cytidylyl-3--5--guanosine and Ovarian-Neoplasms

Several genes that contain the PR (PRDI-BF1 and RIZ) domain have been linked with human cancers. We describe here a new PR-domain-containing gene designated as PRDM5 (PFM2). A PRDM5 cDNA was isolated based on its homology to the PR domain of RIZ1 (PRDM2). The gene encodes an open reading frame of 630 amino acids and contains a PR domain in the NH-terminal region followed by 16 zinc finger motifs. Radiation hybrid analysis mapped PRDM5 to human chromosome 4q26, a region thought to harbor tumor suppressor genes for breast, ovarian, liver, lung, colon, and other cancers. The gene has a CpG island promoter and is silenced in human breast, ovarian, and liver cancers. A recombinant adenovirus expressing PRDM5 caused G2/M arrest and apoptosis upon infection of tumor cells. These results suggest that inactivation of PRDM5 may play a role in carcinogenesis.

Topics: Amino Acid Sequence; Apoptosis; Base Sequence; Breast; Breast Neoplasms; Carcinoma, Hepatocellular; Cell Cycle; Cell Division; Cell Line, Tumor; Dinucleoside Phosphates; DNA-Binding Proteins; Female; Gene Silencing; Humans; Liver; Liver Neoplasms; Molecular Sequence Data; Ovarian Neoplasms; Transcription Factors

Breast cancer is a genetic disease arising from a series of germ-line and/or somatic DNA changes in a variety of genes, including BRCA1 and BRCA2. DNA modifications have been shown to occur by a number of mechanisms that include DNA methylation. In some cases, the aberrant methylation of CpGs within 5' regulatory regions has led to suppression of gene activity. In this report we describe a variation in the pattern of DNA methylation within the regulatory region of the BRCA1 gene. We found no evidence of methylation at CpGs within the BRCA1 promoter in a variety of normal human tissues. However, screening of a series of randomly sampled breast carcinomas revealed the presence of CpG methylation adjacent to the BRCA1 transcription start site. One such methylated CpG occurs at a putative CREB (cAMP-responsive element binding) transcription factor binding site in the BRCA1 promoter. Gelshift assays with methylated and unmethylated BRCA1/CREB binding site oligonucleotides demonstrate that this site is sensitive to site-specific CpG methylation. These data suggest that aberrant DNA methylation at regulatory sequences in the BRCA1 locus may play a role in the transcriptional inactivation of the BRCA1 gene within subclones of breast tumors. This study represents the first evidence suggesting a role for DNA methylation in the transcriptional inactivation of the BRCA1 in human breast cancer.

Topics: Base Sequence; Binding Sites; Breast Neoplasms; Cyclic AMP Response Element-Binding Protein; Dinucleoside Phosphates; DNA Methylation; Exons; Female; Genes, BRCA1; Humans; Models, Genetic; Oligodeoxyribonucleotides; Ovarian Neoplasms; Promoter Regions, Genetic; Reference Values; Regulatory Sequences, Nucleic Acid; Transcription, Genetic

Germline mutations of the BRCA2 gene on chromosome 13q12-q13 predispose to the development of early-onset breast cancer and ovarian cancer. Loss of heterozygosity detected using chromosome 13q markers in the vicinity of BRCA2 is observed in most cancers arising in carriers of germline BRCA2 mutations and also in 30-50% of sporadic breast and ovarian cancers. However, somatic mutations of BRCA2 are extremely rare in sporadic cancers. We have examined the hypothesis that expression of the BRCA2 gene may be suppressed in sporadic breast cancers by a mechanism that is associated with increased methylation of cytosine residues in the promoter region. Using a HpaII/MspI digestion-polymerase chain reaction based assay, the presence of 5-methylcytosine in three CpG dinucleotides within the BRCA2 promoter was assessed in 18 breast or ovarian cancer cell lines, in an SV40 large T antigen immortalized cell line derived from normal breast epithelial cells, in 64 primary sporadic breast cancers and peripheral blood leucocytes from these cases and in a number of other normal human tissues. Methylation was not detected in any of the tissues examined, suggesting that this mechanism of transcriptional repression is unlikely to explain the absence of somatic mutations in sporadic cancers.

Topics: Amino Acid Sequence; BRCA2 Protein; Breast Neoplasms; Dinucleoside Phosphates; DNA Methylation; Female; Genetic Markers; Humans; Molecular Sequence Data; Neoplasm Proteins; Ovarian Neoplasms; Promoter Regions, Genetic; Transcription Factors

Genomic imprinting and monoallelic gene expression appear to play a role in human genetic disease and tumorigenesis. The human H19 gene, at chromosome 11p15, has previously been shown to be monoallelically expressed. Since CpG methylation has been implicated in imprinting, we analyzed methylation of H19 DNA. In fetal and adult organs the transcriptionally silent H19 allele was extensively hypermethylated through the entire gene and its promoter, and, consistent with a functional role for DNA methylation, expression of an H19 promoter-reporter construct was inhibited by in vitro methylation. Gynogenetic ovarian teratomas were found to contain only hypomethylated H19 DNA, suggesting that the expressed H19 allele might be maternal. This was confirmed by analysis of 11p15 polymorphisms in a patient with Wilms tumor. The tumor had lost the maternal 11p15, and H19 expression in the normal kidney was exclusively from this allele. Imprinting of human H19 appears to be susceptible to tissue-specific modulation in somatic development; in one individual, cerebellar cells were found to express only the otherwise silent allele. Implications of these findings for the role of DNA methylation in imprinting and for H19 as a candidate imprinted tumor-suppressor gene are discussed.

Topics: Adult; Alleles; Base Sequence; Chromosomes, Human, Pair 11; Dinucleoside Phosphates; DNA, Neoplasm; Female; Fetus; Humans; Methylation; Molecular Sequence Data; Ovarian Neoplasms; Promoter Regions, Genetic; Teratoma; Wilms Tumor