Identification of the Ca2+/Calmodulin-dependent Protein Kinase II Regulatory Phosphorylation Site in the α-Amino-3-hydroxyl-5-methyl4-isoxazole-propionate-type Glutamate Receptor

doi:10.1074/jbc.272.52.32727

Identification of the Ca²⁺/Calmodulin-dependent Protein Kinase II Regulatory Phosphorylation Site in the α-Amino-3-hydroxyl-5-methyl4-isoxazole-propionate-type Glutamate Receptor*

From the Vollum Institute, Oregon Health Sciences University, Portland, Oregon 97201

Abstract

Ca²⁺/CaM-dependent protein kinase II (CaM-KII) can phosphorylate and potentiate responses of α-amino3-hydroxyl-5-methyl-4-isoxazole-propionate-type glutamate receptors in a number of systems, and recent studies implicate this mechanism in long term potentiation, a cellular model of learning and memory. In this study we have identified this CaM-KII regulatory site using deletion and site-specific mutants of glutamate receptor 1 (GluR1). Only mutations affecting Ser⁸³¹ altered the ³²P peptide maps of GluR1 from HEK-293 cells co-expressing an activated CaM-KII. Likewise, when CaM-KII was infused into cells expressing GluR1, the Ser⁸³¹ to Ala mutant failed to show potentiation of the GluR1 current. The Ser⁸³¹ site is specific to GluR1, and CaM-KII did not phosphorylate or potentiate current in cells expressing GluR2, emphasizing the importance of the GluR1 subunit in this regulatory mechanism. Because Ser⁸³¹ has previously been identified as a protein kinase C phosphorylation site (Roche, K. W., O’Brien, R. J., Mammen, A. L., Bernhardt, J., and Huganir, R. L. (1996) Neuron 16, 1179–1188), this raises the possibility of synergistic interactions between CaM-KII and protein kinase C in regulating synaptic plasticity.

Footnotes

↵* This work was supported in part by Javits Neuroscience Investigator Award NS 27037 from the National Institutes of Health.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵‡ Ph. D. student from the University of Chile.
↵§ To whom correspondence should be addressed: Vollum Inst. L-474, Oregon Health Sciences University, 3181 SW Sam Jackson Park Rd., Portland, OR 97201. Tel.: 503-494-6931; Fax: 503-494-4534; E-mail:soderlit{at}oshu.edu.
↵1 The abbreviations used are: LTP, long term potentiation; AMPA-R, α-amino-3-hydroxyl-5-methyl-4-isoxazole-propionate glutamate receptor; CaM-KII, Ca²⁺/calmodulin-dependent protein kinase II; GluR, glutamate receptor; PKA, protein kinase A; PKC, protein kinase C; PAGE, polyacrylamide gel electrophoresis; GST, glutathione S-transferase; TPA, 12-O-tetradecanoylphorbol-13-acetate.
- Received September 11, 1997.
- Revision received October 25, 1997.