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Transcriptome analysis of early and late pachytene spermatocytes in WT and Zfp541-KO mice (Smart-Seq2)

ABSTRACT: Transcriptome analysis of early and late pachytene spermatocytes in WT and Zfp541-KO mice (Smart-Seq2)

PROVIDER: PRJNA722288 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR14246771_1.fastq.gz	Fastqsanger.gz
	SRR14246771_2.fastq.gz	Fastqsanger.gz
	SRR14246772_1.fastq.gz	Fastqsanger.gz
	SRR14246772_2.fastq.gz	Fastqsanger.gz
	SRR14246773_1.fastq.gz	Fastqsanger.gz

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Transcriptome analysis of early and late pachytene spermatocytes in WT and Zfp541-KO mice (Smart-Seq2)

Project description:The DSB-machinery, which induces the programmed DNA double-strand breaks (DSBs) in leptotene and zygotene stages during meiosis, needs to be kept in silence after the initiation of pachytene stage to prevent the activation of DSB checkpoint that may lead to meiotic arrest or apoptosis of germ cells. However, the mechanisms underlying this repression remain largely unknown. Here, we report that ZFP541, a germ cell-specific zinc finger protein, is responsible for the suppression of DSBs formation at late pachytene. Lack of Zfp541 in mice leads to generation of DSBs in late pachytene spermatocytes by DSB formation related-proteins and causes male infertility due to meiotic failure. Plated-based scRNA-seq of Zfp541-/- spermatocytes revealed that ZFP541 negatively regulates many meiotic prophase genes, including genes for DSB formation and their upstream transcriptional regulators, in late pachytene spermatocytes. These results were confirmed by 10x single-cell RNA-seq data on spermatogenesis of Zfp541-/- testes, which suggested that Zfp541 is required for repressing the activation of pre-pachytene gene expression programs from early to late pachytene. ZFP541 ChIP-seq on pachytene and diplotene spermatocytes demonstrated that ZFP541 occupies the promoters of meiosis initiators (e.g., Meiosin and Rxra) and a subset of their downstream genes to repress their transcription, and thus prevent the reactivation of pre-pachytene gene expression programs in pachytene spermatocytes. Thus, our results not only revealed the role of ZFP541 in maintaining the repression of pre-pachytene transcriptional programs in pachytene spermatocytes but also provide new insight into the regulation of meiotic progression by timely turning off pre-pachytene genes.

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Characterization of ZFP541's occupency sites on genome in pachytene and diplotene spermatocytes

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Characterization of ZFP541's occupency sites on genome in pachytene and diplotene spermatocytes

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Expression profiling of isolated populations of prepachytene spermatocytes, pachytene spermatocytes and spermatids of PWD and B6 males

Project description:Expression profiling of isolated populations of prepachytene spermatocytes (LP), pachytene spermatocytes (RP) and spermatids (ST) from PWD and B6 was performed to study the genome wide variation in gene expression between two mouse subspecies. To evaluate the transcriptional difference between B6 and PWD in during meiosis, we compared their transcriptomes in sorted populations of pre-pachytene primary spermatocytes (Leptonema, Zygotene and Pachytene), pachytene spermatocytes (Mid-late pachytene and diplotene) and spermatids. Populations of pre-pachytene spermatocytes, pachytene spermatocytes and spermatids were isolated from PWD and B6 mice. All populations were isolated in duplicates from two animals. Hence, RNA was prepared from 12 samples (= 2 strains x 3 tissues x 2 replicas) and was then subject to labeling and hybridization on microarray chips.

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Transcriptome analysis of meiotic prophase spermatocytes in Zfp541 +/- and Zfp541 KO mice

Project description:ZFP541 is one of the candidates for a transcriptional regulator during mouse meiotic prophase. In order to address the role of ZFP541, transcriptomes of meiotic prophase-enriched population were compared between Zfp541 +/- and Zfp541 KO testes by RNA-seq. The meiotic prophase-enriched population was isolated from Zfp541 +/- and Zfp541 KO testes at postnatal day 18 (P18), on the Rec8-3FH-GFP KI background by fluorescent sorting of GFP positive cells.

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