Project description:Adenosine-to-inosine RNA editing, a fundamental RNA modification, is regulated by adenosine deaminase (AD) domain containing proteins. Within the testis, RNA editing is catalyzed by ADARB1 and is regulated in a cell-type dependent manner. This study examined the role of two testis-specific AD domain proteins, ADAD1 and ADAD2, on testis RNA editing and male germ cell differentiation. ADAD1, previously shown to localize to round spermatids, and ADAD2 had distinct localization patterns with ADAD2 expressed predominantly in mid- to late-pachytene spermatocytes suggesting a role for both in meiotic and post-meiotic germ cell RNA editing. AD domain analysis showed the AD domain of both ADADs was likely catalytically inactive, similar to known negative regulators of RNA editing. To assess the impact of Adad mutation on male germ cell RNA editing, CRISPR-induced alleles of each were generated in mouse. Mutation of either Adad resulted in complete male sterility with Adad1 mutants displaying severe teratospermia and Adad2 mutant germ cells unable to progress beyond round spermatid. However, mutation of neither Adad1 nor Adad2 impacted RNA editing efficiency or site selection. Taken together, these results demonstrate ADAD1 and ADAD2 are essential regulators of male germ cell differentiation with molecular functions unrelated to A-to-I RNA editing.
Project description:To allow identification of differentially expressed genes and transposons in Dnmt3A mutants at 19dpp and 25dpp; mutant and wildtype RNA-Seq experiments were conducted starting from whole testis RNA.
Project description:Temporal study of the first wave of spermatogenesis in juvenile mouse testis and analysis of four germ cell deficient mouse models.<br> A series of purified testis cell-types (McCarrey libraries)constituting a focussed gene set was exploited to examine gene expression in prepubertal mouse testis. Testis RNA from C57/BL6J mice at various ages post partum was compared to a common control consisting of adult (8 week) testis RNA from the same strain(also corresponding to the last time-point).This generates a time course showing the activation of genes on the array across the first wave of spermatogenesis, which can then be correlated with the appearance of specific germ cell types, allowing assignation of unknown genes to differing cell types. <br> <br> Adult testis RNA from four different genetic models of infertility (XXSxrb, mshi, Bax -/-, bs) were compared to age- and strain-matched normal control testis RNA. These models possess different cellular complements within the testis, which can be interpreted within the framework established by the first wave analysis and used to refine the assignment of genes to cell types.
Project description:Spermatogenesis is a recurring differentiation process that results in the production of male gametes within the testes. During this process, spermatogonial stem cells differentiate to form spermatocytes, which undergo two rounds of meiotic division to form haploid spermatids. Throughout spermiogenesis, round spermatids elongate to form mature sperm. To profile maturing cell types, we generated bulk RNA-seq data from whole testis undergoing the first round of spermatogenesis between post-natal day P6 and P35. We also compared these libraries to adult samples.
Project description:Single cell RNA-seq (scRNA-seq) from Trim28 ovary knockout and wildtype mice ovaries and testis to help elucidate the function of Trim28 in the adult mouse ovaries. The analysis revealed that loss of Trim28 in the adult mouse ovaries lead to a transcriptional repogramming of the Granulosa cells towards the Sertoli cell fate. Therefore, Trim28 has a function to maintain the adult ovarian cell identity
Project description:We used whole larvae of both wildtype and Nxt1 trans-heterozygotes for RNA paired-end sequencing and obtained a read depth between 6.4M to 11.1M reads across all samples. In wandering larvae, stationary larvae and white prepupae, a total of 435, 855, and 611 genes were differentially expressed. A high percentage of down-regulated genes were highly expressed/testis-specific in the Drosophila testis.
Project description:Analysis of testis-specific over-expression of Musashi-1 and Musashi-2 at gene expression level. The hypothesis tested in the present study was that testis-specific over-expression of Musashi-1/Musashi-2 would result in differential gene expression profiles from the Wildtype control. Results provide important information of the genes affected by Musashi-2 overexpression during spermatogenesis. Total RNA obtained from whole adult testis of transgenic testis-specfic Musashi-1 and Musashi-2 overexpression mice compared to Wildtype (control) littermates. 4 replicates, 16 samples, 2 comparisons: Wt (Musashi-1) vs. TgMsi1 & Wt (Musashi-2) vs. TgMsi2
Project description:We precipitated dsRNA using the monoclonal J2 antibody and deep-sequenced the enriched samples from testis of juvenile dicer knock-out mice, age matched controls and adult animals. The dsRNA transcriptome is significantly less complex in juvenile mice as compared to adult controls and, possibly as a consequence, the knock-out of Dicer had only a minor effect of the total number of transcribed regions associated with dsRNA. The genes that potentially generate dsRNA are significantly expressed in isolated sperm cells with particular enrichment in pachytene spermatocytes.
Project description:We designed this experiment to investigate the transcriptional changes in gonads as a result of sex transformation. Here we performed transcriptional profiling of the ovary transformed into testis from the tra loss of function (XX_tra_lof), testis transformed into ovary from the tra gain of function (XY_tra_gof) and ovary transformed into testis in dsxM gain of function (XX_DsxM_gof/lof) Drosophila melanogaster third instar larvae in biological quadruplicates. In addition, as controls we sequenced ovaries and testes from the female and male wildtype larvae respectively. We constructed polyA+ libraries of the gonads, cleaned off the fatbody and performed 50 bp, stranded single-end RNA-Seq.