Adenovirus-mediated increase in HNF-3β or HNF-3α shows differences in levels of liver glycogen and gene expression

Abstract

We previously generated a transgenic mouse line (T-77) in which increased hepatic expression of the hepatocyte nuclear factor-3␤ (HNF-3␤) protein was used to assess its role in hepatocyte-specific gene transcription. The T-77 transgenic mice displayed elevated serum bile acid and bilirubin levels and a complete absence of hepatic glycogen storage. These postnatal liver defects were associated with diminished expression of hepatocyte genes involved in gluconeogenesis and bile acid transport as well as reduced levels of hepatocyte transcription factors. In this study, we show that mouse tail vein injections of adenovirus expressing the rat HNF-3␤ (AdHNF3␤) cDNA efficiently increased its levels throughout the liver lobule and recapitulated the T-77 transgenic liver phenotype within several days postinfection. Likewise, the AdHNF3␤-infected liver phenotype was associated with reduced hepatic expression of genes involved in glucose homeostasis, bile acid transport, and bilirubin conjugation, which were not found with control adenovirus infections. These studies show that adenovirus-mediated gene transfer is an effective method for rapid hepatic increases in transcription factor levels to determine in vivo target genes. In contrast, AdHNF3␣-infected liver displayed only a transient reduction in hepatic glycogen levels and was associated with less severe decreases in hepatic expression of gluconeogenic and bilirubin metabolism genes. Consistent with these findings, only T-77 transgenic and AdHNF3␤-infected liver exhibited diminished hepatic expression of the HNF-6 transcription factor, suggesting that reduced HNF-6 levels contribute to diminished HNF-3␤-specific transcriptional activity. T he liver performs essential functions in the body by transcriptionally regulating expression of hepatocyte-specific genes encoding plasma proteins and enzymes involved in glucose metabolism, gluconeogenesis and glycogen storage, and cholesterol and bile acid homeostasis. 1,2 Functional analysis of numerous hepatocyte-specific regulatory regions reveals that they are composed of multiple cis-acting DNA sequences that bind different families of hepatocyte nuclear factors (HNF) genes. 3,4 Although none of these transcription factors is entirely liver specific, the requirement for combinatorial protein interactions among them to achieve high transcriptional levels plays an important role in maintaining hepatocyte-specific gene expression. Isolation of the cDNA clones encoding these HNF proteins facilitated the identification of their DNA binding and transcriptional activation domains. On the basis of homology within the DNA binding domains, the transcription factors were grouped into related protein families. These include the Pou-Homeodomain HNF-1, the winged helix containing Forkhead Box (Fox) HNF-3, the steroid hormone receptor HNF-4, the basic leucine zipper CCAAT/enhancer binding protein (C/EBP), and the One-Cut Homeodomain HNF-6 families. 3,4 The Fox transcription factors 5 are an extensive protein family that shares homology in th...

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“…The construction and preparation of the replication-defective AdHNF-6 or AdLacZ have been described previously. 11,15,16 Ribonuclease Protection Assay. Total RNA was prepared from whole mouse liver using RNA-STAT-60 (TelTest "B" Inc., Friendswood, TX).…”

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Experimental Models of Transcription Factor-Associated Maturity-Onset Diabetes of the Young

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