Regulation of gap junction coupling through the neuronal connexin Cx35 by nitric oxide and cGMP.
Gap-junctional coupling among neurons is subject to regulation by a number of neurotransmitters including nitric oxide. We studied the mechanisms by which NO regulates coupling in cells expressing Cx35, a connexin expressed in neurons throughout the central nervous system. NO donors caused potent uncoupling of HeLa cells stably transfected with Cx35. This effect was mimicked by Bay 21-4272, an activator of guanylyl cyclase. A pharmacological analysis indicated that NO-induced uncoupling involved both PKG-dependent and PKG-independent pathways. PKA was involved in both pathways, suggesting that PKG-dependent uncoupling may be indirect. In vitro, PKG phosphorylated Cx35 at three sites: Ser110, Ser276, and Ser289. A mutational analysis indicated that phosphorylation on Ser110 and Ser276, sites previously shown also to be phosphorylated by PKA, had a significant influence on regulation. Ser289 phosphorylation had very limited effects. We conclude that NO can regulate coupling through Cx35 and that regulation is indirect in HeLa cells.
Patel
Leena S
LS
Department of Ophthalmology and Visual Science, University of Texas Health Science Center, Houston, 77030, USA.
Mitchell
Cheryl K
CK
Dubinsky
William P
WP
O'Brien
John
J
D002450:Cell Communication; D017630:Connexins / Q000378:metabolism*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D006152:Cyclic GMP / Q000378:metabolism* / Q000494:pharmacology; D017869:Cyclic GMP-Dependent Protein Kinases / Q000378:metabolism*; D005136:Eye Proteins / Q000378:metabolism*; D017629:Gap Junctions / Q000187:drug effects / Q000378:metabolism*; D006162:Guanylate Cyclase / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D008954:Models, Biological; D009474:Neurons / Q000378:metabolism; D009569:Nitric Oxide / Q000378:metabolism* / Q000494:pharmacology; D018408:Patch-Clamp Techniques; D010766:Phosphorylation; D012694:Serine / Q000378:metabolism
D017630:Connexins; D005136:Eye Proteins; C100585:connexin 35 protein, vertebrate; D009569:Nitric Oxide; D012694:Serine; D017868:Cyclic AMP-Dependent Protein Kinases; D017869:Cyclic GMP-Dependent Protein Kinases; D006162:Guanylate Cyclase; D006152:Cyclic GMP