Gap junction proteins connexin32 and connexin43 partially acquire growth-suppressive function in HeLa cells by deletion of their C-terminal tails

Our laboratory has previously reported that transfection of a connexin26 (C x26) gene, but not connexin40 nor connexin43 (Cx43), into HeLa cells expres sing no detectable level of connexins suppressed the tumorigenic phenotype of the HeLa cells both in vivo and in vivo, although all of these connexins induced gap junctional intercellular communication in HeLa cells to a simi lar extent. The most remarkable structural difference between connexin prot eins is the length of the C-terminal cytoplasmic tail, Cx26 having almost n o tail, while Cx43 and connexin32 (Cx32) have long and intermediate ones, r espectively, When Cx32 and Cx43 lose their C-terminal tails, they seem to r esemble Cx26 in structure. To examine whether such truncated connexins beco me tumor suppressive in HeLa cells, we introduced a stop codon into each of the Cx32 and Cx43 cDNAs to remove their C-terminal tails and transfected t hese constructs (Delta Cx) into HeLa cells, Both Delta Cx cDNAs induced GJI C as efficiently as the wild-type counterparts, Although none of the trunca ted connexins affected proliferation rate, the truncated Cx32 and Cx43 prot eins suppressed anchorage-independent cell growth in soft agar, Furthermore , when the transfectants were injected into the backs of nude mice, tumor a ppearance was delayed by 7 days in animals given cells expressing truncated connexins, i.e. tumors became detectable on days 11 and 18 after injection of vector and Delta Cx transfectants, respectively. Although throughout th ese experiments the truncated connexins did not completely eliminate the tu morigenicity of HeLa cells, as Cx26 did, it was evident that deletion of th e C-terminal tails gave both Cx32 and Cx43 a capacity for negative growth c ontrol, suggesting that the C-terminal tails of these two connexins functio n as a regulatory region for connexin-mediated growth control in HeLa cells .