The concept of tumor suppressor gene dysregulation is a central theme in cancer biology. A powerful method for identifying potential tumor suppressor genes is loss of heterozygosity (LOH) analysis. Several recent studies have demonstrated an area of LOH on chromosome 6 band q22 in T-cell leukemias, Non-Hodgkin's lymphomas refractory to chemotherapy, breast cancer and melanoma suggesting the presence of a tumor suppressor gene at this locus. Using a c-myb specific probe, we have previously cloned a rearrangement from this region, designated myb recombination region (MRR). In our current studies, we have identified and begun analysis of a putative tumor supressor gene within this rearrangement. Based on a prototype human cDNA clone, MRR is a novel nuclear WD-protein with an associated SH3 domain. Genomic analysis reveals MRR to be a 250 kB gene containing 28 primary exons in addition to multiple alternative exons and at least two polyadenylation sites. Analysis of MRR expression by RT-PCR reveals evidence of multiple transcript sizes in these cell lines. This, together with Analysis of multiple MRR cDNA clones, indicates the presence of multiple alternatively spliced transcripts for this gene. In normal human tissues, maximum expression is seen in brain, testis and ovary with variable expression in a variety of other tissues. Expression in human tumor cell lines reveals variable exprssion across given tumor types. Florescent in situ hybridization studies localize MRR to a region of 6q deleted in the T-cell leukemic cell line RPMI 8402. Southern blotting experiments using a panel of exon-specific probes demonstrates alterations of MRR in a variety of human leukemia and lymphoma cell lines. Additional alterations identified include a frame shift mutation in a cDNA from a uterine sarcoma and an aberrant mRNA species in a non-small cell lung cancer harboring a t1:6 translocation. Further analysis of ests from a variety of CGAP tumors revels truncations and other message alterations in prostate and other tumor types. These results indicate that an important and novel tumor suppressor gene has been isolated that resides in a region of chromosome 6q that is the site of frequent alteration in a wide range of tumor types. Future work will focus on the function of this gene product and its' alternatively spliced forms in control of normal and tumor cell proliferation/ differentiation.
