Clustering and Enhanced Activity of an Inwardly Rectifying Potassium Channel, Kir4.1, by an Anchoring Protein, PSD-95/SAP90*
- Yoshiyuki Horio‡,
- Hiroshi Hibino‡,
- Atsushi Inanobe§,
- Mitsuhiko Yamada‡,
- Masaru Ishii‡,
- Yoshihiko Tada‡,
- Eisaku Satoh‡¶,
- Yutaka Hata‖,
- Yoshimi Takai‖** and
- Yoshihisa Kurachi‡§‡
- From the ‡Department of Pharmacology II, **Department of Molecular Biology and Biochemistry, Faculty of Medicine, Osaka University, Suita, Osaka 565, Japan, the §Department of Cell Signaling, Yamagata University School of Medicine, Yamagata 993-23, Japan, the ¶Department of Pharmacology, Akita University School of Medicine, Akita 010, Japan, and the ‖Takai Biotimer Project, ERATO, Japan Science and Technology Corporation, Kobe 651-22, Japan
Abstract
An inwardly rectifying potassium channel predominantly expressed in glial cells, Kir4.1/KAB-2, has a sequence of Ser-Asn-Val in its carboxyl-terminal end, suggesting a possible interaction with an anchoring protein of the PSD-95 family. We examined the effects of PSD-95 on the distribution and function of Kir4.1 in a mammalian cell line. When Kir4.1 was expressed alone, the channel immunoreactivity was distributed homogeneously. In contrast, when co-expressed with PSD-95, prominent clustering of Kir4.1 in the cell membrane occurred. Kir4.1 was co-immunoprecipitated with PSD-95 in the co-expressed cells. Glutathione S-transferase-fusion protein of COOH terminus of Kir4.1 bound to PSD-95. These interactions disappeared when the Ser-Asn-Val motif was deleted. The magnitude of whole-cell Kir4.1 current was increased by 2-fold in cells co-expressing Kir4.1 and PSD-95 compared with cells expressing Kir4.1 alone. SAP97, another member of the PSD-95 family, showed similar effects on Kir4.1. Furthermore, we found that Kir4.1 as well as SAP97 distributed not diffusely but clustered in retinal glial cells. Therefore, PSD-95 family proteins may be a physiological regulator of the distribution and function of Kir4.1 in glial cells.
Footnotes
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↵* This work was supported in part by grants from the Ministry of Education, Science, Sports and Culture of Japan and the Research for the Future Program (JSPS-RFTF96L00302) of The Japan Society for the Promotion of Science and the Ichiro Kanehara Foundation.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.
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↵‡ To whom correspondence should be addressed: Dept. of Pharmacology II, Faculty of Medicine, Osaka University, 2-2, Yamadaoka, Suita, Osaka 565, Japan. Tel.: 81-6-879-3510 or 3512; Fax: 81-6-879-3519; E-mail: ykurachi{at}pharma2.med.osaka-u.ac.jp.
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↵1 The abbreviations used are: NMDA,N-methyl-d-aspartate; BCCP, biotin carboxylase carrier protein; DMEM, Dulbecco’s modified Eagle’s medium; FITC, fluorescein isothiocianate; GST, glutathione S-transferase; HEK, human embryonic kidney; PBS, phosphate-buffered saline; PAGE, polyacrylamide gel electrophoresis; F, farad(s); S, Siemens.
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↵2 Y. Hata and Y. Takai, submitted for publication.
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- Received November 11, 1996.
- Revision received February 14, 1997.
