Moreover, enhanced SPTAN1 was linked to tumor progression and malignancy in ovarian cancer and described to be involved in the carcinogenesis of sporadic CRC. Upregulated SPTAN1, e.g., was demonstrated in various types of tumors and shown to be associated with local aggressiveness and metastic behavior of soft tissue carcinomas. Deregulation of spectrins, especially of SPTAN1 seriously affects cellular behavior and promotes tumor progression.
Spectrins contribute to cell adhesion and migration, interact with structural and regulatory proteins and are involved in the regulation of DNA repair. SPTAN1 belongs to a superfamily of F-actin cross-linking proteins (scaffolding proteins) which, first identified as membrane-skeleton components in erythrocytes, are ubiquitously expressed in metazoan cells. Amongst them we identified non-erythroid spectrin αII (SPTAN1) as a novel interaction partner of MLH1 and found evidence for the involvement of both proteins in cytoskeletal and filamental organization. Several MLH1 interacting proteins have been published, which might be essential for signaling DNA damages to different cellular processes. Besides, recent studies suggest that MLH1 also participates in other important fundamental cellular functions beyond its primary role in MMR, e.g., the regulation of cell cycle checkpoints and apoptosis, but also in meiotic reciprocal recombination and meiotic mismatch repair. Looking at functionality, MutLα is mainly involved in the correction of base-base mismatches and insertion-deletion loops resulting from defective DNA replication. In addition, 13-15% of sporadic CRCs are caused by MLH1 deficiency based on somatic promotor hypermethylation. Germline mutations in MLH1 are responsible for 50% of a hereditary form of colorectal cancer (CRC) called Lynch syndrome. MLH1 is the main component of the heterodimer MutLα, formed by MLH1 and PMS2. The most important DNA mismatch repair (MMR) protein commonly dysregulated in colon cancer is MLH1. Therefore, aggressiveness of MLH1-positive CRC might be related to SPTAN1. These data suggest that SPTAN1 levels decreased in concordance with MLH1 reduction and impaired cellular mobility in MLH1 deficient colon cancer cells. Consequently, overexpression of SPTAN1 increased migration of MLH1 deficient cells while knock down of SPTAN1 decreased cellular mobility of MLH1 proficient cells, indicating SPTAN1-dependent migration ability. In addition, cellular motility of MLH1 deficient HCT116 and MLH1 deficient HEK293T cells was impaired compared to the MLH1 proficient sister clones. Moreover, siRNA knock down of MLH1 decreased the mRNA level of SPTAN1 in HeLa, HEK293 as well as in MLH1 positive HCT116 cells, which indicates a co-expression of SPTAN1 by MLH1. MLH1 deficiency is clearly associated with SPTAN1 reduction. Effects on cellular motility were determined in MLH1 deficient HCT116 and MLH1 deficient HEK293T compared to their MLH1 proficient sister cells, respectively. Co-expression of SPTAN1 and MLH1 was analyzed by siRNA knock down of MLH1 in HeLa, HEK293, MLH1 positive HCT116, SW480 and LoVo cells.
Nine cancer cell lines as well as fresh and paraffin embedded colon cancer tissue from 12 patients were used in gene expression studies of SPTAN1 and MLH1. In the current study, the interaction of MLH1 and SPTAN1 and its potential consequences for CRC metastasis was evaluated. We recently identified non-erythroid spectrin αII (SPTAN1), a scaffolding protein involved in cell adhesion and motility, to interact with MLH1. Although MLH1 has been shown to be not only involved in postreplicative MMR but also in several MMR independent processes like cytoskeletal organization, the connection between MLH1 and metastasis remains unclear. Affected tumors generate much less metastatic potential than the MLH1 proficient forms. Defects in the DNA mismatch repair (MMR) protein MLH1 are frequently observed in sporadic and hereditary colorectal cancers (CRC).
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