Karyotypic analysis of human leukemias and lymphomas has led to the observation that isolated chromosomal translocations may be implicated in the pathogenesis of these malignant disorders. Examples of such translocations include the t(9;22) found in chronic granulocytic leukemia and involving the c-abl gene and the t(8;14) of Burkitt's lymphoma which involves the c-myc gene. Such precedents suggest that new genes involved in the control of cellular proliferation might be identified in the regions surrounding other translocation breakpoints not known to be associated with cellular proto-oncogenes. We have identified a novel single translocation breakpoint t(3;4) in the leukemic cells of a patient with a B cell leukemia. This breakpoint is adjacent to a region on chromosome 3 which has been implicated in B cell lymphomas and non-lymphocytic leukemias and to a region on chromosome 4 which has been intensively mapped in the search for the Huntington's Disease gene. The use of genomic probes from this region has permitted mapping the breakpoint to within 120 kb of one of the probes. I propose to clone and characterize the breakpoint and surrounding DNA regions in order to identify gene(s) which may be of pathogenic significance to the malignant transformation of these cells. In order to accomplish this aim, a cell line has been derived from the leukemic cells which contain the same translocation and no other karyotypic abnormality. Fusion of this line with a Chinese Hamster Ovary cell line has resulted in a hybrid line containing the translocated human chromosome. Using chromosome """"""""walking"""""""" techniques and the chromosome 4 probe to cosmid clones containing normal human chromosome 4 sequences which adjoin the breakpoint will be identified. Subclones of this flanking region will be used to identify clones which contain the actual t(3;4) breakpoint from a lambda genomic library made from the hybrid cell line DNA. Clones containing the breakpoint will be sequenced and examined for areas of homology to other sequences in GenBank. These clones and flanking sequences from both chromosomes will be used to screen Northern blots containing RNA from these and other normal and malignant B cell populations for the aberrant expression of corresponding mRNA(s). In addition, DNA from these cells will be examined by Southern blotting techniques for the presence of occult rearrangements or deletions in this region using the same probes. This study should provide valuable additional insights into sequences involved in chromosomal translocations. It could lead to the identification of new transforming genes and could help define their role in this and other leukemias. In addition, it will provide the candidate with valuable experience in the area of chromosome mapping and cloning which should provide the basis for an independent career as a clinical investigator with expertise in the molecular biology of malignancy.
