Project description:Numerous studies have shown the potential of spermatozoal RNAs to delineate failures of spermatogenic pathways in infertile samples. However, the RNA contribution of normal fertile samples still needs to be established in relation to transcripts consistently present in human spermatozoa. We report here the spermatozoal transcript profiles characteristic of 24 normally fertile individuals. RNA was extracted from the purified sperm cells of ejaculate and hybridized to Illumina Human-8 BeadChip Microarrays
Project description:In this study, we tested whether reduction in spermatozoal quality induced by under-nutrition is associated with an increase in germ cell apoptosis and a reduction in spermatogenesis, and whether these effects are regulated by small RNAs. Groups of 8 male sheep were fed for a 10% increase or 10% decrease in body mass over 65 days. Testicular tissue from underfed males had more apoptotic germ cells (TUNEL assay; P < 0.05) and greater (P < 0.05) levels of expression of apoptosis-related genes than well-fed males. We identified 44 miRNAs and 35 piRNAs that were differentially expressed in well-fed and underfed males (FDR < 0.05) and found that they were predominantly related to development of the reproductive system, apoptosis (miRNAs), or sperm production and quality (piRNAs). Furthermore, experimental validation showed that novel-miR-144, a homologue of miR-98, would target three apoptotic genes (TP53, CASP3, FASL). The proportion of miRNAs as a total of small RNAs was greater in well-fed males than in underfed males (P < 0.05) and was positively correlated with the proportion of piRNAs in well-fed males (r = 0.8, P < 0.05) and underfed males (r = 0.8, P < 0.05). We conclude that the reductions in spermatozoal quality induced by under-nutrition are due, at least partly, to increased germ cell apoptosis and to changes in the expression of miRNAs and piRNAs.
Project description:Normal human spermatogenesis concludes with the formation of large numbers of morphologically well developed spermatozoa. While transcriptionally quiescent these cells carry an RNA payload that reflects the final spermiogenic phase of transcription. We report here the spermatozoal transcript profiles characteristic of normally fertile individuals and infertile males suffering from a consistent and severe teratozoospermia in which under 4% of spermatozoa are morphologically normal. RNA was extracted from the purified sperm cells of ejaculate and hybridized to Affymetrix U133 (v2) Microarrays. Spermatozoal RNAs were prepared from the semen samples of 21 individuals. An asymmetric dual block design was adopted with biological replicates in both blocks. 13 semen samples were assessed from normally fertile males who had fathered at least one child. 8 semen samples were assessed from infertile individuals with a severe and consistent heterogeneous teratozoospermia who showed no other abnormal semen parameters.
Project description:Formation of biomolecular condensates have emerged as a critical mechanism for compartmentation in living cells. Despite interactions between distinct condensates have been reported, the biological relevance of such interactions remains elusive. In germ cells, small RNA silencing factors are enriched in germ granules, which distinct factors are organized into sub-compartments with specific functions linked to genome surveillance or transgenerational gene silencing. Here we showed that perinuclear germ granules are coated by P body, another condensate known for housing untranslated mRNAs and mRNA degradation factors. Disruption of P body factors, including CGH-1/DDX6 and CAR-1/LSM14, lead to dispersal of small RNA factors from perinuclear germ granules and disorganization of sub-compartments within germ granules. We further showed that CAR-1 promote the interactions between CGH-1 with germ granule factors and such interactions are critical for CGH-1’s ability to promote piRNA-mediated gene silencing. Importantly, we observed that cgh-1 mutants are competent in triggering gene silencing but exhibit defects in maintaining gene silencing effects in the subsequent generations. Small RNA sequencing further showed that cgh-1 mutants exhibit defects in amplifying secondary small RNAs, a known carrier of gene silencing memory. Together, our results uncover the function of P body factors in small RNA-mediated transgenerational gene silencing and highlight how the formation and function of one condensate can be regulated by an adjacent, interacting condensate in cells.