Subunit Structure and Regulation of Protein Phosphatase-1 in Rat Liver Nuclei

doi:10.1074/jbc.270.29.17257

Subunit Structure and Regulation of Protein Phosphatase-1 in Rat Liver Nuclei (*)

From the Afdeling Biochemie, Faculteit Geneeskunde, Katholieke Universiteit Leuven, B-3000 Leuven, Belgium

^¶ To whom correspondence should be addressed. Tel.: 32-16-345700; Fax: 32-16-345995.

Abstract

The activity of protein phosphatase-1 in rat liver nuclei (PP-1N) was decreased by up to 97% by associated inhibitory polypeptides, depending on the assay and extraction conditions. These inhibitors were rapidly degraded by endogenous proteases, resulting in the accumulation of active heat-stable intermediates. Two major species of PP-1N could be differentiated by fractionation of a nuclear extract. PP-1N contained, besides the -isoform of the catalytic subunit, an inhibitory polypeptide of 111 kDa. PP-1N was found to be an inactive heterodimer between the -isoform of the catalytic subunit and NIPP-1, a nuclear inhibitor of PP-1, which in its undegraded form is heat labile and migrates during SDS-polyacrylamide gel electrophoresis as a polypeptide of 41 kDa. Native hepatic NIPP-1 displayed a reduced affinity for the catalytic subunit after phosphorylation by protein kinase A in vitro and after glucagon-induced phosphorylation in vivo.

Footnotes

↵^§ On leave from the Institute of Biochemistry and Biophysics (Polish Academy of Sciences) in Warsaw, Poland.
↵* This work was supported by the ASLK, by the Belgian Fund for Medical Scientific Research (Grant 3.0041.93), and by a Concerted Research Action of the “Vlaamse Executieve.” 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:

PP-1

protein phosphatase 1

PP-1_C

catalytic subunit of PP-1

NIPP-1

nuclear inhibitor of PP-1

PP-1N

nuclear PP-1

PP-1G

PP-1 associated with glycogen

PP-1M

PP-1 associated with myofibrils

PP-1E

PP-1 associated with endoplasmic reticulum

PP-1S

soluble fraction of PP-1

PP-2A_D

dimeric form of protein phosphatase-2A, previously called PCS_L or PP2A₂

TLCK

1-chloro-3-tosylamido-7-amino-2-heptanone

TPCK

L-1-tosylamido-2-phenylethyl chloromethyl ketone

DIG

digoxygenin

PAGE

polyacrylamide gel electrophoresis

PKA

protein kinase A.
↵²A. Van Eynde, S. Wera, M. Beullens, S. Torrekens, F. Van Leuven, W. Stalmans, and M. Bollen, unpublished data.
- Received March 10, 1995.
- Revision received May 8, 1995.