Extinction of α1-antitrypsin gene expression in somatic cell hybrids: Evidence for multiple controls

Abstract

Expression of the liver-specific α1,-antitrypsin (α1, AT) gene is extinguished in hepatoma/fibroblast hybrids. To define the mechanism of extinction, we identified DNA sequences involved in this process by transiently transfecting mutant α1AT promoters into parental and hybrid cells. The wild-type α1AT promoter (-554 to +44 bp) was highly expressed in rat hepatoma cells, but activity was 100-fold less in fibroblasts or cell hybrids. Mutations in this region failed to activate α1, AT expression in nonhepatic cells, but mutations in the binding site for liver factor B1 (LF-B1) reduced hepatic-specific expression >100-fold. Furthermore, the hybrid cells failed to express LF-B1-binding activity and mRNA. This suggested that α1AT extinction in hybrids might be an indirect, lack-of-activation phenotype mediated primarily through repression of LF-B1. To test this possibility, we stably transfected an LF-B1 expression cassette into parental and hybrid cells and monitored expression of transfected and endogenous α1AT genes. Surprisingly, although constitutive LF-B1 expression could activate α1AT-CAT transgenes in these cells, it neither prevented nor reversed extinction of the chromosomal α1AT genes. We conclude that although extinction of the LF-B1 trans-activator accompanies α1 AT extinction in cell hybrids, it does not play a causal role in this process.