My laboratory has two principal aims: 1) understanding the molecular mechanism of the functionof proto-oncogenes and suppressor genes, and 2) gene therapy. We have been analyzing the mechanism of activation of NF-kB/Rel family of proteins, which are involved in a variety of immune functions, growth regulation and programmed cell death. We have shown that in response to external signals, the IkB protein is modified by phosphorylation, ubiquitinated, and degraded by proteasomes, thereby allowing the release of NF-kB proteins to the nucleus to activate transcription of genes containing kB sites (see Figure). We are currently trying to identify and molecularly clone the inducible IkB kinase and define the proteins involved in the proteasome degradation pathway. We have identified a series of proteins that associate with NF-kB proteins by two hybrid plasmid systems. To study the function of individual members of the NF-kB/IkB family, we have used two methods: 1) generate homozygous null mice, and 2) generation of IkB transdominant negative mutant. We have shown that when Bcl-3, a IkB member, is inactivated, the mice are more susceptible to pathogens and fail to generate specific antibodies due to loss of germinal centers. Using transdominant negative IkBa, we have observed that upon addition of TNFa, the cells undergo programmed cell death (apoptosis). Presently, we are attempting to delineate the pathway of programmed cell death caused by lack of NF-kB activity. We have recently embarked on a program to understand the function of breast cancer genes (BRCA-1 and BRCA-2) involved in hereditary breast cancer. Homozygous deletion of either of these genes is closely linked to hereditary ovarian and breast carcinomas. We have shown that BRCA-1 protein is nuclear in its localization, undergoes extensive phosphorylation during the cell cycle, and likely acts as a transcriptional activator. We are studying whether other cellular proteins interact with BRCA-1 and BRCA-2 proteins. We have only recently generated antibodies to BRCA-2 predicted peptides and plan to use the antisera to localize the BRCA-2 protein. Our long-term goal with these two suppressor genes is to understand their role in development and cell regulation. Our gene therapy project revolves around generating
versatile and more efficient vectors for gene delivery. We have extensively
used retroviral vectors, but they have two major limitations: 1) inability
to infect non-dividing cells, and 2) lack of sustained expression. To
overcome some of these problems, we have used replication-defective
recombinant adenoviral vectors, but they have also serious limitations,
largely immune-problems. We have, therefore, developed lentiviral vectors
which are capable of transducing "non-dividing" cells in vitro
and in vivo. Using VSVG protein, we have been able to expand
the host range of these recombinant lentiviruses. We have been able
to successfully transduce brain, islets, lung and hematopoietic cells.
We are also generating vectors which can be targeted by using single-chain
variable fragment or ligands. Finally, we are generating mice where
the Factor IX gene is "knocked-out" to use as a model system.
Chapman, M. and Verma, I. M. (1996). Transcriptional Activation by Wildtype but not Mutant BRCA-1. Nature (in press). Naldini, L., Blömer, U., Gallay, P., Gage, F. H., Verma, I. M. and Trono, D. (1996). In vivo gene delivery and stable transduction of postmitotic cells by a lentiviral vector. Science 272:263-267. Verma, I. M., Stevenson, J. K., Schwarz, E. M., Van Antwerp, D. and Miyamoto, S. (1995). Rel/NF-kB/IkB family: intimate tales of association and dissociation. Genes Dev. 9:2723-2735. Dai, Y., Schwarz, E. M., Gu, D., Zhang, W.-W., Sarvetnick, N. and Verma, I. M. (1995). Cellular and humoral immune responses to adenoviral vectors containing factor IX gene: tolerization of both factor IX and vector antigens allows for long-term expression. Proc. Natl. Acad. Sci. USA 92:1401-1405. Miyamoto, S., Maki, M., Schmitt,
M.J. and Verma, I.M. (1994). Tumor necrosis factor a-induced phosphorylation
of IkBa is a signal for its degradation but not dissociation from NF-kB.
Proc. Natl. Acad. Sci. USA 91:12740-12744. Inder Verma received his Ph.D. from the Weizmann Institute of Science and was a postdoctoral fellow at MIT. He is an American Cancer Society Professor of Molecular Biology and recipient of an NIH Outstanding Investigator award. Dr. Verma is the Director of the Laboratory of Genetics at the Salk Institute. |
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