Cloning and DNA-binding properties of the rat pancreatic beta-cell-specific factor Nkx6.1

The homeodomain (HD) protein Nkx6.1 is the most beta-cell-specific transcri ption factor known in the pancreas and its function is critical for the for mation of the insulin-producing beta-cells. However, the target genes, DNA- binding site, and transcriptional properties of Nkx6.1 are unknown. Using i n vitro binding site selection we have identified the DNA sequence of the N kx6.1 binding site to be TTAATTG/A. A reporter plasmid containing four copi es of this sequence is activated by an Nkx6.1HD/VP16 fusion construct. Full -length Nkx6.1 fails to activate this reporter plasmid in spite of robust i nteraction with the binding site in vitro. Stable expression of Nkx6.1 in t he glucagon-producing alpha-cell-like MSL-G-AN cells induces expression of the endogenous insulin gene in a subset of the cell population. The express ion of other known beta-cell-specific factors such as Pax4, Pax6, Pdx1, GLU T2 and GLP1-R is unchanged by the introduction of Nkx6.1. (C) 1999 Federati on of European Biochemical Societies.