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Microarray mRNA expression profiling of E16.5 foetal gonocytes

ABSTRACT: Gonocytes were FACS sorted from testes of E16.5 Miwi2 Tom/+ embryos using the tdTomato reporter knocked into the Miwi2 locus. RNA was isolated from sorted cells and transcript expression analyzed by microarray measurement.

ORGANISM(S): Mus musculus

SUBMITTER: Ansgar Zoch

PROVIDER: E-MTAB-7985 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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SPOCD1 is an essential executor of piRNA-directed de novo DNA methylation.

Zoch Ansgar A Auchynnikava Tania T Berrens Rebecca V RV Kabayama Yuka Y Schöpp Theresa T Heep Madeleine M Vasiliauskaitė Lina L Pérez-Rico Yuvia A YA Cook Atlanta G AG Shkumatava Alena A Rappsilber Juri J Allshire Robin C RC O'Carroll Dónal D

Nature 20200716 7822

In mammals, the acquisition of the germline from the soma provides the germline with an essential challenge: the need to erase and reset genomic methylation1. In the male germline, RNA-directed DNA methylation silences young, active transposable elements2-4. The PIWI protein MIWI2 (PIWIL4) and its associated PIWI-interacting RNAs (piRNAs) instruct DNA methylation of transposable elements3,5. piRNAs are proposed to tether MIWI2 to nascent transposable element tran ...[more]

PMID: 32674113

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Affymetrix gene expression array (mouse 2.0) of sorted undifferentiated spermatogonia in inducible Miwi2 knock-out model

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2018-03-21 | E-MTAB-6612 | biostudies-arrayexpress

Profiling of Mili and Miwi2 associated piRNAs in WT, MiliDAH, Miwi2DAH and Miwi2KO from E16.5 gonadocytes.

Project description:Piwi proteins and piRNAs have conserved functions in transposonM- silencing. The murine Piwi proteins Mili and Miwi2 direct epigeneticM- LINE1 (L1) and intracisternal A particle (IAP) transposon silencingM- during genome reprogramming in the embryonic male germline. PiwiM- proteins are proposed to be piRNA-guided endonucleases that initiateM- secondary piRNA biogenesis . However the actual contribution of theirM- endonuclease activities to piRNA biogenesis and transposon silencingM- remain unknown. To investigate the role of Piwi-catalyzedM- endonucleolytic activity, we engineered point mutations in the mouseM- that substitute the second D to an A in the catalytic triad (DDH) ofM- Mili and Miwi2, generating the MiliDAH and Miwi2DAH alleles,M- M- respectively. Analysis of Mili-bound piRNAs from homozygous MiliDAHM- fetal gonadocytes revealed the failure of transposon piRNA amplification resulting in the stark reduction of piRNA bound withinM- Miwi2 ribonuclear particles (RNPs). We find that Mili-mediated piRNA amplification is selectively required for L1 but not IAP silencing.M- The defective piRNA pathway in MiliDAH mice results in spermatogenic failure and sterility. Surprisingly, homozygous Miwi2DAH mice areM- fertile, transposon silencing is established normally and no defectsM- in secondary piRNA biogenesis are observed. In addition, the hallmarks of piRNA amplification are observed in Miwi2-deficient gonadocytes. WeM- conclude that cycles of intra-Mili secondary piRNA biogenesis fuelM- piRNA amplification that is selectively required for L1 silencing.M-

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SPOCD1 is a novel executor of piRNA-directed DNA methylation that links MIWI2 to the de novo DNA methylation machiner

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