Role of COOH-terminal Phosphorylation in the Regulation of Casein Kinase Iδ

doi:10.1074/jbc.270.37.21689

Role of COOH-terminal Phosphorylation in the Regulation of Casein Kinase Iδ (*)

From the Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, Indianapolis, Indiana 46202-5122

^¶To whom correspondence and reprint requests should be addressed:
Dept. of Biochemistry and Molecular Biology, Indiana University School of Medicine, 635 Barnhill Dr., Indianapolis, IN 46202-5122.

↵^§ Present address: Dept. of Cell Biology and Physiology, Washington University School of Medicine, 660 S. Euclid Ave., St. Louis, MO 63110.

Abstract

Casein kinase I is a member of the casein kinase I (CKI) family, a group of second messenger independent protein kinases. We present evidence that the COOH-terminal domain of CKI has regulatory properties. CKI expressed in Escherichia coli was activated by heparin, as found previously, and by treatment with the catalytic subunit of type-1 protein phosphatase (CS1). Concomitant with activation by CS1, there was a reduction in the apparent molecular weight of CKI from 55,000 to 49,000 as judged by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. Truncation of CKI by removal of the COOH-terminal 110 amino acids eliminated the ability of CS1 to activate or to increase electrophoretic mobility. Casein kinase I α, a 37-kDa isoform that lacks an extended COOH-terminal domain, was not activated by CS1 or the presence of heparin. However, a chimeric enzyme consisting of CKIα fused to the COOH-terminal domain of CKI was activated by both heparin and CS1. Analysis of the effects of CS1 on a series of CKI COOH-terminal truncation mutants identified an inhibitory region between His and Pro, which contained six potential phosphorylation sites. From analysis of the specific activites of these truncation mutants, removal of the same region resulted in enzyme with a specific activity nearly 10-fold greater than wild-type. Thus, CKI activity can be regulated by phosphorylation of its COOH terminus, which may serve to create an autoinhibitory domain. This mechanism of regulation could have important consequences in vivo.

Footnotes

↵* This work was supported by National Institutes of Health Grant DK27221. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵¹ The abbreviations used are:

CKI

casein kinase I

CS1

catalytic subunit of type 1 protein phosphatase

PAGE

polyacrylamide gel electrophoresis

bp

base pair(s)

TLCK

N-p-tosyl-L-lysine chloromethyl ketone.
↵²L. Robinson, personal communication.
↵³P. R. Graves and P. J. Roach, unpublished results.
↵⁴L. Zhai, P. R. Graves, and P. J. Roach, unpublished results.
- Received April 27, 1995.
- Revision received July 8, 1995.