Definition of a high affinity growth hormone DNA response element

Rat liver contains a growth hormone inducible nuclear factor complex, GHINF , that binds to the growth hormone response element (GHRE) of the serine pr otease inhibitor (Spi) 2.1 gene. GHINF contains Stat5 and binds to paired g amma-activated sites (GAS) within the GHRE, but poorly to either one alone. By analysis of the sequence of various GAS sites that bind the GHINF compl ex (based on the GHRE 3' GAS motif), we demonstrate that a 13 nucleotide hi gh affinity DNA recognition sequence (haGHRE) for GHINF complex binding is ANTTC(C)/N-T(A)/(G)GAA(A)/(A)(T)/(T). One copy of the haGHRE will replace t he requirement for two GAS elements present in the wild type promoter in su pporting a GH response in primary hepatocyte culture. Mutation of the nativ e Spi 2.1 from a paired GAS site to a single haGHRE does not appreciably ch ange its affinity for binding to the GHINF complex, nor does it alter its s ensitivity to GH concentration. (C) 1999 Elsevier Science Ireland Ltd. All rights reserved.