A systematic analysis of the 3'UTR of HNF4A mRNA reveals an interplay of regulatory elements including miRNA target sites.

Dysfunction of hepatocyte nuclear factor 4α (HNF4α) has been linked to maturity onset diabetes of the young (MODY1), diabetes type II and possibly to renal cell carcinoma (RCC). Whereas diabetes causing mutations are well known, there are no HNF4A mutations found in RCC. Since so far analyses have been constricted to the promoter and open reading frame of HNF4A, we performed a systematic analysis of the human HNF4A 3'UTR.

The nephrogenic potential of the transcription factors osr1, osr2, hnf1b, lhx1 and pax8 assessed in Xenopus animal caps.

Background: The three distinct types of kidneys, pronephros, mesonephros and metanephros, develop consecutively in vertebrates. The earliest form of embryonic kidney, the pronephros, is derived from intermediate mesoderm and the first expressed genes localized in the pronephros anlage are the transcription factors osr1, osr2, hnf1b, lhx1 and pax8, here referred to as the early nephrogenic transcription factors. However, the pathway inducing nephrogenesis and the network of theses factors are poorly understood.

Improved conditional expression systems resulting in physiological level of HNF4alpha expression confirm HNF4alpha induced apoptosis in the pancreatic beta-cell line INS-1.

Background: To analyze gene function in mammalian cells tetracycline inducible expression of a gene-of-interest at a specific genomic location (Flp-In T-REx) is most attractive. However, leakiness of basal transgene expression and artificially high expression level upon tetracycline addition may be disadvantageous.

Findings: To solve these problems, we developed two different approaches to improve our pancreatic beta-cell line INS-1 Flp-In T-REx expressing the tissue restricted transcription factor HNF4alpha under control of tetracycline.

Heat-shock inducible Cre strains to study organogenesis in transgenic Xenopus laevis.

The frog Xenopus is a well established vertebrate model to study the processes involved in embryogenesis and organogenesis, as it can be manipulated easily with a whole series of methods. We have expanded these approaches by establishing two transgenic Xenopus strains that allow specific interference with the activity of defined genes using a heat-shock inducible Cre recombinase that can induce upon heat-shock expression of a reporter gene in crossings to a corresponding reporter strain. We have applied this binary technique of gene interference in Xenopus development to overexpress the mutated HNF1 beta transcription factor at distinct developmental stages.

Transcription factor HNF1beta and novel partners affect nephrogenesis.

Heterozygous mutations of the tissue-specific transcription factor hepatocyte nuclear factor (HNF)1beta, cause maturity onset diabetes of the young (MODY5) and kidney anomalies including agenesis, hypoplasia, dysplasia and cysts. Because of these renal anomalies, HNF1beta is classified as a CAKUT (congenital anomalies of the kidney and urinary tract) gene. We searched for human fetal kidney proteins interacting with the N-terminal region of HNF1beta using a bacterial two-hybrid system and identified five novel proteins along with the known partner DCoH.

Tissue-specific transcription factor HNF4alpha inhibits cell proliferation and induces apoptosis in the pancreatic INS-1 beta-cell line.

Hepatocyte nuclear factor 4alpha (HNF4alpha) is a tissue-specific transcription factor expressed in many cell types, including pancreatic beta-cells. Mutations in the HNF4alpha gene in humans give rise to maturity-onset diabetes of the young (MODY1) characterized by defective insulin secretion by beta-cells. To elucidate the mechanism underlying this disease, we introduced the splice form HNF4alpha2 or HNF4alpha8 into the rat beta-cell line INS-1.

A Kir6.2 mutation causing neonatal diabetes impairs electrical activity and insulin secretion from INS-1 beta-cells.

ATP-sensitive K(+) channels (K(ATP) channels) couple beta-cell metabolism to electrical activity and thereby play an essential role in the control of insulin secretion. Gain-of-function mutations in Kir6.2 (KCNJ11), the pore-forming subunit of this channel, cause neonatal diabetes.

Conditional expression of hepatocyte nuclear factor-1beta, the maturity-onset diabetes of the young-5 gene product, influences the viability and functional competence of pancreatic beta-cells.

Mutations in the gene encoding hepatocyte nuclear factor (HNF)1beta result in maturity-onset diabetes of the young-(MODY)5, by impairing insulin secretory responses and, possibly, by reducing beta-cell mass. The functional role of HNF1beta in normal beta-cells is poorly understood; therefore, in the present study, wild-type (WT) HNF1beta, or one of two naturally occurring MODY5 mutations (an activating mutation, P328L329del, or a dominant-negative form, A263insGG) were conditionally expressed in the pancreatic beta-cell line, insulin-1 (INS-1), and the functional consequences examined. Surprisingly, overexpression of the dominant-negative mutant did not modify any of the functional properties of the cells studied (including insulin secretion, cell growth and viability).

Identification of target genes of the transcription factor HNF1beta and HNF1alpha in a human embryonic kidney cell line.

Hepatocyte nuclear factor 1beta (HNF1beta, TCF2) is a tissue-specific transcription factor whose mutation in humans leads to renal cysts, genital malformations, pancreas atrophy and maturity onset diabetes of the young (MODY5). Furthermore, HNF1beta overexpression has been observed in clear cell cancer of the ovary. To identify potential HNF1beta target genes whose activity may be deregulated in human patients, we established a human embryonic kidney cell line (HEK293) expressing HNF1beta conditionally.

Pattern of genes influenced by conditional expression of the transcription factors HNF6, HNF4alpha and HNF1beta in a pancreatic beta-cell line.

Using the rat insulinoma cell line INS-1 we generated beta-cell clones that are most efficient for gene transfer, as they contain an FRT site for Flp recombinase-mediated, site-directed integration of a single copy transgene. Therefore, the gene-of-interest can be introduced by DNA transfection without the need to select individual cell clones. Additionally, the clones contain the tetracycline repressor allowing tetracycline induction of the transgene.

Tagging muscle cell lineages in development and tail regeneration using Cre recombinase in transgenic Xenopus.

The use of Cre and FLP recombinases to analyze embryogenesis and organogenesis in Xenopus has not been applied so far. We report on the generation of transgenic Xenopus animals containing a Cre-activated reporter gene cassette expressing blue fluorescent protein that can be switched over to yellow fluorescent protein expression upon Cre-mediated recombination. By injecting Cre mRNA into the two-cell stage embryo we show that Cre-mediated activation of the yellow fluorescent protein gene occurs.