INDUCTION OF CELL-TRANSFORMATION BY MUTATED 16K VACUOLAR H-ATPASE (DUCTIN) IS ACCOMPANIED BY DOWN-REGULATION OF GAP JUNCTIONAL INTERCELLULAR COMMUNICATION AND TRANSLOCATION OF CONNEXIN-43 IN NIH3T3 CELLS()

The 16 K subunit of the vacuolar H+-ATPase (ductin) has been suggested to also play a role in gap junction channels. Since mutated 16 K subu nits have transforming ability when transfected into NIH3T3 cells and since aberrant gap junctional intercellular communication (GJIC) is a hallmark of cancer cells, we hypothesized that mutated 16 K subunits m ight transform these cells via alteration of GJIC, When GJIC was measu red by the dye-transfer assay, NIH3T3 cells transfected with the mutan t 16 K protein genes (deletion of the fourth transmembrane domain or a point mutation at codon 143 from glutamic acid to arginine) showed si gnificantly lower levels of GJIC than those transfected with the vecto r alone or with the wild-type 16 K subunit gene. GJIC levels of NIH3T3 cells transformed by v-ras and v-src were not significantly decreased , suggesting that low GJIC levels are not necessarily the result of ce ll transformation pel se. NIH3T3 cells express Cx 43 as a major connex in gene. Although cells transfected with mutated 16 K subunits showed a level of Cx 43 protein expression similar to non-transfectants, thei r Cx43 protein was localized aberrantly, i.e. intracytoplasmically, Th ese results indicate that mutant 16 K subunits with transforming abili ty translocate Cx43 proteins, thus inhibiting GJIC of NIH3T3 cells.