Human substance P receptor expressed in Chinese hamster ovary cells directly activates G(alpha q/11), G(alpha s), G(alpha o).
Substance P receptor (SPR) stably expressed in Chinese hamster ovary (CHO) cells stimulates at least three second messenger systems including phosphoinositide hydrolysis, cyclic AMP (cAMP) formation, and arachidonic acid release. Whether these second messenger systems are activated via single or multiple G proteins is not known. Therefore, in the present study we examined whether human SPR (hSPR) stably expressed in CHO cells activates multiple G proteins. This was achieved by photoaffinity labeling of G(alpha)-subunits with [32P]azidoanilido-GTP ([32P]AA-GTP) upon hSPR stimulation in CHO-hSPR membranes followed by immunoprecipitation of the labeled G(alpha)-subunits with antibodies specific for various G(alpha)-subunits. These experiments reveal that hSPR directly activates G(alpha q/11), G(alpha s) and G(alpha o). While hSPR is known to couple G(alpha q/11), the present study provides the first evidence that hSPR can also activate G(alpha s) and G(alpha o) in a mammalian system.
Roush
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Department of Anesthesiology, Duke University Medical Center, Durham, NC 27710, USA.
Kwatra
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D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies; D000918:Antibody Specificity; D016466:CHO Cells; D002462:Cell Membrane / Q000378:metabolism; D006224:Cricetinae; D019204:GTP-Binding Proteins / Q000032:analysis / Q000737:chemistry / Q000378:metabolism*; D006160:Guanosine Triphosphate / Q000378:metabolism; D006801:Humans; D015151:Immunoblotting; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D018040:Receptors, Neurokinin-1 / Q000096:biosynthesis / Q000502:physiology*; D011994:Recombinant Proteins / Q000096:biosynthesis; D015290:Second Messenger Systems; D014162:Transfection
D000906:Antibodies; D046911:Macromolecular Substances; D018040:Receptors, Neurokinin-1; D011994:Recombinant Proteins; D006160:Guanosine Triphosphate; D019204:GTP-Binding Proteins