Tissue-specific RNAi reveals that WT1 expression in nurse cells controls germ cell survival and spermatogenesis

Abstract

Using a novel tissue-specific RNA interference (RNAi) approach that mimics the principle by which naturally occurring microRNAs (miRNA) are made, we demonstrate that the Wilms' tumor 1 (WT1) transcription factor has an essential role in spermatogenesis. Mice depleted of WT1 in Sertoli nurse cells suffered from increased germ cell apoptosis, loss of adherens junctions, disregulation of adherence junction-associated genes, and impaired fertility. These effects were recapitulated in transgenic mice expressing a dominant-negative form of WT1 in Sertoli cells, demonstrating the validity of our RNAi approach. Our results indicate that the tumor suppressor WT1 promotes Sertoli cell-germ cell signaling events driving spermatogenesis.

Incentive Learning Underlying Cocaine-Seeking Requires mGluR5 Receptors Located on Dopamine D1 Receptor-Expressing Neurons

“…A similar approach has been reported previously for interference with other genes in nurse cells (Rao et al, 2006) and, together with a very recent report (Garbett et al, 2010), our findings show that this technique can be used successfully in the brain. Compared with conditional gene deletion this approach has the advantage that it involves only one mouse line and offers the perspective to be used, in modified forms, in other organisms in which targeted mutagenesis is not feasible.…”

Multi‐gene engineering: Simultaneous expression and knockdown of six genes off a single platform

“…For genetic function analysis, multi-gene constructs could be used to investigate combinatorial effects, to determine the presence of functional redundancy, or the degree of synergistic or antagonistic interplay between molecular components (Chung et al, 2006;Rao et al, 2006;Xia et al, 2006). In addition, the ability to co-ordinate transgene expression with gene silencing also renders the technology useful for therapeutic applications which may require the silencing (via intronic siRNA) of a faulty gene or allele followed by substitution with a wild-type or functional gene (via exon transgene expression) (Samakoglu et al, 2006).…”

Transcription and post-transcriptional regulation of spermatogenesis

“…RNA expression profiling and follow-up transfection experiments identified several genes misregulated in response to perturbed WT1 in adult Sertoli cells (Rao et al 2006). Among these were Eps8 and Icap1-a, which encode crucial signalling molecules at the apical ES.…”

“…WT1 expression persists in Sertoli cells after gonad formation, but until recently, its functional role in this context was not known, as Wt1-mutant mice die early during embryogenesis. The role of WT1 in Sertoli cells after the bipotential gonad stage was recently addressed by two studies (Gao et al 2006;Rao et al 2006). Gao et al (2006) used a conditional knockout approach to disrupt the Wt1 gene in foetal Sertoli cells at approximately e14.5 and found that this severely disrupted testis cord formation in male embryos and subsequently led to the formation of abnormal tubular architecture and reduced testes size in adult males.…”

“…This result clearly demonstrated that WT1 is crucial for developmental events that convert the bipotential gonad into a male gonad. Rao et al (2006) used an RNA interference approach to address whether WT1 also has a role in spermatogenesis (Rao et al 2006). Using a Sertoli cell promoter (from the Rhox5 homeobox gene; see below) to express an artificial microRNA (miRNA) targeting WT1 in postnatal and adult (but not foetal) Sertoli cells, they found that this led to disruption of the apical ectoplasmic specialization (ES), a junctional complex formed between Sertoli cells and elongating/elongated spermatids.…”