Oxidation treated small RNA-seq for studying roles of chicken piRNAs in germ cell development
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ABSTRACT:
ORGANISM(S): Gallus gallus
PROVIDER: GSE98005 | GEO | 2018/05/21
REPOSITORIES: GEO
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Project description:Background: Genes, RNAs, and proteins play important roles during germline development. However, the functions of non-coding RNAs (ncRNAs) on germline development remain unclear in avian species. Recent high-throughput techniques have identified several classes of ncRNAs, including micro RNAs (miRNAs), small-interfering RNAs (siRNAs), and PIWI-interacting RNAs (piRNAs). These ncRNAs are functionally important in the genome, however, the identification and annotation of ncRNAs in a genome is challenging. The aim of this study was to identify different types of small ncRNAs particularly piRNAs, and the role of piRNA pathway genes in the protection of chicken primordial germ cells (PGCs). Results: At first, we performed next-generation sequencing to identify ncRNAs in chicken PGCs, and we performed ab initio predictive analysis to identify putative piRNAs in PGCs. Then, we examined the expression of three repetitive sequence-linked piRNAs and 14 genic-transcript-linked piRNAs along with their linked genes using real-time PCR. All piRNAs and their linked genes were highly expressed in PGCs. Subsequently, we knocked down two known piRNA pathway genes of chicken, PIWI-like protein 1 (CIWI) and 2 (CILI), in PGCs using siRNAs. After knockdown of CIWI and CILI, we examined their effects on the expression of six putative piRNA-linked genes and DNA double-strand breakage in PGCs. The knockdown of CIWI and CILI upregulated chicken repetitive 1 (CR1) element and RAP2B, a member of RAS oncogene family, and increased DNA double-strand breakage in PGCs. Conclusions: Our results increase the understanding of PGC-expressed ncRNAs and the role of piRNA pathway genes in the protection of germ cells. Small ncRNA expression profiling in SSEA-1 positive PGCs, SSEA-1 negative gonadal stromal cells (GSCs), Stage X blastoderms, and chicken embryonic fibroblast (CEFs)
2015-04-27 | E-GEOD-60400 | biostudies-arrayexpress
Project description:The small non-coding RNAs (sncRNAs) are considered as postranscriptional key regulators of male germ cell development. In addition to microRNAs (miRNAs) and PIWI-interacting RNAs (piRNAs), other sncRNAs generated from small nucleolar RNAs (snoRNAs), tRNAs or rRNAs processing may also play important regulatory roles in spermatogenesis. By next generation sequencing (NGS), we characterized the different sncRNA populations detected at three milestone stages in male germ differentiation: primordial germ cells (PGCs) at 13.5 dpc, pubertal spermatogonia cells, and mature spermatozoa. In order to assess the potential transmission of the sncRNAs through the mature spermatozoa during fertilization, the sncRNA population detected in male germ cells was also compared with sncRNAs detected in unfertilized mouse oocytes and zygotes. Combining the data obtained by NGS and microarrays from whole PGC and spermatogonia transcriptome, we defined the potential regulatory roles of specific miRNAs and their validated targets. Similar to miRNAs, both the small RNAs derived from snoRNAs and the piRNAs, could be involved in the postranscriptional regulation of mRNA transcripts during the male germ development. Finally, our results strongly suggest that the small RNAs-derived from tRNAs and rRNAs are interacting with PIWI proteins, and specifically with MILI. These new classes of piRNAs are not generated by the ping-pong pathway and could be the source of primary piRNAs. mRNA analysis of Primordial Germ Cells (PGCs), Spermatogonia cells (SPG), adult testis (AdT) and Gonad-less (GL) embryos. Indirect comparisons were made across multiple arrays with raw data pulled from different channels for data analysis and comparison to the control data.
2015-03-26 | E-GEOD-59252 | biostudies-arrayexpress
Project description:Background: Genes, RNAs, and proteins play important roles during germline development. However, the functions of non-coding RNAs (ncRNAs) on germline development remain unclear in avian species. Recent high-throughput techniques have identified several classes of ncRNAs, including micro RNAs (miRNAs), small-interfering RNAs (siRNAs), and PIWI-interacting RNAs (piRNAs). These ncRNAs are functionally important in the genome, however, the identification and annotation of ncRNAs in a genome is challenging. The aim of this study was to identify different types of small ncRNAs particularly piRNAs, and the role of piRNA pathway genes in the protection of chicken primordial germ cells (PGCs). Results: At first, we performed next-generation sequencing to identify ncRNAs in chicken PGCs, and we performed ab initio predictive analysis to identify putative piRNAs in PGCs. Then, we examined the expression of three repetitive sequence-linked piRNAs and 14 genic-transcript-linked piRNAs along with their linked genes using real-time PCR. All piRNAs and their linked genes were highly expressed in PGCs. Subsequently, we knocked down two known piRNA pathway genes of chicken, PIWI-like protein 1 (CIWI) and 2 (CILI), in PGCs using siRNAs. After knockdown of CIWI and CILI, we examined their effects on the expression of six putative piRNA-linked genes and DNA double-strand breakage in PGCs. The knockdown of CIWI and CILI upregulated chicken repetitive 1 (CR1) element and RAP2B, a member of RAS oncogene family, and increased DNA double-strand breakage in PGCs. Conclusions: Our results increase the understanding of PGC-expressed ncRNAs and the role of piRNA pathway genes in the protection of germ cells.
2015-04-27 | GSE60400 | GEO
Project description:The small non-coding RNAs (sncRNAs) are considered as postranscriptional key regulators of male germ cell development. In addition to microRNAs (miRNAs) and PIWI-interacting RNAs (piRNAs), other sncRNAs generated from small nucleolar RNAs (snoRNAs), tRNAs or rRNAs processing may also play important regulatory roles in spermatogenesis. By next generation sequencing (NGS), we characterized the different sncRNA populations detected at three milestone stages in male germ differentiation: primordial germ cells (PGCs) at 13.5 dpc, pubertal spermatogonia cells, and mature spermatozoa. In order to assess the potential transmission of the sncRNAs through the mature spermatozoa during fertilization, the sncRNA population detected in male germ cells was also compared with sncRNAs detected in unfertilized mouse oocytes and zygotes. Combining the data obtained by NGS and microarrays from whole PGC and spermatogonia transcriptome, we defined the potential regulatory roles of specific miRNAs and their validated targets. Similar to miRNAs, both the small RNAs derived from snoRNAs and the piRNAs, could be involved in the postranscriptional regulation of mRNA transcripts during the male germ development. Finally, our results strongly suggest that the small RNAs-derived from tRNAs and rRNAs are interacting with PIWI proteins, and specifically with MILI. These new classes of piRNAs are not generated by the ping-pong pathway and could be the source of primary piRNAs. Comparative analysis from deep sequencing of piRNAs and endo-siRNAs in mouse oocytes, spermatozoa and zygotes
2015-03-26 | E-GEOD-59254 | biostudies-arrayexpress
Project description:Although mitochondria are widely studied organelles, the recent interest in the role of mitochondrial small non-coding RNAs (sncRNA) is providing new functional perspectives in germ cell development and differentiation. PIWI-interacting RNAs (piRNAs) are single-stranded sncRNAs of 20-35 nt generated from the processing of pre-piRNAs longer molecules to be functionally bound to PIWI proteins. Initially ascribed to germ cells, as protectors against transposons, we now know that they are also expressed in somatic cells. In mammals, germ cells differentiate at early stages of embryonic development in the primitive gonads as primordial germ cells (PGCs), initiating divergent pathways in both sexes, accompanied by somatic cells (SCs) of the ovary or testis. Previously, we identified mitochondrial piRNAs (mito-piRNAs) in mouse germ and somatic cells. However, a comparative analysis of mito-piRNAs between PGCs and SCs, between sexes and during early gonadal development has not been performed. We leverage NGS data obtained from PGCs and SCs purified from early differentiating embryonic ovaries and testis from E11.5 to E13.5. Using bioinformatic tools, here we unravel: the origin (from nuclear or mitochondrial genome), the levels of expression, the potential role of mito-piRNAs, as well as their association with genomic regions encoding other sncRNAs (such as tRNAs and rRNAs) and the mitochondrial regulatory region (D-loop). Finally, our results indicate nucleo-mitochondrial communication at both anterograde and retrograde signaling, mediated by mito-piRNAs.
2021-07-01 | GSE179299 | GEO
Project description:In metazoan gonads, transposable elements (TEs) mobilization is limited by PIWI-interacting RNAs (piRNAs). These small RNAs originate from specific source loci, the piRNA clusters. piRNAs are known to silence TEs in the cells where they are produced. Endogenous retroviruses (ERVs), a subclass of TEs, pose a particular threat because they are capable of transiting from cell to cell. In this study, we reveal that piRNAs produced locally in germ cells counteract invasion by ERVs arriving from adjacent somatic cells. We reactivated the Drosophila ERV ZAM in somatic gonadal cells by deleting, using CRISPR-Cas9 genome editing, its single copy in the somatic flamenco piRNA cluster, while keeping the piRNA pathway fully functional. Upon reactivation, ZAM invaded the oocytes, resulting in transposition and severe fertility defects. We show that once ZAM-piRNAs are produced in germ cells they counter the invasion. Our study sheds new light on the mechanisms of recognition and regulation of invasive genetic elements, which is essential for the maintenance of genome integrity.
2022-09-30 | GSE213456 | GEO
Project description:In metazoan gonads, transposable elements (TEs) mobilization is limited by PIWI-interacting RNAs (piRNAs). These small RNAs originate from specific source loci, the piRNA clusters. piRNAs are known to silence TEs in the cells where they are produced. Endogenous retroviruses (ERVs), a subclass of TEs, pose a particular threat because they are capable of transiting from cell to cell. In this study, we reveal that piRNAs produced locally in germ cells counteract invasion by ERVs arriving from adjacent somatic cells. We reactivated the Drosophila ERV ZAM in somatic gonadal cells by deleting, using CRISPR-Cas9 genome editing, its single copy in the somatic flamenco piRNA cluster, while keeping the piRNA pathway fully functional. Upon reactivation, ZAM invaded the oocytes, resulting in transposition and severe fertility defects. We show that once ZAM-piRNAs are produced in germ cells they counter the invasion. Our study sheds new light on the mechanisms of recognition and regulation of invasive genetic elements, which is essential for the maintenance of genome integrity.
2022-09-30 | GSE213368 | GEO
Project description:PIWI-interacting RNAs (piRNAs) protect the germ line by targeting transposable elements (TEs) through base-pair complementarity. We do not know how piRNAs co-evolve with TEs in chickens. Here we reported that all active TEs in the chicken germ line are targeted by piRNAs, and as TEs lose their activity, the corresponding piRNAs erode away. We observed de novo piRNA birth as host responds to a recent retroviral invasion. Avian leukosis virus (ALV) has endogenized prior to chicken domestication, remains infectious, and threatens poultry industry. Domestic fowl produced piRNAs targeting ALV from a genomic locus that was known to render its host ALV resistant. This genomic locus does not produce piRNAs in undomesticated wild chickens. Our findings uncover rapid piRNA evolution reflecting contemporary TE activity, identify a new piRNA acquisition modality by activating a pre-existing genomic locus, and extend piRNA defense roles to include the period when endogenous retroviruses are still infectious.
2017-04-07 | GSE93559 | GEO
Project description:Small RNAs exert an effect through diverse RNA interference pathways to transcriptionally or post-transcriptionally silence their targets. The Piwi-interacting RNAs (piRNAs) represent a germline-specific small RNA pathway where Piwi proteins themselves are thought to mediate piRNA biosynthesis. Here, we provide strong evidence for a piRNA amplification loop in zebrafish, in which Ziwi and Zili bind piRNAs of opposite polarity. Furthermore, we describe a function for Zili in transposon defense and germ cell differentiation, as well as a crucial function in meiosis, significantly extending the function of Piwi proteins beyond the control of transposable elements in vertebrates. small RNA from total gonads or Argonaute IPs were cloned and sequenced using 454 GS FLX system.
2008-11-04 | E-GEOD-12547 | biostudies-arrayexpress
Project description:The germ cell lineage ensures the creation of new individuals perpetuating the genetic information across generations. Primordial germ cells (PGCs) are pioneers of gametes and exist transiently during development until they differentiate into oogonia in females, or spermatogonia in males. Little is known about the molecular characteristics of PGCs in cattle. By performing single-cell RNA-sequencing of bovine fetal gonads we evaluated the molecular signatures of bovine germ cells and their niche at 50 days of development. Our results indicate that PGCs and somatic cells were in very early stages of development, and that bovine and human germ cells share expression of transcriptional regulators of germline specification, and several genes coding for membrane-bound proteins. Additionally, the expression of members of Notch, Nodal/Activin, and BMP signaling cascades in the bovine fetal ovary, suggests that these pathways are involved in the interaction between germ cells and their niche. Results of this study provide insights into the mechanisms involved in the development of bovine PGCs and put in evidence similarities between the bovine and human germline.
2021-01-29 | GSE162952 | GEO