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Transcriptomic analysis of wild-type, spn-E and aub mutant 0-2h- and 2-4h-embryos

ABSTRACT: We used microarrays to identify unstable mRNAs in wild-type embryos by comparing the transcriptomes derived from 0-2h- and 2-4h-embryos. We then analyzed which of these unstable mRNAs are stabilized in embryos from aub or spn-E mutant mothers (aub or spn-E mutant embryos) by comparing mutant and wild-type embryonic transcriptomes

ORGANISM(S): Drosophila melanogaster

PROVIDER: GSE70775 | GEO | 2015/08/19

SECONDARY ACCESSION(S): PRJNA289519

REPOSITORIES: GEO

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RNA helicase Spn-E is required to maintain Aub and AGO3 protein levels for piRNA silencing in the germline of Drosophila

Project description:Germline-specific RNA helicase Spindle-E (Spn-E) is known to be essential for piRNA silencing in Drosophila that takes place mainly in the perinuclear nuage granules. Loss-of-function spn-E mutations lead to tandem Stellate genes derepression in the testes and retrotransposon mobilization in the ovaries. However, Spn-E functions in the piRNA pathway are still obscure. Analysis of total library of short RNAs from the testes of spn-E heterozygous flies revealed the presence of abundant piRNA ping-pong pairs originating from Su(Ste) transcripts. The abundance of these ping-pong pairs were sharply reduced in the library from the testes of spn-E mutants. Thus we found that ping-pong mechanism contributed to Su(Ste) piRNA generation in the testes. The lack of Spn-E caused a significant drop of protein levels of key ping-pong participants, Aubergine (Aub) and AGO3 proteins of PIWI subfamily, in the germline of both males and females, but did not disrupt of their assembly in nuage granules. We found that observed decline of the protein expression was not caused by suppression of aub and ago3 transcription as well as total transcription, indicating possible contribution of Spn-E to post-transcriptional regulation.

2016-06-08 | GSE50984 | GEO

Modulation of Aub–Tdrd interactions elucidates piRNA amplification and germ plasm formation

Project description:Aub protein guided by piRNAs ensures genome integrity by cleaving retrotransposons and genome propagation by trapping mRNAs to form the germ plasm that instructs germ cell formation. The amino terminus of Aub (Aub-NT) is rich in arginines (Aub-NTRs), which are symmetrically dimethylated (sDMAs), and interacts with Tudor protein and other Tudor domain containing proteins (Tdrds). Aub-Tdrd interactions play critical roles in suppressing active retrotransposons via piRNA amplification and in germ plasm formation via generation of Aub-Tudor ribonucleoproteins. Here we show that Aub-NTRs are dispensable for primary piRNA biogenesis but essential for piRNA amplification and transposon control and that sDMAs in Aub-NT are essential for germ plasm formation and germ cell specification, but largely dispensable for piRNA amplification and transposon control.

2020-12-14 | GSE155874 | GEO

Aubergine protein interactors in 0-2 hour-Drosophila embryos

Project description:To identify Aub function in mRNA regulation in the Drosophila embryo, we have performed mass spectrometry analysis of Aub interactors, following immunoprecipitation of GFP-Aub in 0-2 hour-embryos. Immunoprecipitation of GFP alone was used as negative control. Because Aub accumulates at high levels in the germ plasm, GFP-Aub immunoprecipitation was also performed in oskar mutant embryos that do not assemble the germ plasm. Proteins coprecipitating with GFP-Aub were similar in wild-type and oskar mutant embryos. Translation factors were enriched among proteins coprecipitating with Aub.

2020-04-27 | PXD016399 | Pride

Aubergine iCLIP reveals direct regulation of mRNAs involved in germ cell development by piRNAs in the early embryo

Project description:The Piwi-interacting RNA (piRNA) pathway plays an essential role in the repression of transposons in the germline. Other functions of piRNAs such as post-transcriptional regulation of mRNAs are now emerging. Here, we perform iCLIP with the PIWI protein Aubergine (Aub) and identify hundreds of maternal mRNAs interacting with Aub in the early Drosophila embryo. Gene expression profiling reveals that a proportion of these mRNAs undergoes Aub-dependent destabilization during the maternal-to-zygotic transition. Strikingly, Aub-dependent unstable mRNAs encode germ cell determinants. iCLIP with an Aub mutant that is unable to bind piRNAs confirms that binding of Aub to mRNAs is piRNA dependent. Base-pairing between piRNAs and mRNAs can induce mRNA cleavage and decay that are essential for embryonic development. These results reveal a general regulation of maternal mRNAs by Aub and piRNAs, which plays a key developmental role in the embryo, by mediating decay and localization of mRNAs encoding germ cell determinants.

2015-08-19 | GSE70777 | GEO

Drosophila melanogaster

Project description:Transcriptomic analysis of wild-type, spn-E and aub mutant 0-2h- and 2-4h-embryos

| PRJNA289519 | ENA

Binding of guide piRNA triggers methylation of the unstructured N-terminal region of Aub leading to assembly of the piRNA amplification complex

Project description:In Drosophila, Piwi proteins use guide piRNAs to repress selfish genomic elements, protecting the genomic integrity of gametes and ensuring the fertility of animal species. Efficient transposon repression depends on amplification of piRNA guides in the ping-pong cycle, which in Drosophila entails tight cooperation between two Piwi proteins, Aub and Ago3. Here we show that post-translational modification, symmetric dimethylarginine (sDMA), of Aub is essential for piRNA biogenesis, transposon silencing and fertility. Methylation is triggered by loading of a piRNA guide into Aub, which exposes its unstructured N-terminal region to the PRMT5 methylosome complex. Thus, sDMA modification is a signal that Aub is loaded with piRNA guide. Amplification of piRNA in the ping-pong cycle requires assembly of a tertiary complex scaffolded by Krimper, which simultaneously binds the N-terminal regions of Aub and Ago3. To promote generation of new piRNA, Krimp uses its two Tudor domains to bind Aub and Ago3 in opposite modification and piRNA-loading states. Our results reveal essential functions of post-translational modifications in unstructured regions of Piwi proteins and of Tudor domains capable of discriminating between modification states in piRNA biogenesis and silencing.

2020-06-25 | GSE153156 | GEO

miR-309-dependent unstable maternal mRNAs are stabilized in smaug mutants

Project description:The behavior of 410 miR-309-dependent maternal mRNAs (from Bushati et al., 2008) in embryos from wild type and smg-mutant females relative to wild-type stage 14 oocyte reference RNA. All are unstable in wild-type while almost 85% are stabilized in smg mutants

2009-04-23 | GSE13288 | GEO

Characterization of expression changes in armi,rhino,aub,ago3 mutants by tiling array

Project description:We characterized changes of transposon and mRNA expressions in armi, rhino ,aub, ago3 mutants with respect to wild type using Affy tiling array. In most of these mutants, mRNA expressions were mostly unchanged but increased expressions was observed for many transposons indicating the role of these proteins in silencing transposons in Drosophila ovaries Keywords: Tiling array transcriptome profiling

2009-05-03 | GSE14370 | GEO

Krimper enforces an antisense bias on piRNA pools by binding AGO3 in the Drosophila germline

Project description:Piwi-interacting RNAs (piRNAs) suppress transposon activity in animal germ cells. In the Drosophila ovary, primary Aubergine (Aub)-bound antisense piRNAs initiate the ping-pong cycle to produce secondary AGO3-bound sense piRNAs. This increases the number of secondary Aub-bound antisense piRNAs that can act to destroy transposon mRNAs. Here we show that Krimper (Krimp), a Tudor-domain protein, directly interacts with piRNA-free AGO3 to promote symmetrical dimethylarginine (sDMA) modification, ensuring sense piRNA-loading onto sDMA-modified AGO3. In aub mutant ovaries, AGO3 associates with ping-pong signature piRNAs, suggesting AGO3’s compatibility with primary piRNA loading. Krimp sequesters ectopically expressed AGO3 within Krimp bodies in cultured ovarian somatic cells (OSCs), in which only the primary piRNA pathway operates. Upon krimp-RNAi in OSCs, AGO3 loads with piRNAs, further showing the capacity of AGO3 for primary piRNA loading. We propose that Krimp enforces an antisense bias on piRNA pools by binding AGO3 and blocking its access to primary piRNAs. In order to investigate function of Krimp in piRNA pathway, sequencing of Piwi subfamily protein associated small RNAs was performed using adult Drosophila ovaries and Ovarian Somatic Cells (OSCs) depleted for Krimp or Aub.

2015-05-28 | E-GEOD-69293 | biostudies-arrayexpress

Transcriptional response of monocyte-derived macrophages to stimulation with wildtype or pneumolysin-deficient Streptrococcus pneumoniae in the presence or absence of cytochalasin D

Project description:We used genome-wide transcriptional profiling by microarray to assess the contribution of pneumolysin on macrophage innate immune responses to the TIGR4 strain of Streptococcus pneumoniae (Spn). We focused on the early transcriptional responses at 4 hours after inoculation of human blood monocyte-derived macrophage cultures with Spn at a multiplicity of 10 bacteria to each cell. We compared transcriptomes in the presence and absence of wildtype or pneumolysin-deficient TIGR4 Spn, and also in the presence and absence of cytochalasin D to assess whether there is a differential effect of pneumolysin on innate immune responses with and without bacterial internalisation.

2021-03-31 | E-MTAB-8947 | biostudies-arrayexpress

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