Bcl-2 Prevents CD95 (Fas/APO-1)-induced Degradation of Lamin B and Poly(ADP-ribose) Polymerase and Restores the NF-κB Signaling Pathway

doi:10.1074/jbc.271.48.30354

Bcl-2 Prevents CD95 (Fas/APO-1)-induced Degradation of Lamin B and Poly(ADP-ribose) Polymerase and Restores the NF-κB Signaling Pathway*

From the Departments of ^‡ Medicine
^§ Cellular and Molecular Physiology, and
^¶ Microbiology and Immunology, Pennsylvania State University College of Medicine, Hershey, Pennsylvania 17033 and
the ^∥ Division of Oncology Research, Mayo Medical School, Rochester, Minnesota 55905

** To whom correspondence and reprint requests should be addressed: Present address:
Laboratory of Cell Growth Regulation, Box 036, UT MD Anderson Cancer Center, 1515 Holcombe Blvd., Houston, TX 77033.

Abstract

In the study presented here, we investigated the possible interactions between CD95 (Fas/APO-1) and Bcl-2 by studying the effects of Bcl-2 on the modulation of cellular pathways activated by CD95 using HeLa cells as a model system. We report that stable expression of Bcl-2 in HeLa cells is associated with multiple phenotypic changes. Treatment of HeLa cells with anti-CD95 monoclonal antibody (mAb) resulted in preferential degradation of lamin B compared with lamins A and C. Significant lamin B degradation was detected as early as 1 h after anti-CD95 mAb treatment. In contrast, lamins A and C as well as actin remained unchanged until 4 h after treatment with anti-CD95 mAb, a time point that correlated with the period of DNA fragmentation. These results indicate that selective degradation of lamin B is an early cellular event in response to activation of the CD95 pathway and that it precedes DNA fragmentation. Overexpression of Bcl-2 resulted in prevention of lamin B degradation and DNA fragmentation into oligonucleosome fragments in response to the apoptotic signal by anti-CD95 mAb. In addition, in Bcl-2-overexpressing cells that were protected against apoptosis, anti-CD95 mAb-induced cleavage of poly(ADP-ribose) polymerase was completely blocked. Overexpression of Bcl-2 also resulted in restoration of the CD95-mediated signaling pathway involving activation of the transcription factor NF-κB (p50/RelA). These findings suggest that Bcl-2 prevents apoptosis in part by preventing the degradation of major nuclear polypeptides such as lamin B and poly(ADP-ribose) polymerase. In addition, our results demonstrate that CD95-mediated signaling involves activation of NF-κB (p50/RelA).

Footnotes

↵* This work was supported in part by National Institutes of Health Grant CA 56564 and American Institute for Cancer Research Grants 94B93 and 96A077 (to R. 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 abbreviations used are:

PARP

poly(ADP-ribose) polymerase

TNF

tumor necrosis factor

mAb

monoclonal antibody

ICE

interleukin-1β-converting enzyme

EMSA

electrophoretic mobility shift assay

BAPTA

1,2-bis(2-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid

TPCK

L-1-tosylamido-2-phenylethyl chloromethyl ketone.
- Received May 30, 1996.
- Revision received August 6, 1996.