INVOLVEMENT OF HEPATOCYTE NUCLEAR FACTOR-3 IN ENDODERM DIFFERENTIATION OF EMBRYONIC STEM-CELLS

The transcription factors of the hepatocyte nuclear factor 3 (HNF3) fa mily, which are active in the liver, are expressed early during endode rm differentiation, To study their involvement in early murine develop ment, we examined their role in embryonic stem (ES) cells, HNF3 alpha or HNF3 beta mRNA transcripts were not detected in ES cells before dif ferentiation, and only low levels of HNF3 beta mRNA were detected at a late stage of differentiation of ES cells to embryoid bodies (EB) (20 days after induction of differentiation). To examine the consequences of overexpressing HNF3 alpha or -beta in ES cells, we transfected the two genes into these cells and determined the levels of expression of tissue-specific genes during EB differentiation, Specifically, we exa mined expression of albumin, cystic fibrosis transmembrane conductance regulator (CFTR), phosphoenolpyruvate carboxykinase (PEPCK), alpha 1- antitrypsin, transthyretin, zeta-globin, and neurofilament 68kd as mar kers for different cell lineages, Overexpression of HNF3 beta (and to a lesser extent of HNF3 alpha) induced the expression of genes associa ted with endodermal lineage, namely, the genes for CFTR and albumin, b ut did not induce the expression of genes involved in late endoderm di fferentiation, such as the genes for PEPCK and alpha 1-antitrypsin, Mo reover, expression of HNF1 beta was highly induced in HNF3-overexpress ing cells, while expression of HNF1 alpha and HNF4 was only mildly ind uced in these cells, Therefore, HNF3 alpha and -beta seem to be involv ed in early endoderm differentiation of ES cells and together with oth er developmental factors are apparently needed for the induction of th e endodermal lineage in vivo.