The proposed research deals with the control of oncogene expression by DNA-binding proteins - particularly as manifested in cells of the colonic epithelium. It is based on the premise that transcription of oncogenic and proto-oncogenic DNA sequences is regulated by proteins which control the accessibility, topology, and template function of the DNA. Our experimental approaches focus on the alterations in nuclear protein composition and metabolism that precede or accompany malignant transformation in vivo and in cultured cells. We have shown previously that the synthesis and post-synthetic modifications of specific colonic nuclear proteins are altered in a characteristic fashion at very early stages of tumor induction by the alkylating carcinogen, 1,2-dimethylhydrazine (DMH). The relevance of such changes to oncogenesis and oncogene expression is being assessed by careful comparisons of the responses of rodent strains that differ markedly in their susceptibilities to DMH. To further understand the significance of such changes and their relevance to altered oncogene expression, new procedures have been developed for analyzing chromatin structure at the nucleosomal level and for the isolation of nucleosomes containing the transcribed DNA sequences. These methods are being employed to monitor changes in oncogene expression in systems in which particular oncogenes or proto-oncogenes are sequentially activated or repressed. We are particularly interested in proteins entering the nucleus and binding to oncogenic DNA sequences when transcription is suppressed. Primary attention is directed at c-myc, because myc-encoded protein(s) appear to be associated with the nuclear matrix, and the latter structure appears to play a key role in the topological constraints on chromosomal 'loop' domains. The role of protein phosphorylation in matrix binding will be investigated, as will the more general problem of how aberrant phosphorylation of DNA-binding proteins might affect the structure and function of oncogenic DNA sequences. A related aspect of our research deals with the mechanisms by which DNA-regulatory proteins are transported into the cell nucleus following their synthesis in the cytoplasm. """"""""Signal"""""""" sequences believed to be involved in directing karyophilic proteins across the nuclear envelope have been prepared by solid-phase peptide synthesis. They are being coupled to antibodies for microinjection experiments on living cancer cells to ascertain whether specific targeting of the nucleus can be achieved.
| Boffa, L C; Carpaneto, E M; Mariani, M R et al. (1997) Contrasting effects of PNA invasion of the chimeric DMMYC gene on transcription of its myc and PVT domains. Oncol Res 9:41-51 |
| Boffa, L C; Morris, P L; Carpaneto, E M et al. (1996) Invasion of the CAG triplet repeats by a complementary peptide nucleic acid inhibits transcription of the androgen receptor and TATA-binding protein genes and correlates with refolding of an active nucleosome containing a unique AR gene sequence. J Biol Chem 271:13228-33 |
