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  Gdiff


  Director : Mary C. Weiss (mweiss@pasteur.fr)


  abstract

 

The unit studies the mechanims implicated in the regulation of expression of hepatic genes and in the roles of the transcription factors that are necessary for the establishment and maintenance of hepatic differentiation. In addition, we establish new experimental models of liver cell differentiation and for the study of a key regulatory subunit of cAMP dependent protein kinase.



  report

cale

The Unit of Genetics of Differentiation uses cell genetics to analyse the roles of different Liver Enriched Transcription Factors (LETF) in the establishment and maintenance of the differentiated hepatic phenotype. It has been shown that in dedifferentiated rat hepatoma cells, the forced expression of HNF4a cDNA is sufficient to provoke the re-expression of genes that are considered markers of hepatic differentiation. In addition, in some cell lines of hepatic origin which fail to express either the LETF or hepatocyte differentiation, the forced expression of either HNF1a or of HNF4a cDNA is sufficient to ensure the expression of the endogenous genes for both factors, indicating the existence of a reciprocal regulatory loop between these two key transcription factors.

At present, our efforts are concentrated on the study of HNF4a, identified as a key factor for execution of the liver differentiation program. The HNF4a gene has two alternative promoters whose use is regulated during the course of liver development. The protein resulting from use during embryogenesis of the distal promoter, HNF4a7, shows avid transactivation in tranfection tests of genes that are expressed early during development. In contrast, HNF4a1 is produced in abundant fashion only at birth, and this protein shows strong activity on promoters of genes that are expressed neonatally. Thus, the timing of expression and the activity of the two HNF4a isoforms are complementary.

We have also analyzed an enhancer situated —3.5 kb upstream of the HNF4a1 promoter. It possesses functional binding sites for essentially all of the LETF, including HNF4a and HNF1a, as well as several GR half sites (glucocorticoid receptor). The site whose mutation provokes the most dramatic loss of activity binds C/EBPa. Thus, the properties of this enhancer are in perfect agreement with the expression pattern of the gene in the liver: weak expression during embyrogenesis, in agreement with sites that are weakly tranactivated by HNF3, HNF4a and HNF1, followed by robust expression at birth, explainable by the upregulation of C/EBPa and of circulating glucocorticoids.

Analysis of the expression of RIa, reputed to be a cytosolic regulatory subunit of cAMP dependent protein kinase (PKA), has permitted us to identify five alternative non-coding first exons and their cognate promoters. The different promoters are subject to both ubiquitous and tissue spécific expression. RIa is anchored at different sites in the cell, including the neruo-muscular junction (NMJ) and microtubules. Using the technique of intramuscular injection of plasmids encoding fusion proteins of RIa and green fluorescent protein, we have been able to identify three amino acids in the dimerization domain that are necessary for RIa localization to the NMJ. Finally, the over-expression of RIa is associated with high frequencies of abnormal mitoses, implying a role for PKA associated with microtubules in mitotoic progression.

Objectives

Activation of expression of the transcription factors enriched in the liver, in particular of HNF4. Filiation of liver cell types. Roles of R1a.



  publications

puce Publications of the unit on Pasteur's references database


  personnel

  Office staff Researchers Scientific trainees Other personnel
 

Papelard Solange, papelard@pasteur.fr

WEISS Mary C., IP, CNRS, mweiss @pasteur.fr

BAILLY Alain, INSERM, abailly@pasteur.fr

FAUST Daniela, IP, dfaust@pasteur.fr

HAYHURST Graham, CNRS, hayhurst@pasteur.fr

IMAIZUMI-SCHERRER Tereza, CNRS, mayumi@pasteur.fr

BRIANCON Nadège, Ph.D. student, nbrianco@pasteur.fr

STRICK Hélène, Ph.D. student, hstrick@pasteur.fr

TORRES PADILLA Maria Elena, Ph.D. student, mtorres@pasteur.fr

CATHERIN Anne-Marie, IP, acatheri@pasteur.fr

DESCHATRETTE Catherine, CNRS,Graduate engineer, cdeschat@pasteur.fr

MULET Céline, IP, Laboratory research support, cmulet@pasteur.fr


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