Regulation of Phosphatidylinositol 4-Phosphate 5-Kinase fromSchizosaccharomyces pombe by Casein Kinase I

doi:10.1074/jbc.274.2.1147

Regulation of Phosphatidylinositol 4-Phosphate 5-Kinase fromSchizosaccharomyces pombe by Casein Kinase I*

From the Department of Biological Sciences, St. John’s University, Jamaica, New York 11439 and the ^‡Department of Cell and Molecular Biology, Northwestern University Medical School, Chicago, Illinois 60611-3008

Abstract

Phosphatidylinositol (4)P 5-kinase (PtdIns(4)P 5-kinase) catalyzes the last step in the synthesis of phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P₂). PtdIns(4,5)P₂ is a precursor of diacylglycerol and inositol 1,4,5-trisphosphate and is also involved in regulation of actin cytoskeleton remodeling and membrane traffic. To satisfy such varied demands in several aspects of cell physiology, synthesis of PtdIns(4,5)P₂ must be stringently regulated. In this paper we describe extraction, purification, and characterization of PtdIns(4)P 5-kinase from the plasma membranes of Schizosaccharomyces pombe. We also provide evidence that PtdIns(4)P 5-kinase is phosphorylated and inactivated by Cki1, the S. pombe homolog of casein kinase I. Phosphorylation by Cki1 in vitro decreases the activity of PtdIns(4)P 5-kinase. In addition, and most importantly, overexpression of Cki1 in S. pombe results in a reduced synthesis of PtdIns(4,5)P₂ and in a lower activity of PtdIns(4)P 5-kinase associated with the plasma membrane. These results suggest that PtdIns(4)P 5-kinase is a target of Cki1 in S. pombe and that Cki1 is involved in regulation of PtdIns(4, 5)P₂ synthesis by phosphorylating and inactivating PtdIns(4)P 5-kinase.

Footnotes

↵* This work was supported by American Cancer Society Grant BE-239 (to A. V.).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.
↵§ To whom correspondence should be addressed: Tel.: 718-990-6287; Fax: 718-990-5958; E-mail: vancuraa{at}stjohns.edu.
Abbreviations:

PtdIns

phosphatidylinositol

PtdIns(4)P

phosphatidylinositol 4-phosphate

PtdIns(4

5)P₂, phosphatidylinositol 4,5-bisphosphate

DG

diacylglycerol

CK1

casein kinase I

PAGE

polyacrylamide gel electrophoresis

Cki1

casein kinase I from Schizosaccharomyces pombe (product of cki1⁺ gene)

PA

phosphatidic acid

PS

phosphatidylserine

GAP

GTPase-activating protein

MES

4-morpholineethanesulfonic acid

CHAPS

3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonic acid

CHAPSO

3-[(3-cholamidopropyl)dimethylammonio]-2-hydroxy-1- propanesulfonic acid

MOPS

4-morpholinepropanesulfonic acid

GTPγS

guanosine 5′-O-(thiotriphosphate).
- Received August 14, 1998.
- Revision received October 29, 1998.
The American Society for Biochemistry and Molecular Biology, Inc.