I propose to analyze the chromatin structure and transcriptional expression of the endogenous cellular oncogene, c-myc, in normal chicken cells and in bursal lymphoma cell lines. The c-myc gene is the cellular homologue of the transforming gene of MC29 virus, is conserved in evolution, and is expressed at high levels in most avian bursal lymphomas and in some human tumors of lymphoid and nonlymphoid origin. Enhanced myc expression in avians usually results from regional activation by the long-terminal repeat (LTR) of the avian leukosis virus (ALV) either through downstream promotion or through other undefined cis-acting field effects. A peculiar feature of the myc region is that transcription occurs from both DNA strands and involves several as yet undefined transcriptional units. I will use molecular clones, including single-strand clones in M13 phage, to determine if the levels of mRNA, the transcriptional units, and the chromatin structure of c-myc are modulated in the course of normal development. By comparing patterns of opposite strand transcription in lymphoid and non-lymphoid tissues I will test the hypothesis that opposite-strand transcription is restricted to B cells. Transcription from the region will be analyzed for both DNA strands by quantitative dot-blot hybridization, Northern blot analysis, and in situ hybridization. The transcriptional domains will be defined by nuclear run-off transcription and cDNA cloning of myc mRNAs followed by heteroduplex mapping. Utilizing ALV transformed cell lines that contain only one c-myc locus linked to a nearby LTR, I will define the alterations in chromatin structure and DNA methylation that are imposed by LTR integration. Several experiments are proposed which will attempt to determine whether enhanced myc expression is necessary to maintain transformation in lymphoma cell lines or whether it is a property of rapidly proliferating cells. The techniques and probes generated in this study will be used to extend the analysis to the expression of c-myc in normal and leukemic human hematopoietic cells.
