Fragile site FRA1H and transcriptional profiling of genes located within this region in tumor-derived cell-lines.

Common fragile sites (CFSs) are chromosome regions extending for hundreds or thousands of kilobases that exhibit gaps and breaks when the cells are exposed to replication stress and to some DNA binding compounds [1]. CFS (n=88) are located at specific loci on the chromosomes of all analysed individuals but the frequency of their expression varies among different individuals. Partial or complete molecular characterization of about twenty CFSs were performed but the basis of their fragility has not been definitively clarified. CFSs are regions of genomic instability frequently involved in sister chromatid exchanges in viral integrations and in chromosome mutations, and gene amplifications recurrent in cancer [rev. in 2]. In cancer cells, the analysis of the integrity and of the correct expression level of genes codified in the sequences of some characterized CFSs revealed in most of them the presence of rearrangements, loss of heterozygosity and modification or loss of expression of known or potential oncogenes and tumor suppressor genes [e.g. 3]. Recently, we analysed the FRA1H site (1q41-q42.1) [4] (Fig. 1). We found that it is one of the largest characterized CFSs extending over about 10 Mb. FRA1H is the first characterized CFS which expression is not induced by aphidicolin but instead by DAPI (4’,6-diamidino-2-phenylindole), by 5-azacytidine (5-azaC) and 5-azadeoxycytidine (5-azadC) and by infection of primary cells with adenovirus 12 (Ad12). Forty-nine genes are mapped at the FRA1H region. They include two microRNA (miRNA) genes, MIRN194-1 and MIRN215, codified as a cluster of 389 bp within an intron sequence of the gene IARS2, and two very large genes, USH2 (Usher syndrome 2A; 801 kb) and ESRRG (estrogen-related receptor gamma; 587 kb) (Fig. 2). Both miRNAs and very large genes have frequently been found located at fragile sites and there is evidence that the expression level or the structure of some of them are altered in cancer cells [5-8]. The FRA1H region is recurrently deleted in various neoplasms, mainly leukemias and lymphomas [9]. In this work, nine genes, localized in the FRA1H region, spaced to cover the complete CFS and with a possible role in transformation or regulation of cell proliferation, were investigated by PCR for homozygous deletions and by Real-time PCR for modifications or loss of gene expression in a panel of 19 cancer cell lines.