RNA helicase D1PAS1 resolves R-loops and forms a complex for mouse pachytene piRNA biogenesis required for male fertility
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ABSTRACT:
PROVIDER: PRJNA970354 | ENA |
REPOSITORIES: ENA
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| SRR24467594.fastq.gz | Fastqsanger.gz |
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Project description:To investigate the function of RNA helicase D1PAS1 in mouse spermatogenesis, we established D1Pas1 knockout (KO) and D1Pas1-FLAG knockin (KI) lines in which the D1Pas1 gene has been knocked out by deleting the transcription start site and the 3XFLAG sequences added at the end of C-term by CRISPR/Cas9. We then performed gene expression profiling analysis using data obtained from RNA-seq of 3 different biological replicates at three different developmental stages. Investigated small RNA abundance from WT and KO testis via small RNA-seq. To profile the genome-wide D1PAS1 RNA helicase dependent R-loop loci, we performed BisMapR sequencing analysis with D1PAS1 KI and KO spermatocytes.
2024-11-06 | GSE231983 | GEO
Project description:In the fetal mouse testis, PIWI Interacting RNAs (piRNAs) guide PIWI proteins to silence transposons, but after birth, most post-pubertal pachytene piRNAs map to genome uniquely and are thought to regulate genes required for male fertility. In human males, the developmental classes, precise genomic origins, and transcriptional regulation of post-natal piRNAs remain undefined. Here, we demarcate the genes and transcripts that produce post-natal piRNAs in human juvenile and adult testes. As in mouse, A‑MYB in humans drives transcription of both pachytene piRNA precursor transcripts and the mRNAs encoding piRNA biogenesis factors. Although human piRNA genes are syntenic to those in other placental mammals, their sequences are poorly conserved. In fact, pachytene piRNA loci are rapidly diverging even among modern humans. Our findings suggest that during mammalian evolution, pachytene piRNA genes are under fewer selective constraints. We speculate that pachytene piRNA diversity may provide a hitherto unrecognized driver of reproductive isolation.
2019-12-23 | GSE135791 | GEO
Project description:Purpose: piRNAs are a class of germline-specific, small non-coding RNAs required to maintain genome integrity and preserve RNA homeostasis essential for male gametogenesis. During mouse spermatogenesis, male germ cells transcribe piRNAs at pre-pachytene anad pachytene stages of development from genetically distince loci. The most well-studied and conserved function of pre-pachytene piRNAs is repression of transposons to ensure the integrity of the germline genome. In contrast to pre-pachytene piRNAs, the mode of action and function of pachytene piRNA is much less understood. To investigate and characterize the functional importance of pachytene piRNAs in spermatogenesis and male fertility, we have used CRISPR/Cas9 and established mutant mice which have deleted a bi-directional promoter of the pachytene piRNA cluster on Chr18. Mutant mice have a male sterile phenotype with acrosome overgrowth and defective motility.
2019-08-30 | GSE136603 | GEO
Project description:Pachytene piRNAs control male fertility across metazoans, yet mechanisms through which they govern meiosis I in pachytene remain unclear. We demonstrate, in C. elegans, that homolog pairing, and crossover formation are compromised in the absence of piRNA function. We identify several protein coding genes as targets of piRNAs including Polo like kinase 3 (PLK-). Pachytene piRNAs spatially restrict and exclude PLK-3 expression from meiosis I. Expansion of PLK-3 expression into the pachytene region, upon loss of piRNAs, is reflected in meiotic defects, which are partially rescued by removal of ectopic PLK-3. Pachytene piRNAs regulate PLK-3 through both translational repression and post-transcriptional degradation establishing the spatiotemporal control of gene expression. Given the conserved role of pachytene piRNAs, we propose that these mechanisms may be applicable to mammals.
2023-10-14 | GSE245023 | GEO
Project description:Piwi-interacting small RNAs (piRNAs) of fetal prospermatogonia of mice have been strongly implicated in transposon control. In contrast, little is known about biogenesis and function of abundant piRNAs from adult testes expressed in late spermatocytes and round spermatids. These so-called "pachytene" piRNAs are processed from long non-coding piRNA precursors and have no defined RNA targets in the transcriptome even though their binding partner Piwi, MIWI, is essential for spermiogenesis and fertility. Here we report that 129SvJae mice lacking Maelstrom (MAEL), a conserved piRNA pathway protein, exhibit spermiogenic arrest with defects in acrosome and flagellum formation. Further analysis revealed MAEL association with RNPs containing MIWI, TDRD6, and processed intermediates of pachytene piRNA precursors of various length. Loss of MAEL causes a 10-fold drop in pachytene piRNA levels but an increase in piRNAs from abundantly expressed mRNAs. These results suggest a MAEL-dependent mechanism for the selective processing of pachytene piRNA precursor into piRNAs. Strikingly, ribosome profiling of Mael-null testes revealed that reduced piRNA production is accompanied by reduced translation of over 800 spermiogenic mRNAs including those encoding acrosome and flagellum proteins. In light of recent reports of piRNA-independent protection of translationally repressed mRNPs by MIWI and piRNA-dependent turnover of MIWI, we propose that pachytene piRNAs function by controlling the availably of MIWI for the translational repression of spermiogenic mRNAs.
2014-07-25 | GSE50983 | GEO
Project description:Ribosomes guide pachytene piRNA formation on long intergenic piRNA precursors
| PRJNA275185 | ENA
Project description:In the male germ cells of eutherian animals, 26–30-nt-long PIWI-interacting RNAs (piRNAs) emerge when spermatocytes enter the pachytene phase of meiosis. These pachytene piRNAs derive from ~100 discrete autosomal loci resemble canonical protein-coding genes and long non-coding RNA-producing genes—they are transcribed by RNA polymerase II, bearing 5´ caps and 3´ poly(A) tails, and their transcripts often contain introns that are removed before nuclear export and processing into piRNAs. However, it is unclear which genic and epigenetic features distinguish pachytene piRNA genes from other types of genes and dictate their germline-specific expression. We report that an unusually long first exon (≥ 10 kb) or a long gene absent of introns altogether is highly correlated with both the germline-specific production of piRNA precursor transcripts from mouse pachytene piRNA loci. We also found that these precursor transcripts are enriched in the binding by THOC1 (also known as HPR1) and THOC2, subunits of the THO complex critical for transcription elongation and nuclear export of mRNAs. Our integrative analysis of transcriptome, piRNA, and epigenome datasets across multiple species reveals that a long first exon is an evolutionarily conserved feature of pachytene piRNA loci. We further found that a highly methylated promoter, often containing a low or intermediate level of CG dinucleotides, correlates with germline expression and somatic silencing of pachytene piRNA loci.
2020-10-24 | GSE147724 | GEO
Project description:Ricochet transcription of pachytene piRNA promoters in mammals
| PRJEB3240 | ENA
Project description:Piwi-interacting small RNAs (piRNAs) of fetal prospermatogonia of mice have been strongly implicated in transposon control. In contrast, little is known about biogenesis and function of abundant piRNAs from adult testes expressed in late spermatocytes and round spermatids. These so-called "pachytene" piRNAs are processed from long non-coding piRNA precursors and have no defined RNA targets in the transcriptome even though their binding partner Piwi, MIWI, is essential for spermiogenesis and fertility. Here we report that 129SvJae mice lacking Maelstrom (MAEL), a conserved piRNA pathway protein, exhibit spermiogenic arrest with defects in acrosome and flagellum formation. Further analysis revealed MAEL association with RNPs containing MIWI, TDRD6, and processed intermediates of pachytene piRNA precursors of various length. Loss of MAEL causes a 10-fold drop in pachytene piRNA levels but an increase in piRNAs from abundantly expressed mRNAs. These results suggest a MAEL-dependent mechanism for the selective processing of pachytene piRNA precursor into piRNAs. Strikingly, ribosome profiling of Mael-null testes revealed that reduced piRNA production is accompanied by reduced translation of over 800 spermiogenic mRNAs including those encoding acrosome and flagellum proteins. In light of recent reports of piRNA-independent protection of translationally repressed mRNPs by MIWI and piRNA-dependent turnover of MIWI, we propose that pachytene piRNAs function by controlling the availably of MIWI for the translational repression of spermiogenic mRNAs. piRNA sequencing, RNA immunoprecipitation, and expression measurements (RNA-Seq and ribosome profiling) in wild-type and Mael -/- testes
2014-07-25 | E-GEOD-50983 | biostudies-arrayexpress
Project description:PIWI-interacting RNAs (piRNAs) are a class of small non-coding RNAs essential for fertility. In adult mouse testes, most piRNAs are derived from long single-stranded RNAs lacking annotated open reading frames (ORFs). The mechanisms underlying how piRNA sequences are defined during the cleavages of piRNA precursors remain elusive. Here, we show that 80S ribosomes translate the 5'-proximal short ORFs (uORFs) of piRNA precursors. The MOV10L1/Armitage RNA helicase then facilitates the translocation of ribosomes into the uORF downstream regions (UDRs). The ribosome-bound UDRs are targeted by piRNA processing machinery, with the processed ribosome-protected regions becoming piRNAs. The dual modes of interaction between ribosomes and piRNA precursors underlie the distinct piRNA biogenesis requirements at uORFs and UDRs. Ribosomes also mediate piRNA processing in roosters and green lizards, implying that this mechanism is evolutionarily conserved in amniotes. Our results uncover a function for ribosomes on non-coding regions of RNAs and reveal the mechanisms underlying how piRNAs are defined.
| S-EPMC8041231 | biostudies-literature