Overexpression Of Brm Influences The Alternate Splicing Of HTERT In Human Gastric Cancer Cell Lines

OBJECTIVE:Regulation of telomerase activity primarily depends on transcriptional control of hTERT (human telomerase reverse transcriptase). The specific pattern of hTERT mRNA variants in human development provides evidence that alternative splicing is non-random and play a major role in the regulation of telomerase activity. The SWI/SNF (switch/sucrose non-fermentable) complex contributes to the regulation of gene expression by altering chromatin structures, and plays many important roles during epigenetic regulation in many organisms. Therefore, we aimed to demonstrate that Brm, catalytic ATPase subunit of the SWI/SNF complex, could modulate the alternate splicing patterns of hTERT mRNA, at least in part, contribute to provide the useful information for diagnoses and therapy the early stages of gastric tumor development.METHODS:Human gastric adenocarcinoma MGC-803 cells were transfected with hBrm cDNA expression vector (pCG-Brm-CMV). Then the expression levels of Brm mRNA and protein before and after transfection were detected by RT-PCR and Western-blot assay. Two alternative splicing sites (a,β) were selected, and the expression of 4 hTERT alternative splicing variants (ASVs) was analyzed in human gastric adenocarcinoma MGC-803 cells before and after transfection by half nested-PCR assay.RESULTS:1. Brm expression is frequently low in human gastric adenocarcinoma MGC-803 cells. After these cells were transfected with hBrm cDNA expression vector (pCG-Brm-CMV), Brm mRNA expression in the Brm-deficient gastric adenocarcinoma MGC-803 cells was significantly higher.2. After transfected with hBrm cDNA expression vector (pCG-Brm-CMV), Brm protein expression was significantly higher than before.3. a+β+ ASV expression was lower in the Brm-deficient gastric adenocarcinoma MGC-803 cells than that transfected with hBrm cDNA expression vector (pCG-Brm-CMV). a+β-ASV was not expressed after transfected with hBrm cDNA expression vector (pCG-Brm-CMV) and its positive rate was higher in the Brm-deficient gastric adenocarcinoma MGC-803 cells.4. a-β+ASV and a-β- ASV were not detected in the present study.CONCLUSION:The pattern of hTERT mRNA alternative splicing is different during multistep gastric carcinogenesis and play a major role in the regulation of telomerase activity. The a-deletion isoform appear to be a dominant inhibitor of telomerase activity when over-expressed. In particular, telomerase activity is down-regulated in many tissues by a shift to B-deletion splicing mode. Brm, catalytic subunits of the SWI/SNF chromatin remodeling complex, could modulate the splicing patterns of hTERT mRNA. It can rescue the expression of a+β+ASV and inhibit the expression of a+B"ASV. These findings suggest that Brm has significant tumor suppressive effects, particularly against gastric oncogenesis.