Astudy on the mechanism of oral carcinoma progression upon the suppression of NF kB transcriptional activity by the nucleus-localization MALT1.

鳞状细胞癌是口腔最常见的恶性肿瘤，预后较差。然而，导致该病攻击行为的分子机制(S)仍不清楚。我们先前发现粘膜相关淋巴组织1(MALT1)在口腔癌中不表达。在本研究中，我们探讨了MALT1基因表达缺失的机制(S)及其对肿瘤细胞表型变化的影响。通过亚硫酸氢盐修饰序列、甲基化特异性聚合酶链式反应和5-氮杂氮去甲基化处理，分析了经常抑制多种抑癌基因表达的MALT1基因启动子甲基化状态。转录起始点的特定胞嘧啶被甲基化，而不是转录起始点的其他胞嘧啶，这种-256c甲基化是导致MALT1表达缺失的原因。为了了解MALT1失活对癌细胞表型的影响，我们利用野生型表达MALT1的癌细胞、表达显性阴性MALT1的癌细胞和MALT1基因特异性的siRNA，进行了生物学实验、体外侵袭实验、3D胶原凝胶侵袭实验、明胶酶谱分析、逃避实验、伤口愈合实验和小鼠肿瘤形成实验。野生型MALT1的外源表达降低了小鼠的增殖和细胞外基质降解活性、迁移行为和肿瘤生长。然而，显性负性MALT1的表达显著增强了这些细胞活性。此外，阻断内源和外源野生型MALT1的反义MALT1 siRNA的转染也增强了它们的表达。综上所述，这些结果有力地表明，MALT1特异性地抑制口腔癌细胞的侵袭性特征，MALT1基因的表观遗传失活是导致口腔癌进展和患者预后不良的主要原因之一。

Squamous cell carcinoma is a most frequent malignant neoplasm in the oral cavity with worse prognosis. However, a molecular mechanism(s) responsible for aggressive behaviors of the disease remains unknown. We previously found that mucosa-associated lymphoid tissue 1 (MALT1) is not expressed in oral carcinomas in a high frequency. In the present study, we examined the mechanism(s) of loss of the MALT1 expression and its effects on phenotypic alterations of carcinoma cells.Promoter methylation status of MALT1 gene, which frequently suppresses various tumor suppressor gene expression, was analyzed by bisulfite-modified sequence, methylation-specific PCR and demethylation treatment by 5-aza. Specific cytosine at -256 bp but not other cytosines from the transcription start site was methylated, and this -256C methylation was responsible for loss of the MALT1 expression. To understand a role of MALT1 inactivation on carcinoma cell phenotypes, we used wild-type MALT1-expressing carcinoma cells, dominant-negative MALT1-expressing carcinoma cells and siRNA specific to MALT1 gene, and performed biological assays ; in vitro invasion assay, 3D collagen gel invasion assay, gelatin zymography, evasion assay, wound healing assay and tumor formation assay in mice. Exogenous expression of wild-type MALT1 reduced proliferation and extracellular matrix-degrading activities, migratory behavior and tumor growth in mice. However, expression of dominant-negative MALT1 dramatically enhanced these cellular activities. In addition, transfection of anti-MALT1 siRNA, which blocks endogenous and exogenous wild-type MALT1, also enhanced them. Altogether, these results strongly suggest that MALT1 specifically suppresses aggressive features of oral carcinoma cells and that epigenetic inactivation of MALT1 gene is one of major causative resulting in carcinoma progression and worse prognosis of patients suffering from the disease.