Dynamics of notch pathway expression during mouse testis post-natal development and along the spermatogenic cycle

Articles in International JournalsThe transcription and expression patterns of Notch pathway components (Notch 1–3, Delta1 and 4, Jagged1) and effectors (Hes1, Hes2, Hes5 and Nrarp) were evaluated (through RT-PCR and IHC) in the mouse testis at key moments of post-natal development, and along the adult spermatogenic cycle. Notch pathway components and effectors are transcribed in the testis and expressed in germ, Sertoli and Leydig cells, and each Notch component shows a specific cell-type and timewindow expression pattern. This expression at key testis developmental events prompt for a role of Notch signaling in prepubertal spermatogonia quiescence, onset of spermatogenesis, and regulation of the spermatogenic cycle

Dicer1 Depletion in Male Germ Cells Leads to Infertility Due to Cumulative Meiotic and Spermiogenic Defects

Background: Spermatogenesis is a complex biological process that requires a highly specialized control of gene expression. In the past decade, small non-coding RNAs have emerged as critical regulators of gene expression both at the transcriptional and post-transcriptional level. DICER1, an RNAse III endonuclease, is essential for the biogenesis of several classes of small RNAs, including microRNAs (miRNAs) and endogenous small interfering RNAs (endo-siRNAs), but is also critical for the degradation of toxic transposable elements. In this study, we investigated to which extent DICER1 is required for germ cell development and the progress of spermatogenesis in mice.Principal Findings: We show that the selective ablation of Dicer1 at the early onset of male germ cell development leads to infertility, due to multiple cumulative defects at the meiotic and post-meiotic stages culminating with the absence of functional spermatozoa. Alterations were observed in the first spermatogenic wave and include delayed progression of spermatocytes to prophase I and increased apoptosis, resulting in a reduced number of round spermatids. The transition from round to mature spermatozoa was also severely affected, since the few spermatozoa formed in mutant animals were immobile and misshapen, exhibiting morphological defects of the head and flagellum. We also found evidence that the expression of transposable elements of the SINE family is up-regulated in Dicer1-depleted spermatocytes.Conclusions/Significance: Our findings indicate that DICER1 is dispensable for spermatogonial stem cell renewal and mitotic proliferation, but is required for germ cell differentiation through the meiotic and haploid phases of spermatogenesis

SPO11-Independent DNA Repair Foci and Their Role in Meiotic Silencing

In mammalian meiotic prophase, the initial steps in repair of SPO11-induced DNA double-strand breaks (DSBs) are required to obtain stable homologous chromosome pairing and synapsis. The X and Y chromosomes pair and synapse only in the short pseudo-autosomal regions. The rest of the chromatin of the sex chromosomes remain unsynapsed, contains persistent meiotic DSBs, and the whole so-called XY body undergoes meiotic sex chromosome inactivation (MSCI). A more general mechanism, named meiotic silencing of unsynapsed chromatin (MSUC), is activated when autosomes fail to synapse. In the absence of SPO11, many chromosomal regions remain unsynapsed, but MSUC takes place only on part of the unsynapsed chromatin. We asked if spontaneous DSBs occur in meiocytes that lack a functional SPO11 protein, and if these might be involved in targeting the MSUC response to part of the unsynapsed chromatin. We generated mice carrying a point mutation that disrupts the predicted catalytic site of SPO11 (Spo11YF/YF), and blocks its DSB-inducing activity. Interestingly, we observed foci of proteins involved in the processing of DNA damage, such as RAD51, DMC1, and RPA, both in Spo11YF/YFand Spo11 knockout meiocytes. These foci preferentially localized to the areas that undergo MSUC and form the so-called pseudo XY body. In SPO11-deficient oocytes, the number

A High Incidence of Meiotic Silencing of Unsynapsed Chromatin Is Not Associated with Substantial Pachytene Loss in Heterozygous Male Mice Carrying Multiple Simple Robertsonian Translocations

Meiosis is a complex type of cell division that involves homologous chromosome pairing, synapsis, recombination, and segregation. When any of these processes is altered, cellular checkpoints arrest meiosis progression and induce cell elimination. Meiotic impairment is particularly frequent in organisms bearing chromosomal translocations. When chromosomal translocations appear in heterozygosis, the chromosomes involved may not correctly complete synapsis, recombination, and/or segregation, thus promoting the activation of checkpoints that lead to the death of the meiocytes. In mammals and other organisms, the unsynapsed chromosomal regions are subject to a process called meiotic silencing of unsynapsed chromatin (MSUC). Different degrees of asynapsis could contribute to disturb the normal loading of MSUC proteins, interfering with autosome and sex chromosome gene expression and triggering a massive pachytene cell death. We report that in mice that are heterozygous for eight multiple simple Robertsonian translocations, most pachytene spermatocytes bear trivalents with unsynapsed regions that incorporate, in a stage-dependent manner, proteins involved in MSUC (e.g., γH2AX, ATR, ubiquitinated-H2A, SUMO-1, and XMR). These spermatocytes have a correct MSUC response and are not eliminated during pachytene and most of them proceed into diplotene. However, we found a high incidence of apoptotic spermatocytes at the metaphase stage. These results suggest that in Robertsonian heterozygous mice synapsis defects on most pachytene cells do not trigger a prophase-I checkpoint. Instead, meiotic impairment seems to mainly rely on the action of a checkpoint acting at the metaphase stage. We propose that a low stringency of the pachytene checkpoint could help to increase the chances that spermatocytes with synaptic defects will complete meiotic divisions and differentiate into viable gametes. This scenario, despite a reduction of fertility, allows the spreading of Robertsonian translocations, explaining the multitude of natural Robertsonian populations described in the mouse

The Specification and Global Reprogramming of Histone Epigenetic Marks during Gamete Formation and Early Embryo Development in C. elegans

In addition to the DNA contributed by sperm and oocytes, embryos receive parent-specific epigenetic information that can include histone variants, histone post-translational modifications (PTMs), and DNA methylation. However, a global view of how such marks are erased or retained during gamete formation and reprogrammed after fertilization is lacking. To focus on features conveyed by histones, we conducted a large-scale proteomic identification of histone variants and PTMs in sperm and mixed-stage embryo chromatin from C. elegans, a species that lacks conserved DNA methylation pathways. The fate of these histone marks was then tracked using immunostaining. Proteomic analysis found that sperm harbor ?2.4 fold lower levels of histone PTMs than embryos and revealed differences in classes of PTMs between sperm and embryos. Sperm chromatin repackaging involves the incorporation of the sperm-specific histone H2A variant HTAS-1, a widespread erasure of histone acetylation, and the retention of histone methylation at sites that mark the transcriptional history of chromatin domains during spermatogenesis. After fertilization, we show HTAS-1 and 6 histone PTM marks distinguish sperm and oocyte chromatin in the new embryo and characterize distinct paternal and maternal histone remodeling events during the oocyte-to-embryo transition. These include the exchange of histone H2A that is marked by ubiquitination, retention of HTAS-1, removal of the H2A variant HTZ-1, and differential reprogramming of histone PTMs. This work identifies novel and conserved features of paternal chromatin that are specified during spermatogenesis and processed in the embryo. Furthermore, our results show that different species, even those with diverged DNA packaging and imprinting strategies, use conserved histone modification and removal mechanisms to reprogram epigenetic information