Defects in pulmonary vasculature and perinatal lung hemorrhage in mice heterozygous null for the Forkhead Box f1 transcription factor.

Decreased pulmonary expression of Forkhead Box f1 (Foxf1) transcription factor was associated with lethal alveolar hemorrhage in 55% of the Foxf1 +/- newborn mice. The severity of the pulmonary abnormalities correlates with the levels of Foxf1 mRNA. Defects in alveolarization and vasculogenesis were observed in subsets of the Foxf1 +/- mice with relatively low levels of expression from the normal Foxf1 allele. Lung hemorrhage was coincident with disruption of the mesenchymal-epithelial cell interfaces in the alveolar and bronchiolar regions of the lung parenchyma and was associated with increased apoptosis and reduced surfactant protein B (SP-B) expression. Finally, the lung defect associated with the Foxf1 +/- mutation was accompanied by reduced expression of vascular endothelial growth factor (VEGF), the VEGF receptor 2 (Flk-1), bone morphogenetic protein 4 (Bmp-4), and the transcription factors of the Brachyury T-Box family (Tbx2-Tbx5) and Lung Kruppel-like Factor. Reduction in the level of Foxf1 caused neonatal pulmonary hemorrhage and abnormalities in alveologenesis, implicating this transcription factor in the regulation of mesenchyme-epithelial interaction critical for lung morphogenesis.

Elevated levels of hepatocyte nuclear factor 3beta in mouse hepatocytes influence expression of genes involved in bile acid and glucose homeostasis.

The winged helix transcription factor, hepatocyte nuclear factor-3beta (HNF-3beta), mediates the hepatocyte-specific transcription of numerous genes important for liver function. However, the in vivo role of HNF-3beta in regulating these genes remains unknown because homozygous null HNF3beta mouse embryos die in utero prior to liver formation. In order to examine the regulatory function of HNF-3beta, we created transgenic mice in which the -3-kb transthyretin promoter functions to increase hepatocyte expression of the rat HNF-3beta protein. Postnatal transgenic mice exhibit growth retardation, depletion of hepatocyte glycogen storage, and elevated levels of bile acids in serum. The retarded growth phenotype is likely due to a 20-fold increase in hepatic expression of insulin-like growth factor binding protein 1 (IGFBP-1), which results in elevated levels in serum of IGFBP-1 and limits the biological availability of IGFs required for postnatal growth. The defects in glycogen storage and serum bile acids coincide with diminished postnatal expression of hepatocyte genes involved in gluconeogenesis (phosphoenolpyruvate carboxykinase and glycogen synthase) and sinusoidal bile acid uptake (Ntcp), respectively. These changes in gene transcription may result from the disruptive effect of HNF-3beta on the hepatic expression of the endogenous mouse HNF-3alpha,-3beta, -3gamma, and -6 transcription factors. Furthermore, adult transgenic livers lack expression of the canalicular phospholipid transporter, mdr2, which is consistent with ultrastructure evidence of damage to transgenic hepatocytes and bile canaliculi. These transgenic studies represent the first in vivo demonstration that the HNF-3beta transcriptional network regulates expression of hepatocyte-specific genes required for bile acid and glucose homeostasis, as well as postnatal growth.

The nuclear receptor fetoprotein transcription factor is coexpressed with its target gene HNF-3beta in the developing murine liver, intestine and pancreas.

During organogenesis, the winged helix hepatocyte nuclear factor 3beta (HNF-3beta) protein participates in regulating gene transcription in the developing esophagus, trachea, liver, lung, pancreas, and intestine. Hepatoma cell transfection studies identified a critical HNF-3beta promoter factor, named UF2-H3beta, and here, we demonstrate that UF2-H3beta is identical to the fetoprotein transcription factor (FTF). In situ hybridization studies of mouse embryos demonstrate that FTF expression initiates in the foregut endoderm during liver and pancreatic morphogenesis (day 9) and that earlier expression of FTF is observed in the yolk sac endoderm, branchial arch and neural crest cells (day 8). Abundant FTF hybridization signals are observed throughout morphogenesis of the liver, pancreas, and intestine and its expression continues in the epithelial cells of these adult organs. In day 17 mouse embryos and adult pancreas, however, expression of FTF becomes restricted to the exocrine acinar and ductal epithelial cells.

In situ hybridization with 33P-labeled RNA probes for determination of cellular expression patterns of liver transcription factors in mouse embryos.

Murine hepatocyte nuclear factor-3beta (HNF-3beta) protein is a member of a large family of developmentally regulated transcription factors that share homology in the winged helix/fork head DNA binding domain and that participate in embryonic pattern formation. HNF-3beta also mediates cell-specific transcription of genes important for the function of hepatocytes, intestinal and bronchiolar epithelium, and pancreatic acinar cells. We have previously identified a hepatocyte and pancreatic cut-homeodomain transcription factor, HNF-6, which is required for HNF-3beta promoter activity. In this study, we used in situ hybridization studies of stage-specific embryos to demonstrate that HNF-6 and its target gene, HNF-3beta, are coexpressed in the foregut endoderm and in the pancreatic and hepatic diverticulum. More detailed analysis of HNF-6 and HNF-3beta's developmental expression patterns provides evidence of colocalization in hepatocytes, intestinal epithelium, and pancreatic ductal epithelium and exocrine acinar cells. In support of the role of HNF-6 in regulating HNF-3beta expression in developing hepatocytes, their liver expression levels are both transiently reduced between 14 and 15 days of gestation. At day 18 of gestation and in adult pancreas, HNF-6 and HNF-3beta transcripts remain colocalized in the exocrine acinar cells, but their expression patterns diverge in endocrine cells. HNF-3beta expression is restricted to the endocrine cells of the islets of Langerhans, whereas the ductal epithelium expresses HNF-6. We discuss these expression patterns with respect to specification of hepatocytes and differentiation of the endocrine and exocrine pancreas.

Two naturally occurring alpha2,6-sialyltransferase forms with a single amino acid change in the catalytic domain differ in their catalytic activity and proteolytic processing.

The alpha2,6-sialyltransferase (ST) is a Golgi glycosyltransferase that adds sialic acid residues to glycoprotein N-linked oligosaccharides. Here we show that two forms of alpha2,6-sialyltransferase are expressed by the liver and are encoded by two different RNAs that differ by a single nucleotide. The ST tyr possesses a Tyr at amino acid 123, whereas the ST cys possesses a Cys at this position. The ST tyr is more catalytically active than the ST cys; however, both are functional when introduced into tissue culture cells. The proteolytic processing and turnover of the ST tyr and ST cys proteins differ dramatically. The ST cys is retained intact in COS-1 cells, whereas the ST tyr is rapidly cleaved and secreted. Analysis of the N-linked oligosaccharides of these proteins demonstrates that both proteins enter the late Golgi. However, differences in ST tyr and ST cys proteolytic processing may be related to differences in their localization, because ST tyr but not ST cys is expressed at low levels on the cell surface. The possibility that the ST tyr is cleaved in a post-Golgi compartment is supported by the observation that a 20 degrees C temperature block, which stops protein transport in the trans Golgi network, blocks both cleavage and secretion of the ST tyr.