A highly conserved retinoic acid responsive element controlswt1aexpression in the zebrafish pronephros

Abstract

The Wilms' tumor suppressor gene Wt1 encodes a zinc-finger transcription factor that plays an essential role in organ development, most notably of the kidney. Despite its importance for organogenesis, knowledge of the regulation of Wt1 expression is scarce. Here, we have used transgenesis in zebrafish harboring two wt1 genes, wt1a and wt1b, in order to define regulatory elements that drive wt1 expression in the kidney. Stable transgenic lines with approximately 30 kb of the upstream genomic regions of wt1a or wt1b almost exactly recapitulated endogenous expression of the wt1 paralogs. In the case of wt1b, we have identified an enhancer that is located in the far upstream region that is necessary and sufficient for reporter gene expression in the pronephric glomeruli. Regarding wt1a, we could also identify an enhancer that is located approximately 4 kb upstream of the transcriptional start site that is required for expression in the intermediate mesoderm. Interestingly, this intermediate mesoderm enhancer is highly conserved between fish and mammals, is bound by members of the retinoic acid receptor family of transcription factors in gel shift experiments and mediates responsiveness to retinoic acid both in vivo and in cell culture. To our knowledge, this is the first functional demonstration of defined regulatory elements controlling Wt1 expression in vivo. The identification of kidney-specific enhancer elements will help us to better understand the integration of extracellular signals into intracellular networks in nephrogenesis.

A paradoxical teratogenic mechanism for retinoic acid

“…Previous studies have shown that Wt1 and Raldh2 are coexpressed in several tissues, including the metanephros, during embryogenesis (15,16). More recently, a RARE has been discovered in the zebrafish Wt1a, a homolog of the mammalian gene (18). The RARE consensus sequence is highly conserved among zebrafish, human, and mouse.…”

Murine Double Minute-2 Prevents p53-Overactivation-Related Cell Death (Podoptosis) of Podocytes

“…12 Therefore, we crossed the transparent zebrafish line Casper 33 (provided by Uwe Strähle, Karlruhe, Germany) with the wt1a:EGFP strain (provided by Christoph Englert, Jena, Germany) that expresses EGFP in podocytes. 34 Furthermore, we crossed the transgenic l-fabp:DBP-EGFP zebrafish line (provided by Anand-Apte) expressing DBP coupled to EGFP in the blood plasma 35 with the zebrafish strain Casper. This zebrafish line is referred to as CADE.…”

Two-Photon Microscopy Reveals Stationary Podocytes in Living Zebrafish Larvae

“…24 The wt1a:eGFP zebrafish strain 19 expresses eGFP under the wt1a promoter in podocytes and parietal epithelial cells (kindly provided by Dr. C. Englert, Jena, Germany). The translucent zebrafish strain casper 18 (kindly provided by Dr. U. Strähle, Karlsruhe, Germany) was crossed with the wt1a:eGFP strain.…”

“…For this purpose, the translucent zebrafish (mitfa w2/w2 ;roya 9/a9 [casper]) lacking black melanophores and reflecting iridophores 18 was intercrossed with the wt1a:eGFP transgenic zebrafish (Tg [-35.1wt1a:EGFP]), 19 resulting in mitfa w2/w2 ;roya 9/a9 ;Tg(235.1wt1a:eGFP). The zebrafish strain was abbreviated as ET (ET-EGFP/Translucent).…”