GTPase activating specificity of RGS12 and binding specificity of an alternatively spliced PDZ (PSD-95/Dlg/ZO-1) domain.
Regulator of G-protein signaling (RGS) proteins increase the intrinsic guanosine triphosphatase (GTPase) activity of G-protein alpha subunits in vitro, but how specific G-protein-coupled receptor systems are targeted for down-regulation by RGS proteins remains uncharacterized. Here, we describe the GTPase specificity of RGS12 and identify four alternatively spliced forms of human RGS12 mRNA. Two RGS12 isoforms of 6.3 and 5.7 kilobases (kb), encoding both an N-terminal PDZ (PSD-95/Dlg/ZO-1) domain and the RGS domain, are expressed in most tissues, with highest levels observed in testis, ovary, spleen, cerebellum, and caudate nucleus. The 5.7-kb isoform has an alternative 3' end encoding a putative C-terminal PDZ domain docking site. Two smaller isoforms, of 3.1 and 3.7 kb, which lack the PDZ domain and encode the RGS domain with and without the alternative 3' end, respectively, are most abundantly expressed in brain, kidney, thymus, and prostate. In vitro biochemical assays indicate that RGS12 is a GTPase-activating protein for Gi class alpha subunits. Biochemical and interaction trap experiments suggest that the RGS12 N terminus acts as a classical PDZ domain, binding selectively to C-terminal (A/S)-T-X-(L/V) motifs as found within both the interleukin-8 receptor B (CXCR2) and the alternative 3' exon form of RGS12. The presence of an alternatively spliced PDZ domain within RGS12 suggests a mechanism by which RGS proteins may target specific G-protein-coupled receptor systems for desensitization.
Snow
B E
BE
Amgen Institute, Toronto, Ontario M5G 2C1, Canada.
Hall
R A
RA
Krumins
A M
AM
Brothers
G M
GM
Bouchard
D
D
Brothers
C A
CA
Chung
S
S
Mangion
J
J
Gilman
A G
AG
Lefkowitz
R J
RJ
Siderovski
D P
DP
D048868:Adaptor Proteins, Signal Transducing; D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D017931:DNA Primers; D000074288:Discs Large Homolog 1 Protein; D000076122:Disks Large Homolog 4 Protein; D004789:Enzyme Activation; D020558:GTP Phosphohydrolases / Q000378:metabolism*; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins / Q000235:genetics*; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000235:genetics*; D010750:Phosphoproteins / Q000235:genetics*; D011506:Proteins / Q000235:genetics* / Q000378:metabolism*; D020710:RGS Proteins; D051381:Rats; D012441:Saccharomyces cerevisiae / Q000235:genetics; D013379:Substrate Specificity; D062826:Zonula Occludens-1 Protein
D048868:Adaptor Proteins, Signal Transducing; C513314:DLG1 protein, human; D017931:DNA Primers; D000074288:Discs Large Homolog 1 Protein; D000076122:Disks Large Homolog 4 Protein; C501119:Dlg1 protein, rat; C501449:Dlg4 protein, rat; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D010750:Phosphoproteins; D011506:Proteins; D020710:RGS Proteins; C106375:RGS12 protein, human; C491401:Rgs12 protein, rat; C569042:TJP1 protein, human; C569044:Tjp1 protein, rat; D062826:Zonula Occludens-1 Protein; C033652:postsynaptic density proteins; D020558:GTP Phosphohydrolases