The orphan homeobox gene Hex was first identified in hematopoetic cells. We have shown that Hex has a limited domain of expression in the developing mouse which includes the liver and biliary tree (Bogue, et al; Ped Res; 1995, p336). By in situ hybridization, it is highly expressed in the liver at E9.5 and in the gall bladder at E10.5, the earliest stages at which these organs can be identified as distinct structures. At E12.5, liver parenchymal expression persists, but at this age it is higher in epithelial cells of ducts in the region of the developing bile and pancreatic ducts. Northern analysis showed that Hex expression persists in the liver into adult life. Furthermore, in situ analysis of adult liver showed a homogeneous pattern of expression. We began to evaluate a potential late, liver-specific function for Hex. To determine which cell types in the liver express Hex, we separately isolated adult rat hepatocytes and cholangiocytes and performed RT-PCR using primers specific for the mouse cDNA sequence. Both rat hepatocytes and cholangiocytes expressed Hex. Additionally, RT-PCR with primers specific for human HEX demonstrated expression in human liver and HepG2 cells. In order to determine if Hex protein can trans-activate a developmentally-regulated liver-specific promoter, we co-transfected a CMV-driven mouse Hex cDNA construct(pCMVHex) with a luciferase reporter plasmid driven by the rat NTCP promoter(JBC 271:15211, 1996). Activity in HepG2 cells of a 1.3 kb 5'-upstream fragment (nt -1237/+47) from the rat NTCP gene promoter was enhanced 3-5 fold by co-transfected pCMVHex. Co-transfection with a plasmid consisting of vector alone did not alter reporter gene activity. Moreover, pCMVHex did not affect activity of an SV40 promoter-driven luciferase construct. Finally, 5'-deletion analysis of the rat NTCP promoter localized a Hex-response element between-758 and -158 nt from the start of NTCP transcription. We conclude that regulation of NTCP transcription is a potential liver-specific function of Hex. Experiments are ongoing to finely map the Hex response element in the NTCP promoter and to better define the role of Hex in liver development.