Related Experiment Videos
Induction of apoptosis by human Nbk/Bik, a BH3-containing protein that interacts with E1B 19K
1Center for Advanced Biotechnology and Medicine, Rutgers University, Piscataway, New Jersey 08854, USA.
Abstract:
The E1B 19-kilodalton protein (19K protein) is a potent apoptosis inhibitor and the adenovirus homolog of Bcl-2 (E. White, Genes Dev. 10:1-15, 1996). To obtain a better understanding of the biochemical mechanism by which the E1B 19K protein regulates apoptosis, proteins that interact with 19K have been identified; one of these is Bax (J. Han, P. Sabbatini, D. Perez, L. Rao, D. Mohda, and E. White, Genes Dev. 10:461-477, 1996), and another is Bak (S. N. Farrow, J. H. M. White, I. Martinou, T. Raven, K.-T. Pun, C. J. Grinham, J.-C. Martinou, and R. Brown, Nature (London) 374:731-733, 1995). Bax and Bak are Bcl-2 family members which contain Bcl-2 homology regions 1, 2, and 3 (BH1, BH2, and BH3), which interact with E1B 19K and Bcl-2 and promote apoptosis. Like Bax and Bak, Nbk was cloned from a yeast two-hybrid screen for proteins that interact with E1B 19K. Nbk contained BH3 but not BH1 or BH2. It also interacted with Bcl-2 but not with Bax. Both Bcl-2 and E1B 19K interacted with Nbk in vitro, and this interaction was highly specific. In vivo, the Nbk and E1B 19K proteins may colocalize with cytoplasmic and nuclear membranes. Nbk expression functionally antagonized 19K-mediated inhibition of apoptotic cell death and completely prevented transformation by E1A and E1B 19K. Nbk was sufficient for induction of apoptosis in the presence of mutant p53 and thus low levels of Bax, suggesting that Nbk functions independently of Bax to induce apoptosis. Nbk may therefore represent a novel death regulator which contains only a BH3 that interacts with and antagonizes apoptosis inhibitors such as the E1B 19K protein.
Insights
The novel Nbk protein antagonizes apoptosis inhibitors like the E1B 19K protein by interacting with its BH3 domain. Nbk induces apoptosis independently of Bax, suggesting a new role in regulating cell death.
Area of Science:
- Molecular Biology
- Cell Biology
- Virology
Background:
- The E1B 19-kilodalton (19K) protein inhibits apoptosis and is an adenovirus homolog of Bcl-2.
- Bcl-2 family members Bax and Bak contain BH1, BH2, and BH3 domains, interacting with 19K and Bcl-2 to promote apoptosis.
Purpose of the Study:
- To elucidate the biochemical mechanism of E1B 19K protein's regulation of apoptosis.
- To identify novel proteins interacting with the E1B 19K protein.
Main Methods:
- Yeast two-hybrid screening to identify proteins interacting with E1B 19K.
- In vitro interaction assays to confirm binding between Nbk, Bcl-2, and E1B 19K.
- In vivo studies to assess Nbk's effect on apoptosis and viral transformation.
Main Results:
- Nbk, a novel protein interacting with E1B 19K, contains only a BH3 domain.
- Nbk interacted with Bcl-2 but not Bax, and specifically with E1B 19K in vitro.
- Nbk expression antagonized 19K-mediated apoptosis inhibition and prevented E1A/E1B 19K-induced transformation.
- Nbk induced apoptosis independently of Bax, even with mutant p53.
Conclusions:
- Nbk represents a novel death regulator that antagonizes apoptosis inhibitors like E1B 19K.
- Nbk's function, mediated by its BH3 domain, is independent of Bax.
- Nbk may colocalize with cellular membranes in vivo, influencing apoptosis regulation.