Reversible tumorigenesis induced by deficiency of vasodilator-stimulated phosphoprotein

Random homozygous knockout (RHKO) is an antisense RNA strategy capable of i dentifying genes whose homozygous functional inactivation yields a selectab le phenotype in cells growing in culture. Using this approach, we isolated NIH 3T3 fibroblast clones that showed the ability to form colonies on 0.5% agar and tumors in nude mice, The gene inactivated in one of these clones w as found to encode VASP (vasodilator-stimulated phosphoprotein), a previous ly identified protein that binds to components of the cadherin-catenin junc tional complex and has been implicated in cell-cell interactions, the forma tion of actin filaments, and the transmission of signals at the cytoskeleto n-membrane interface. Fibroblasts made deficient in VASP by RHKO showed los s of contact inhibition, and consequently, continued cell division past con fluence. Restoration of VASP function by reversal of RHKO yielded cells tha t had lost the neoplastic capabilities acquired during RHKO. Overproduction of VASP mRNA in the sense or antisense orientation from expression constru cts introduced by transfection into naive NM 3T3 fibroblasts also resulted in neoplastic transformation, implying that normal cell growth may require the maintenance of VASP expression within a narrow range, Our results impli cate VASP in tumorigenesis and/or cancer progression.