy; D015536:Down-Regulation / Q000235:genetics; D005347:Fibroblasts / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism; D006801:Humans; D007231:Infant, Newborn; D008246:Lysophospholipids / Q000378:metabolism / Q000494:pharmacology; D008297:Male; D011485:Protein Binding / Q000502:physiology; D051571:Protein Kinase C-alpha / Q000378:metabolism; D034622:RNA Interference; D051381:Rats; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D015854:Up-Regulation / Q000187:drug effects / Q000502:physiology
D054758:A Kinase Anchor Proteins; C491162:AKAP12 protein, human; C487775:AKAP5 protein, human; D048868:Adaptor Proteins, Signal Transducing; D018797:Cell Cycle Proteins; D008246:Lysophospholipids; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D051571:Protein Kinase C-alpha; D000262:Adenylyl Cyclases; C406262:adenylyl cyclase 4; C032881:lysophosphatidic acid