112464

PRKCDBP

CAVIN3|HSRBC|SRBC|cavin-3;protein kinase C, delta binding protein;PRKCDBP;protein kinase C, delta binding protein

protein kinase C delta-binding protein|sdr-related gene product that binds to c-kinase|serum deprivation response factor-related gene product that binds to C-kinase

11p15.4

protein-coding

A cDNA clone encoding human SRBC [serum deprivation response factor (sdr)-related gene product that binds to c-kinase] was isolated in a yeast two-hybrid screening, with amino acids 1-304 of BRCA1 as the probe. The human SRBC gene (hSRBC) was mapped to chromosome region 11p15.5-p15.4, close to marker D11S1323, at which frequent loss of heterozygosity (LOH) has been observed in sporadic breast, lung, ovarian, and other types of adult cancers as well as childhood tumors. hSRBC-coding region mutations including frame shift and truncation mutations were detected in a few ovarian and lung cancer cell lines. More significantly, the expression of hSRBC protein was down-regulated in a large fraction [30 (70%) of 43] of breast, lung, and ovarian cancer cell lines, whereas strong expression of hSRBC protein was detected in normal mammary and lung epithelial cells. The down-regulation of hSRBC expression in cancer cells was associated with hypermethylation of CpG dinucleotides in its promoter region, and 3 (60%) of 5 primary breast tumors and 11 (79%) of 14 primary lung tumors were also found to be hypermethylated. Treatment of breast cancer MCF7 cells with 5azacytidine and Trichostatin A resulted in expression of hSRBC, confirming DNA methylation as the mode of inactivation. Our results suggest that epigenetic or mutational inactivation of hSRBC may contribute to the pathogenesis of several types of human cancers, marking hSRBC as a candidate tumor suppressor gene.

Recently, the human SRBC (hSRBC) gene, a candidate tumor suppressor gene (TSG), has been mapped to the chromosomal region 11p 15.5--p15.4 where frequent allele loss has been described in lung cancer. Aberrant methylation (referred to as methylation) of the promoter region of TSGs has been identified as an important mechanism for gene silencing. Loss of hSRBC protein expression occurs frequently in lung cancer cell lines and sodium bisulfite sequencing of the promoter region of hSRBC in several lung cancer cell lines suggested that methylation plays an important role in inactivating hSRBC. To determine the methylation status of hSRBC in a large collection of primary lung cancer samples, corresponding nonmalignant lung tissues and lung cancer cell lines (N=52), we designed primers for a methylation-specific PCR assay. Methylation was detected in 41% of primary non-small-cell lung cancers (NSCLC) (N=107) and in 80% of primary small-cell lung cancers (SCLC) (N=5), but was seen only in 4% of corresponding nonmalignant lung tissues (N=103). In all, 79% of lung cancer cell lines were methylated and the frequency of hSRBC methylation was significantly higher in SCLC (100%) than in NSCLC (58%) cell lines. Normal hSRBC protein expression was detected in only 18% of primary NSCLCs (N=93) by immunostaining and a significant association between loss of protein expression and methylation was found. hSRBC re-expression was observed after treatment of lung cancer cells with the demethylating agent 5-aza-2-deoxycytidine. In addition, 45% of the 76 hSRBC immunostaining-negative NSCLCs did not have hSRBC promoter methylation, indicating that other mechanisms of hSRBC expression silencing also exist. Both hSRBC immunostaining and methylation results did not correlate with clinicopathological characteristics of these patients. Our findings suggest that hSRBC is a candidate TSG involved in lung cancer pathogenesis, where expression is frequently inactivated by methylation and other mechanisms.

hSRBC is a putative tumor suppressor located at 11p15.4, at which frequent genomic loss has been observed in several human malignancies. To explore the candidacy of hSRBC as a suppressor of gastric tumorigenesis, we analyzed the expression and mutation status of hSRBC in gastric tissues and cell lines. hSRBC transcript was expressed in all normal and benign tumor tissues examined, but undetectable or very low in 73% (11/15) cancer cell lines and 41% (46/111) primary tumors. Loss or reduction of hSRBC expression was tumor-specific and correlated with stage and grade of tumors. While allelic loss or somatic mutations of the gene were infrequent, its expression was restored in tumor cells by 5-aza-2-deoxycytidine treatment and aberrant hypermethylation of 23 CpG sites in the promoter region showed a tight association with altered expression. Transient or stable expression of hSRBC led to a G(1) cell cycle arrest and apoptosis of tumor cells, and strongly suppresses colony forming ability and xenograft tumor growth. In addition, hSRBC elevated apoptotic sensitivity of tumor cells to genotoxic agents, such as 5-FU, etoposide and ultraviolet. Interestingly, hSRBC increased the protein stability of p53 and expression of p53 target genes, such as p21(Waf1), PUMA and NOXA, while hSRBC-mediated cell cycle arrest and apoptosis were abolished by blockade of p53 function. Our findings suggest that hSRBC is a novel tumor suppressor whose epigenetic inactivation contributes to the malignant progression of gastric tumors, in part, through attenuated p53 response to stresses.