ANALYSIS OF CONNEXIN43 EXPRESSION UNDER THE CONTROL OF A METALLOTHIONEIN PROMOTER

Transfection of C6 glioma cells with connexin43 (Cx43) cDNA under a co nstitutive promoter resulted in expression of Cx43 protein, an increas e in functional gap junctions, and reduced growth under in vitro and i n vivo conditions (D, Zhu et al., Proc. Natl, Acad. Sci. USA, 88: 1883 -1887, 1991). To allow for precise temporal and quantitative control o f Cx43 gene expression, the Cx43 cDNA was inserted into an expression vector [pSV(2)M(2)6] containing a modified metallothionein promoter. U pon transfection of this vector into C6 cells, clones were isolated th at expressed increased levels of inducible Cx43 protein and dye coupli ng, The level of induction of Cx43 expression increased with increasin g concentration of Zn2+, thus enabling the use of the same clone with different levels of gap junctions present. Although we observed no cha nge in cell growth under in vitro conditions following exposure to Zn2 + or Cd2+, clones with inducible expression of Cx43 were characterized by reduced growth in vivo. Within tumors, the level of expression of Cx43 mRNA and protein corresponded to that seen in vitro following the addition of Zn2+. The suppression of tumor growth in vivo correlated with the level of induced Cx43 expression.