A cells fate as well as its differentiative state and phenotype are highly dependent on signals from the extracellular matrix. The liver, and hepatocyte cultures in particular, represent and appropriate and convenient model system to study cellular growth and differentiation since several signaling pathways and liver-enriched transcription factors have been identified which acre essential to initiation and maintenance of liver differentia6on/development. Hepatic growth and differentiation is mediated primarily by the combinatorial actions of liver-enriched transcription factors, foremost of which is HNF-4.. To more clearly understand the molecular mechanisms by which extracellular matrix proteins regulate growth, regeneration and differentiation of hepatocytes, this study aims to investigate the matrixspecific alterations in HNF-4 expression, DNA binding, and transcriptional activity in differentiating hepatocytes. Specifically, this study will look at (1) the mRNA and protein expression profiles, during differentiation, of the various HNF-4 isoforms as well as other factors known to modulate HNF4 activity through Northern, RT-PCR and Western analysis. Likewise, (2) the DNA binding activities of these transcription factors will be assessed through mobility shift and reporter gene assays. Additionally, (3) the functionality of these factors in promoting and/or inhibiting differentiation will be assessed via transfection assays utilizing reporter genes, antisense oligonucleotides, plasmids over- expressing wild type or mutant factors. Lastly, we will investigate specific matrix components and signaling cascades responsible for matrix-induced differentiation. The information derived from the proposed studies will heighten our understanding of the role of extracellular matrix in regulation of cellular differentiation involved in development, regeneration, and cancer.
