Beyond the Identification of Transcribed Sequences: Functional and Expression Analysis

9th Annual Workshop, October 28-31, 1999

Co-sponsored by the U.S. Department of Energy


KAPANADZE

Sequence homology between human and mouse genomic regions to identify the tumor suppressor gene involved in B cell chronic lymphocytic leukemia

Bagrat Kapanadze, Nataliy Makeeva, Olle Sangfelt, Martin Corcoran, Anna Baranova, Eugene Zabarovsky, Nick Yankovsky, Dan Grander and Stefan Einhorn

CCK, Research Laboratory of Radiumhemmet, Karolinska Hospital, Stockholm, Sweden

Previous studies have indicated the presence of a putative tumor suppressor gene on human chromosome 13q14, commonly deleted in patients with B-cell chronic lymphocytic leukemia (B-CLL). We have recently identified a minimally deleted region (MDR) of less than 10 kb, encompassing parts of two adjacent genes, termed Leu1 and Leu2 (leukemia-associated gene 1 and 2). Mutational analysis of Leu1 and Leu2 in 170 CLL samples revealed no small intragenic deletions or point mutations.  Subsequent examination of the genomic sequence around the MDR revealed several additional expressed transcripts (ESTs). In addition 50kb centromeric to this region another gene, Leu5 has been identified. This gene encodes a zing-finger protein  and shares homology to known genes involved in tumorogenesis. In order to further understand the genomic organisation of such a complex gene rich region, we decided to directly compare the human sequence with that of the mouse, as this may indicate the most important genes in the region, since critical genes tend to be highly conserved between mouse and human. A mouse genomic PAC library was screened with a number of probes covering a 100 kb distance in the human 13q14.3 region, including the MDR. Southern hybridization of subcloned fragments covering the MDR revealed several highly conserved areas, including exon1 and exon 2 of Leu2. Interestingly, human Leu1 does not seem to be conserved in mouse, by sequence analysis, whereas  Leu2 sequence was found to be highly conserved. In addition, the Leu5 protein encoding exon has > 95% homology with mouse sequence. In conclusion, following this analysis, the strongest candidates for a conserved tumor suppressor gene in this region are Leu5 and Leu2. Further work is required to elucidate their  mechanism of action in this disease and to further identify which gene or genes is the real tumor suppressor gene in BCLL.


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