Conserved Usage of Alternative 5′ Untranslated Exons of the GATA4 Gene

BACKGROUND:GATA4 is an essential transcription factor required for the development and function of multiple organs. Despite this important role, our knowledge of how the GATA4 gene is regulated remains limited. To better understand this regulation, we characterized the 5' region of the mouse, rat, and human GATA4 genes. METHODOLOGY/PRINCIPAL FINDINGS:Using 5' RACE, we identified novel transcription start sites in all three species. GATA4 is expressed as multiple transcripts with varying 5' ends encoded by alternative untranslated first exons. Two of these non-coding first exons are conserved between species: exon 1a located 3.5 kb upstream of the GATA4 ATG site in exon 2, and a second first exon (exon 1b) located 28 kb further upstream. Expression of both mRNA variants was found in all GATA4-expressing organs but with a preference for the exon 1a-containing transcript. The exception was the testis where exon 1a- and 1b-containing transcripts were similarly expressed. In some tissues such as the intestine, alternative transcript expression appears to be regionally regulated. Polysome analysis suggests that both mRNA variants contribute to GATA4 protein synthesis. CONCLUSIONS/SIGNIFICANCE:Taken together, our results indicate that the GATA4 gene closely resembles the other GATA family members in terms of gene structure where alternative first exon usage appears to be an important mechanism for regulating its tissue- and cell-specific expression

An Ebox Element in the Proximal Gata4 Promoter Is Required for Gata4 Expression In Vivo

GATA4 is an essential transcription factor required for the development and function of multiple tissues, including a major role in gonadogenesis. Despite its crucial role, the molecular mechanisms that regulate Gata4 expression in vivo remain poorly understood. We recently found that the Gata4 gene is expressed as multiple transcripts with distinct 5′ origins. These co-expressed alternative transcripts are generated by different non-coding first exons with transcripts E1a and E1b being the most prominent. Moreover, we previously showed that an Ebox element, located in Gata4 5′ flanking sequences upstream of exon 1a, is important for the promoter activity of these sequences in cell lines. To confirm the importance of this element in vivo, we generated and characterized Gata4 Ebox knockout mice. Quantitative PCR analyses realized on gonads, heart and liver at three developmental stages (embryonic, pre-pubertal and adult) revealed that the Ebox mutation leads to a robust and specific decrease (up to 89%) of Gata4 E1a transcript expression in all tissues and stages examined. However, a detailed characterization of the gonads revealed normal morphology and GATA4 protein levels in these mutants. Our qPCR data further indicate that this outcome is most likely due to the presence of Gata4 E1b mRNA, whose expression levels were not decreased by the Ebox mutation. In conclusion, our work clearly confirms the importance of the proximal Ebox element and suggests that adequate GATA4 protein expression is likely protected by a compensation mechanism between Gata4 E1a and E1b transcripts operating at the translational level

Characterization and Regulation of Epididymal 4-ene Steroid 5a-Reductase Messenger Ribonucleic Acids and Protein

Note:Tissue distribution, endocrine, developmental, and aging studies were used to characterize the regulation of the Sa-reductase mRNAs, types 1 and 2, in the rat epididymis. Sa-Reductase type 1 mRNA was predominantly localized to the initial segment of the epididymis. Bilateral orchidectomy markedly reduced type 1 m RNA levels; testosterone replacement could maintain these levels in all epididymal regions with the exception of the initial segment. Unilateral orchidectomy or efferent duct ligation caused a sharp decrease in type 1 mRNA concentrations but only in the initial segment of the epididymis. Sa-Reductase type 1 mRNA expression was developmentally regulated; age-dependent changes in type 1 mRNA expression were specific to the initial segment of the epididymis. […]Ce travail avait pour objectif de caractériser la distribution et la régulation des ARNms codant pour les isotypes 1 et 2 de la Sa-réductase dans l'epididyme du rat. Chez le rat adulte, I'ARNm de la Sa-réductase de type 1 a été principalement localise dans le segment initial de l'epididyme. L'orchiectomie bilatérale a considérablement réduit les quantités d'ARNm de type 1; à l'exception du segment initial de l'epididyme, le remplacement par la testostérone a pu maintenir l'expression de I'ARNm de la Sa-réductase de type 1. L'orchiectomie unilatérale ou la ligation des canaux efférents a entrainé une diminution très nette des quantités d'ARNm de la Sa-réductase de type 1 uniquement dans le segment initial de l'epididyme. […]