Protein Kinase C-mediated Interphase Lamin B Phosphorylation and Solubilization

doi:10.1074/jbc.272.34.21274

Protein Kinase C-mediated Interphase Lamin B Phosphorylation and Solubilization*

From the ^‡Department of Biochemistry, Norwegian College of Veterinary Medicine, 0033 Oslo, Norway, the ^¶Sealy Center for Oncology, University of Texas Medical Branch, Galveston, Texas 77555, the ^**Department of Biology, Amherst College, Amherst, Massachusetts 01002, and the ^‡Institut Jacques Monod, CNRS, Université Paris VII, 75251 Paris Cedex 5, France

Abstract

Disassembly of the sperm nuclear envelope at fertilization is one of the earliest events in the development of the male pronucleus. We report that nuclear lamina disassembly in interphase sea urchin egg cytosol is a result of lamin B phosphorylation mediated by protein kinase C (PKC). Lamin B of permeabilized sea urchin sperm nuclei incubated in fertilized egg G₁ phase cytosolic extract is phosphorylated within 1 min of incubation and solubilized prior to sperm chromatin decondensation. Phosphorylation is Ca²⁺-dependent. It is reversibly inhibited by the PKC-specific inhibitor chelerythrine, a PKC pseudosubstrate inhibitor peptide, and a PKC substrate peptide, but not by inhibitors of PKA, p34^cdc2 or calmodulin kinase II. Phosphorylation is inhibited by immunodepletion of cytosolic PKC and restored by addition of purified rat brain PKC. Sperm lamin B is a substrate for rat brain PKC in vitro, resulting in lamin B solubilization. Two-dimensional phosphopeptide maps of lamin B phosphorylated by the cytosolic kinase and by purified rat PKC are virtually identical. These data suggest that PKC is the major kinase required for interphase disassembly of the sperm lamina.

Footnotes

↵* This work was supported in part by National Science Foundation Grant IBN-9304394 and an Amherst College Faculty Research Award (to D. L. P.), National Institutes of Health Grant CA56869 (to A. P. F.), and Ligue Contre le Cancer (to J.-C. C.).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: Dept. of Biochemistry, Norwegian College of Veterinary Medicine, P. O. Box 8146 Dep., 0033 Oslo, Norway. Tel.: 47 22 96 45 69; Fax: 47 22 60 09 85; E-mail:philippe.collas{at}veths.no.
↵‖ Leukemia Society of America Scholar.
↵1 The abbreviations used are: NE, nuclear envelope; BAPTA, 1,2-bis(2-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid; DMAP, 6-dimethylaminopurine; NB, nuclear buffer; PAGE, polyacrylamide gel electrophoresis; PKA, protein kinase A (cAMP-dependent protein kinase); PKC, protein kinase C (Ca²⁺-dependent protein kinase); PKI, PKA inhibitor; CaM, calmodulin.
↵2 P. Collas, manuscript in preparation.
- Received March 10, 1997.
- Revision received May 16, 1997.