Phosphorylation of the recombinant rho1 GABA receptor.
Gamma-aminobutyric acid (GABA) is the major inhibitory neurotransmitter in the mammalian brain. While a growing body of literature indicates that postsynaptic GABA receptors are regulated by phosphorylation, there is discrepancy as to the specific effects of phosphorylation on GABA receptor function. Here, we have identified phosphorylation sites on the human rho1 GABA receptor for six protein kinases widely expressed in the brain: protein kinase C (PKC); cAMP-dependent protein kinase (PKA); calmodulin-dependent kinase (CaMKII); casein kinase (CKII); mitogen-activated protein kinase (MAPK); and cGMP-dependent protein kinase (PKG). We demonstrate that in nearly all cases, the consensus sites and actual phosphorylation sites do not agree supporting the risk of relying on a sequence analysis to identify potential phosphorylation sites. In addition, of the six kinases examined, only CKII phosphorylated the human rho2 subunit. Site-directed mutagenesis of the phosphorylation sites, or activation/inhibition of select kinase pathways, did not alter the receptor sensitivity or maximal GABA-activated current of the rho1 GABA receptor expressed in Xenopus laevis oocytes suggesting phosphorylation of rho1 does not directly alter receptor properties. This study is a first and necessary step towards elucidating the regulation of rho1 GABA receptors by phosphorylation.
Sedelnikova
Anna
A
Department of Neurobiology, University of Alabama at Birmingham, Birmingham, AL 35294-0021, USA.
Weiss
David S
DS
D000818:Animals; D001665:Binding Sites / Q000502:physiology; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism; D047388:Casein Kinases; D002490:Central Nervous System / Q000201:enzymology*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D017869:Cyclic GMP-Dependent Protein Kinases / Q000378:metabolism; D005260:Female; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D009865:Oocytes; D010766:Phosphorylation; D010770:Phosphotransferases / Q000378:metabolism*; D011493:Protein Kinase C / Q000378:metabolism; D011494:Protein Kinases / Q000378:metabolism; D017434:Protein Structure, Tertiary / Q000502:physiology; D018080:Receptors, GABA-B / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D014982:Xenopus laevis; D005680:gamma-Aminobutyric Acid / Q000378:metabolism*
D018080:Receptors, GABA-B; D011993:Recombinant Fusion Proteins; D005680:gamma-Aminobutyric Acid; D010770:Phosphotransferases; D011494:Protein Kinases; D047388:Casein Kinases; D017868:Cyclic AMP-Dependent Protein Kinases; D017869:Cyclic GMP-Dependent Protein Kinases; D011493:Protein Kinase C; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D019950:Mitogen-Activated Protein Kinase 1