Heteronuclear Ribonucleoproteins C1 and C2, Components of the Spliceosome, Are Specific Targets of Interleukin 1β-converting Enzyme-like Proteases in Apoptosis

doi:10.1074/jbc.271.46.29335

Heteronuclear Ribonucleoproteins C1 and C2, Components of the Spliceosome, Are Specific Targets of Interleukin 1β-converting Enzyme-like Proteases in Apoptosis*

From the ^‡ Queensland Cancer Fund Research Unit, Queensland Institute of Medical Research, P. O. Royal Brisbane Hospital, Herston, Brisbane, Queensland 4029, Australia,
^§ Department of Surgery, University of Queensland, St. Lucia, Queensland 4072, Australia,
^¶ The Hanson Centre for Cancer Research, P. O. Box 14, Rundle Mall, Adelaide, South Australia 5000, Australia,
^∥ Commonwealth Scientific and Industrial Research Organization, Division of Biomolecular Engineering, Parkville, Victoria 3052, Australia,
** Howard Hughes Medical Institute Research Laboratories, University of Pennsylvania School of Medicine, Department of Biochemistry and Biophysics, Philadelphia, Pennsylvania 19104-6148, and
^‡‡ Department of Pharmacology and the Jefferson Cancer Institute, Thomas Jefferson University, Philadelphia, Pennsylvania 19107

^§§To whom correspondence should be addressed. Tel: 61-7-3362-0335; Fax: 61-7-3362-0106; E-mail: dianneW{at}qimr.edu.au.

Abstract

Apoptosis induced by a variety of agents results in the proteolytic cleavage of a number of cellular substrates by enzymes related to interleukin 1β-converting enzyme (ICE). A small number of substrates for these enzymes have been identified to date, including enzymes involved in DNA repair processes: poly(ADP-ribose) polymerase and DNA-dependent protein kinase. We describe here for the first time the specific cleavage of the heteronuclear ribonucleoproteins (hnRNPs) C1 and C2 in apoptotic cells induced to undergo apoptosis by a variety of stimuli, including ionizing radiation, etoposide, and ceramide. No cleavage was observed in cells that are resistant to apoptosis induced by ionizing radiation. Protease inhibitor data implicate the involvement of an ICE-like protease in the cleavage of hnRNP C. Using recombinant ICE-like proteases and purified hnRNP C proteins in vitro, we show that the C proteins are cleaved by Mch3α and CPP32 and, to a lesser extent, by Mch2α, but not by ICE, Nedd2, Tx, or the cytotoxic T-cell protease granzyme B. The results described here demonstrate that the hnRNP C proteins, abundant nuclear proteins thought to be involved in RNA splicing, belong to a critical set of protein substrates that are cleaved by ICE-like proteases during apoptosis.

Footnotes

↵* This work was supported by grants from the Queensland Cancer Fund and the University of Queensland Cancer Research Fund (to D. W. and M. F. L.) and the Royal Adelaide Hospital Research Fund and the Wellcome Trust (to S. K.). 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.
↵¹ The abbreviation used are:

ICE

interleukin 1β-converting enzyme

PK

protein kinase

hnRNP

heteronuclear ribonucleoprotein

TPCK

L-1-tosylamido-2-phenylethyl chloromethyl ketone

TLCK

N^α-p-tosyl-L-lysine chloromethyl ketone

PMSF

phenylmethylsulfonyl fluoride

CHAPS

3-[(3-cholamidopropyl)dimethylamino]-1-propanesulfonate

DTT

dithiothreitol

PAGE

polyacrylamide gel electrophoresis

YVAD-CMK

Tyr-Val-Ala-Asp-chloromethylketone

DEVD-CHO

Ac-Asp-Glu-Val aspartic acid aldehyde

PBS

phosphate-buffered saline

VAD-FMK

Z-Val-Ala-Asp-CH₂F

PIPES

1,4-piperazinediethanesulfonic acid

HPLC

high performance liquid chromatography

CPP32

cysteine protease protein of molecular mass 32 kDa

Mch

mammalian Ced-3 homolog

MACH

MORT 1-associated CED-3/ice homolog

FADD

Fas-associated death domain protein

FLICE

FADD-like ICE

sn

small nuclear.
- Received August 9, 1996.