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 recent work, we have demonstrated that MRR1 is the human homolog of the recently described Ahi-1 (Abelson helper integration) gene altered by viral insertion in a variety of murine T and B cell neoplasms. Based on a prototype human cDNA clone, the human MRR1/Ahi-1 gene encodes a novel nuclear WD-protein with an associated SH3 domain. Genomic analysis reveals MRR1/Ahi-1 to be a 250 kB gene containing 28 primary exons in addition to multiple alternative exons and at least two polyadenylation sites. Analysis of MRR1/Ahi-1 expression by RT-PCR revealed evidence of multiple alternatively spliced transcript forms in normal human tissues and cell lines. In normal human tissues, maximum expression is seen in brain, testis and ovary with variable expression in other tissues. Expression in human tumor cell lines reveals variable exprssion across given tumor types along with expression of T-cell tumor specific splice forms. Southern blotting experiments using a panel of exon-specific probes demonstrates a minimal frequency of gross structural alterations of MRR1/Ahi-1 in a variety of human leukemia and lymphoma cell lines. Additional alterations have been identified including 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 reveals truncations and other message alterations in acute myeloid leukemia, prostate cancer and other tumor types. These results together 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. This is further confirmed by the recent finding of tumor cell growth suppression by consititutive expression of prototype MRR1/Ahi-1 and a number of domain specific deletion constructs. Work has also begun on the expression of the MRR1/Ahi-1 proteins. Work to date demonstrates both nuclear and cytoplasic anti-peptide antibody specific forms of MRR1/Ahi-1 proteins are expressed with a spectrum of molecular weights consistent with the finding of multiple mRNA forms produced based on RT-PCR data. The prototype form of the protein appears to be expressed predominantly in the cytoplasm though translocation to the nucleus occurs on induction of apoptosis by a variety of drugs. A form lacking the SH3 domain produced through use of an internal polyadenylation site within the gene appears to be expressed predominantly in the nucleus. In a range of human T-cell leukemia/lymphoma and breast carcinoma cell lines, expression of one or more forms of MRR1/Ahi-1 protein appears to be lacking lending further credence to a possible role for this gene in these tumor types. Future work will continue to focus on the function of this gene product and its' alternatively spliced forms in control of normal and tumor cell proliferation/ differentiation and characterization of the proteins with which it interacts..
NIH ExPorter
NIH ExPorter