Gelsolin functions as a metasatsis suppressor in B16-BL6 mouse melanoma cells and requirement of the carboxyl-terminus for its effect

Gelsolin, an actin-binding protein, is implicated as a critical regulator i n cell motility. In addition, we have reported that cellular levels of gels olin are decreased in various tumor cells, and overexpression of gelsolin b y gene transfer suppresses tumorigenicity. We sought to assess the effects of gelsolin overexpression on metastasis and to determine the Importance of a carboxyl-terminus that confers Ca2+ dependency on gelsolin for effects o f its overexpression. Expression vectors with cDNA encoding either full-len gth wild-type or His321 mutant form, isolated from a flat revertant of Ras- transformed cells and a carboxyl-terminal truncate, C-del of gelsolin, were transfected into a highly metastatic murine melanoma cell line, B16-BL6. E xpression of Introduced cDNA in transfectants was confirmed using Western b lotting, 2-dimensional gel electrophoresis and reverse transcription-polyme rase chain reaction (RT-PCR). We characterized phenotypes of transfectants, such as growth rate, colony formation in soft agar, cell motility and meta stasis formation in vivo. Transfectants expressing the wild-type, His321 mu tant and C-del gelsolin exhibited reduced growth ability in soft agar. Alth ough expression of integrin beta1 or alpha4 on the cell surface of transfec tants was not changed, wild-type and His321 mutant gelsolin, except for C-d el gelsolin, exhibited retardation of cell spreading, reduced chemotatic mi gration to fibronectin and suppressed lung colonization in spontaneous meta stasis assay. Gelsolin may function as a metastasis suppressor as well as a tumor suppressor gene. The carboxyl-terminus of gelsolin is important for retardation of cell spreading, reduced chemotasis and metastasis suppressio n. (C) 2001 Wiley-Liss, Inc.