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J Biol Chem, Vol. 274, Issue 16, 10685-10688, April 16, 1999

COMMUNICATION
Interaction of Heterotrimeric G Protein Galpha o with Purkinje Cell Protein-2
EVIDENCE FOR A NOVEL NUCLEOTIDE EXCHANGE FACTOR

Yuan Luo and Bradley M. Denker

From the Renal Division, Department of Medicine, Brigham and Women's Hospital and Harvard Medical School, Boston, Massachusetts 02115

The heterotrimeric G protein Galpha o is ubiquitously expressed throughout the central nervous system, but many of its functions remain to be defined. To search for novel proteins that interact with Galpha o, a mouse brain library was screened using the yeast two-hybrid interaction system. Pcp2 (Purkinje cell protein-2) was identified as a partner for Galpha o in this system. Pcp2 is expressed in cerebellar Purkinje cells and retinal bipolar neurons, two locations where Galpha o is also expressed. Pcp2 was first identified as a candidate gene to explain Purkinje cell degeneration in pcd mice (Nordquist, D. T., Kozak, C. A., and Orr, H. T. (1988) J. Neurosci. 8, 4780-4789), but its function remains unknown as Pcp2 knockout mice are normal (Mohn, A. R., Feddersen, R. M., Nguyen, M. S., and Koller, B. H. (1997) Mol. Cell. Neurosci. 9, 63-76). Galpha o and Pcp2 binding was confirmed in vitro using glutathione S-transferase-Pcp2 fusion proteins and in vitro translated [35S]methionine-labeled Galpha o. In addition, when Galpha o and Pcp2 were cotransfected into COS cells, Galpha o was detected in immunoprecipitates of Pcp2. To determine whether Pcp2 could modulate Galpha o function, kinetic constants kcat and koff of bovine brain Galpha o were determined in the presence and absence of Pcp2. Pcp2 stimulates GDP release from Galpha o more than 5-fold without affecting kcat. These findings define a novel nucleotide exchange function for Pcp2 and suggest that the interaction between Pcp2 and Galpha o is important to Purkinje cell function.

Copyright © 1999 by The American Society for Biochemistry and Molecular Biology, Inc.


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