Unknown,Transcriptomics,Genomics,Proteomics

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Expression data from shMiwi/shCaf1 transduced mouse elongating spermatids

ABSTRACT: We reasoned that the pi-RISC, by virtue of the enormous sequence portfolio of piRNAs, might mediate elimination of a large variety of mRNAs in late stages of spermiogenesis. Both Miwi-null and Caf1-null mice showed early spermiogenic arrest, preventing us from determining the role of MIWI and CAF1 in elongating spermatids using the existing genetic models. We therefore used microarrays to detail the global effect of MIWI or CAF1 on mRNA levels in mouse elongating spermatids. Using GFP+ elongating spermatids sorted from mouse testes transduced with shMiwi:GFP, shCaf1:GFP, or control pSilencer:GFP, we performed transcriptome profiling on Affymetrix mouse arrays.

ORGANISM(S): Mus musculus

SUBMITTER: Lantao Gou

PROVIDER: E-GEOD-59291 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Pachytene piRNAs are PIWI-interacting small RNAs abundantly expressed in pachytene spermatocytes and spermatids in adult mouse testes. Both MIWI and MILI-bound pachytene piRNAs have been found enriched in round spermatids. Miwi-null male mice are sterile due to spermiogenic arrest. In C. elegans, sperm-borne piRNAs appear to have an epigenetic role during fertilization and development because progeny of offspring derived from piRNA-deficient sperm display a progressive fertility loss after several generations. In mice, it remains unknown whether MIWI-bound pachytene piRNA-deficient round spermatids can produce offspring, and whether the progeny of offspring derived from MIWI-bound pachytene piRNA-deficient round spermatids also exhibit transgenerational loss of fertility. Here, we report that Miwi KO round spermatids could fertilize both wild type (WT) and Miwi KO oocytes through round spermatid microinjection (ROSI), and produce healthy and fertile offspring despite the aberrant pachytene piRNA profiles in those Miwi KO spermatids. Progeny of ROSI-derived heterozygotes, both male and female, displayed normal fertility for at least three generations when bred with either WT or Miwi KO females. Our data indicate that aberrant MIWI-bound pachytene piRNAs profiles in spermatids do not affect fertilization, early embryonic development, or fertility of the offspring, suggesting a normal pachytene piRNAs profile is not required for paternal transgenerational epigenetic inheritance in mice. Method: Round spermatids were purified from WT and Miwi KO adult testes using a mini-STA-PUT method[Methods Enzymol 1993; 225:84-113.]，the purity of round spermatids was >90% based on our previous report[ J Biol Chem 2012; 287:25173-25190.]. Small RNA was isolated from round spermatids using the mirVana RNA isolation kit (Ambion) according to the manufacturer’s instructions. RNA quality and quantity were assessed using the Agilent 2100 Bioanalyzer. Small RNA-Seq was performed on an Ion Proton sequencer (Life Technologies). Libraries were prepared using the Ion Total RNA-Seq Kit v2 (Invitrogen) with biological triplicates for WT and Miwi KO samples. Resutls:Our data indicate that aberrant MIWI-bound pachytene piRNAs profiles in spermatids do not affect fertilization, early embryonic development, or fertility of the offspring, suggesting a normal pachytene piRNAs profile is not required for paternal transgenerational epigenetic inheritance in mice.

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Expression data from shMiwi/shCaf1 transduced mouse elongating spermatids

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