TGFbeta regulates the expression of G alpha(i2) via an effect on the localization of ras.
The negative chronotropic response of the heart to parasympathetic stimulation is mediated via the interaction of M(2) muscarinic receptors, Galpha(i2) and the G-protein coupled inward rectifying K(+) channel, GIRK1. Here TGFbeta(1) is shown to decrease the expression of Galpha(i2) in cultured chick atrial cells in parallel with attenuation of the negative chronotropic response to parasympathetic stimulation. The response to the acetylcholine analogue, carbamylcholine, decreased from a 95+/-2% (+/-SEM, n=8) inhibition of beat rate in control cells to 18+/-2% (+/-SEM,n =8) in TGFbeta(1) treated cells. Data support the conclusion that TGFbeta regulation of Galpha(i2) expression was mediated via an effect on Ras. TGFbeta(1) inhibited Galpha(i2) promoter activity by 56+/-6% (+/-SEM, n=4) compared to control. A dominant activating Ras mutant reversed the effect of TGFbeta on Galpha(i2) expression and stimulated Galpha(i2) promoter activity 1.7 fold above control. A dominant negative Ras mutant mimicked the effect of TGFbeta(1) on Galpha(i2) promoter activity. TGFbeta had no effect on the ratio of GDP/GTP bound Ras, but markedly decreased the level of membrane associated Ras and increased the level of cytoplasmic Ras compared to control. Furthermore, farnesol, a precursor to farnesylpyrophosphate, the substrate for the farnesylation of Ras, not only reversed TGFbeta(1) inhibition of Ras localization to the membrane, but also reversed TGFbeta(1) inhibition of Galpha(i2)promoter activity. FTI-277, a specific inhibitor of the farnesylation of Ras, mimicked the effect of TGFbeta(1) on Ras localization and Galpha(i2) promoter activity. These data suggest a novel relationship between TGFbeta signaling, regulation of Ras function and the autonomic response of the heart.
Ward
Simone M
SM
Cardiovascular Division, Department of Medicine, Brigham and Women's Hospital and Harvard Medical School, Boston, Massachusetts 02115, USA.
Gadbut
Albert P
AP
Tang
Dongjiang
D
Papageorge
Alexander G
AG
Wu
Leeying
L
Li
Guangde
G
Barnett
Joey V
JV
Galper
Jonas B
JB
D000818:Animals; D002217:Carbachol / Q000494:pharmacology; D002316:Cardiotonic Agents / Q000494:pharmacology; D002478:Cells, Cultured; D002642:Chick Embryo; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005204:Farnesol / Q000494:pharmacology; D051960:GTP-Binding Protein alpha Subunit, Gi2; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000378:metabolism*; D018507:Gene Expression Regulation, Developmental / Q000187:drug effects*; D006325:Heart Atria / Q000166:cytology / Q000196:embryology / Q000378:metabolism*; D006339:Heart Rate / Q000187:drug effects; D008715:Methionine / Q000031:analogs & derivatives* / Q000494:pharmacology; D032383:Myocytes, Cardiac / Q000187:drug effects; D010275:Parasympathetic Nervous System / Q000187:drug effects; D011401:Promoter Regions, Genetic; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D016212:Transforming Growth Factor beta / Q000494:pharmacology*; D018631:ras Proteins / Q000187:drug effects / Q000378:metabolism*
D002316:Cardiotonic Agents; D004791:Enzyme Inhibitors; C096856:FTI 277; D011518:Proto-Oncogene Proteins; D016212:Transforming Growth Factor beta; D005204:Farnesol; D002217:Carbachol; D008715:Methionine; D051960:GTP-Binding Protein alpha Subunit, Gi2; D019206:GTP-Binding Protein alpha Subunits, Gi-Go; D018631:ras Proteins