In many Balb/c mouse myeloma cell lines the c-myc oncogene normally located on chromosome 15 is often translocated to a H chain constant region locus on chromosome 12. In the majority of myelomas studied, the translocation of c-myc is to the alpha constant region sequence (C-alpha) of the H chain """"""""non-productive"""""""" gene (the mRNA coding strand of DNA for the c-myc gene is on the anti-sense strand for the C-alpha gene). While unrearranged C-alpha alleles are transcriptionally silent, C-alpha genes to which the c-myc oncogene has translocated are transcriptionally active. Several discrete cytoplasmic RNA species hybridizing to a alpha-C region probe can be detected. Some of these appear to be transcribed from the C-alpha sense strand. In a mouse myeloma MOPC 315 variant line (V-1), which has lost the alpha-H chain gene, cotranscription of c-myc and C-alpha sequences has been detected. In this application the products of the c-myc-C-alpha transcriptional unit in V-1 cells will be examined. A genomic clone of this locus has been isolated. The initiation and termination sites will be mapped, and the products of transcription analyzed. The sizes of C-alpha and c-myc proteins will be determined using hybrid-selection and cell-free translation in a rabbit reticulocyte system. The steps in synthesis and processing of C-alpha proteins will be determined. Finally, the possible relationship of the translocation of c-myc to C-alpha to the loss of normal immunosuppression of cell growth will be studied. Methods will include tissue culture, RNA and DNA blot hybridization, DNA sequencing, cell-free translation, and acrylamide and agarose gel electrophoresis. The experiments will characterize the RNA and protein products of the c-myc oncogene-C-alpha gene locus and help define their possible roles in the neoplastic transformation of myeloma cells. (X)
Showing the most recent 10 out of 56 publications
