Chromosomal mutations are the underlying cause of most inherited diseases and many developmental abnormalities. Mutations can also lead to alterations in gene expression in somatic cells, leading to loss of the normal differentiated phenotype and ultimately to cellular transformation. We are studying the mechanism of mutagenesis and DNA repair using SV40-based vectors as a probes to investigate the molecular mechanisms by which agents that damage DNA induce mutations in mammalian cells and how these mutations may be prevented by cellular DNA repair processes. Through use of the pZ189 shuttle vector, we have extensively characterized the types of mutations that occur in mammalian cells either spontaneously or in response to DNA damage. Analysis of the sequence specificity of these mutations has led to models which explain how the mammalian DNA polymerase introduces errors during DNA synthesis, causing mutations. Studies with the vector in in vitro DNA replication system indicate that cellular factors, in addition to DNA polymerase, appear to influence replication fidelity. Further studies should allow a characterization of these factors on the biochemical level. We are also using the SV40-based shuttle vector system as well as CAT expression vectors to assess the effects of cell wide responses to DNA-damaging treatments. We are also using in vitro DNA repair systems to investigate DNA repair at the molecular level. Understanding the mechanisms of regulation of cellular proliferation and differentiation is basic to understanding development of multicellular organisms. For the past several years, we have been studying an antimutagenic growth factor secreted by hamster cells transformed by SV40. This mitogenic inhibitor (Ml) strongly inhibits a proliferative response in untransformed hamster cells and normal rat cells stimulated with serum mitogens. Ml also inhibits a mitogenic response by normal hamster spleen lymphocytes stimulated with lectins that activate T cells (concanavalin A) or B cells (pokeweed mitogen). We have proposed that Ml might contribute to the high oncogenicity of the SV40-transformed cells by interfering with mobilization of immune effector cells at the site of tumor growth. We are also using SV40 to study the genetic basis of viral tissue tropism. We find that subcutaneously injected small t-antigen mutants of SV40 often induce abdominal lymphomas in hamsters, rather than the subcutaneous fibrosarcomas induced by wild-type SV40. The mutants may fail to produce a growth factor required for the in vivo transformation of non-proliferating cells.
Showing the most recent 10 out of 73 publications
