Activation of the cyclin D1 Gene by the E1A-associated Protein p300 through AP-1 Inhibits Cellular Apoptosis

Chris Albanese,Mark D'Amico,Anne T Reutens,Maofu Fu,Genichi Watanabe,Richard J Lee,Richard N Kitsis,Berthold Henglein,Maria Avantaggiati,Kumaravel Somasundaram,Bayar Thimmapaya,Richard G Pestell

doi:10.1074/jbc.274.48.34186

Abstract

The adenovirus E1A protein interferes with regulators of apoptosis and growth by physically interacting with cell cycle regulatory proteins including the retinoblastoma tumor suppressor protein and the coactivator proteins p300/CBP (where CBP is the CREB-binding protein). The p300/CBP proteins occupy a pivotal role in regulating mitogenic signaling and apoptosis. The mechanisms by which cell cycle control genes are directly regulated by p300 remain to be determined. The cyclin D1 gene, which is overexpressed in many different tumor types, encodes a regulatory subunit of a holoenzyme that phosphorylates and inactivates PRB. In the present study E1A12S inhibited the cyclin D1 promoter via the amino-terminal p300/CBP binding domain in human choriocarcinoma JEG-3 cells. p300 induced cyclin D1 protein abundance, and p300, but not CBP, induced the cyclin D1 promoter. cyclin D1 or p300 overexpression inhibited apoptosis in JEG-3 cells. The CH3 region of p300, which was required for induction of cyclin D1, was also required for the inhibition of apoptosis. p300 activated the cyclin D1 promoter through an activator protein-1 (AP-1) site at -954 and was identified within a DNA-bound complex with c-Jun at the AP-1 site. Apoptosis rates of embryonic fibroblasts derived from mice homozygously deleted of the cyclin D1 gene (cyclin D1(-/-)) were increased compared with wild type control on several distinct matrices. p300 inhibited apoptosis in cyclin D1(+/+) fibroblasts but increased apoptosis in cyclin D1(-/-) cells. The anti-apoptotic function of cyclin D1, demonstrated by sub-G(1) analysis and annexin V staining, may contribute to its cellular transforming and cooperative oncogenic properties.

Highlights

During the mid to late G1 phase of the cell cycle, the mammalian cell encounters a critical point where an irrevocable commitment takes place to either enter replicative DNA synthesis or to withdraw from the cell cycle with an unduplicated genome
Co-transfection experiments were conducted to determine whether E1A12S regulated the cyclin D1 promoter activity
The inhibition of cyclin D1 promoter activity by E1A12S contrasted with the effect of E1A12S on the cyclin A promoter which was induced (2.5-fold) in JEG-3 cells (Fig. 1A)

Summary

Introduction

During the mid to late G1 phase of the cell cycle, the mammalian cell encounters a critical point where an irrevocable commitment takes place to either enter replicative DNA synthesis or to withdraw from the cell cycle with an unduplicated genome. It was proposed that the inhibition of cyclin D1 protein levels by E1A represented a function evolved by E1A to overcome the G1/S checkpoint function of cyclin D1 that blocks unregulated S-phase entry and apoptosis [29, 37, 38] It was not known whether the cyclin D1 gene was a direct transcriptional target of E1A nor were the mechanisms by which E1A regulated cyclin D1 transcription known. Ribozyme ablation of p300 or CBP demonstrated distinct roles for p300 and CBP in cell cycle control and differentiation and suggested a requirement for p300 in cellular proliferation [41] These studies indicated that the identification of direct transcriptional targets of p300/CBP involved in cell cycle progression would be critical for understanding p300/CBP function in different cell types. These findings indicate that the induction of the cyclin D1 gene by p300 may play an important role in the inhibition of cellular apoptosis

Methods

Results

Conclusion