Control of Post-Meiotic Gene Expression by H2A.L.2
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE284749 | GEO | 2026/01/05
REPOSITORIES: GEO
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Project description:Spermiogenesis is marked by an almost genome-wide histone-to-protamine transition. Protamines are sperm-specific, small and highly positively charged proteins mainly responsible for genome compaction at this step. According to previous investigations from our lab and other groups, it is known that just before their removal, histones become hyperacetylated, which creates binding sites for the first bromodomain of a testis-specific member of the BET double bromodomain protein family, Brdt. Brdt’s first bromodomain is necessary for transition proteins (TPs) and PRMs incorporation and subsequent histone removal. Histone hyperacetylation, specifically on H4 lysine residues 5 and 8 which are recognized by Brdt, is controlled by a post-meiotic enhancer of CBP/p300 acetyltransferase activity, known as Nut. The process of TP assembly itself is dependent on a prior incorporation of a histone variant known as H2A.L.2, expressed in the same cells and at the same time as TPs. To characterize the proteins associating with TP2, we performed a mass spectrometry-based proteomics analysis of TP2 co-immunoprecipitations elutes from condensing spermatids extract. The immunoprecipitation was performed using extract from stages 12-16 spermatids from wild-type and H2A.L.2-KO mice. In parallel, immunopurifications were performed using a non-specific antibody (pre-immune goat serum) to allow the identification of proteins specifically purified with TP2 as opposed to proteins non-specifically precipitated.
2022-02-14 | PXD031665 | Pride
Project description:Spermatogenesis is a complex process of sperm generation, including mitosis, meiosis, and spermiogenesis. During spermiogenesis, histones in post-meiotic spermatids are removed from chromatin and replaced by protamines. Although histone-to-protamine exchange is important for sperm nuclear condensation, the underlying regulatory mechanism is still poorly understood. Here, we identify PHD finger protein 7 (PHF7) as an E3 ubiquitin ligase for histone H3K14 in post-meiotic spermatids. Generation of Phf7-deficient mice and Phf7 C160A knockin mice with impaired E3 ubiquitin ligase activity reveals defects in histone-to-protamine exchange caused by dysregulation of histone removal factor Bromodomain, testis-specific (BRDT) in early condensing spermatids. Surprisingly, E3 ubiquitin ligase activity of PHF7 on histone ubiquitination leads to stabilization of BRDT by attenuating ubiquitination of BRDT. Collectively, our findings identify PHF7 as a critical factor for sperm chromatin condensation and contribute to mechanistic understanding of fundamental phenomenon of histone-to-protamine exchange and potential for drug development for the male reproduction system.
2020-07-28 | GSE136752 | GEO
Project description:Histone replacement by transition proteins (TPs) and protamines (Prms) constitutes an essential step for the successful production of functional male gametes, yet nothing is known on the underlying functional interplay between histones, TPs and Prms. Here, by studying spermatogenesis in the absence of H2A.L.2, a spermatid-specific histone variant, we identify a new step, which is essential for the transformation of nucleosomes to nucleoprotamines. H2A.L.2, synthesized at the same time as TPs, enables the invasion of nucleosomes by TPs, which themselves drive the recruitment and processing of Prms. H2A.L.2 incorporation initiates a series of successive transitional states that ultimately shift to a more stable state characterized by nucleoprotamine assembly and histone-TP displacement, leading to the generation of mature spermatozoa. Hence, the current view of histone-to-nucleoprotein transition should be revisited and include a new step with H2A.L.2 assembly prior to the action of TPs and Prms.
2017-04-24 | PXD005863 | Pride
Project description:Unique chromatin remodeling factors orchestrate dramatic changes in nuclear morphology during differentiation of the mature sperm head. A critical step in this process is histone-to-protamine exchange, which must be executed correctly to avoid sperm DNA damage, embryonic lethality, and male sterility. Here, we define an essential role for the histone methyltransferase DOT1L in the histone-to-protamine transition. We show that DOT1L is abundantly expressed in meiotic and postmeiotic germ cells and that methylation of histone H3 lysine 79 (H3K79), the modification catalyzed by DOT1L, is enriched in developing spermatids in the initial stages of histone replacement. Elongating spermatids lacking DOT1L fail to fully replace histones and exhibit aberrant protamine recruitment, resulting in deformed sperm heads and male sterility. Loss of DOT1L results in transcriptional dysregulation coinciding with the onset of histone replacement and affecting genes required for histone-to-protamine exchange. DOT1L also deposits H3K79me2 and promotes accumulation of elongating RNA Polymerase II at the testis-specific bromodomain gene Brdt. Together, our results indicate that DOT1L is an important mediator of transcription during spermatid differentiation and an indispensable regulator of male fertility.
2023-03-22 | GSE217931 | GEO
Project description:Histone ubiquitination has been suggested to serve as a “tag” for nucleosome removal during histone-to-protamine exchange that is essential for chromatin packaging in round spermatids. Here, we screen for putative E3 ligase and identify that the PHF7, containing both RING finger and PHD domains, is critical for H2A ubiquitination and histone removal. Mechanistically, its PHD domain as a histone code reader can specifically bind H3K4me3/me2 and its RING domain as a histone writer can ubiquinate H2A. PHF7 deficiency results in male infertility in mice by generating Phf7 knockout mice using CRISPR-Cas9 technology. Futhermore, we find impaired histone-to-protamine exchange leads to reduced chromatin compaction in Phf7 knockout sperm. Immunostaining and western blot further confirme abnormal retention of core histones and reduced levels of protamine PRM1 and PRM2 in Phf7 knockout sperm. In order to exclude the possibility that Phf7 regulate the expession of protamine, we sorte the round spermatids from WT and Phf7 knockout females for RNA-seq. The transcriptomes in round spermatids are very similar between WT and Phf7 knockout mice. There are few differentially expressed genes in the two groups and the expression of protamine is also unchanged even if PHF7 deficiency. Therefore we find PHF7 has no effects on gene regulation in histone-to-protamine exchange.
2019-07-05 | GSE119701 | GEO
Project description:Histone replacement by small basic proteins, transition proteins (TPs) and protamines (Prms) constitutes an essential step for the successful production of male gametes, yet nothing is known on the underlying functional interplay between histones, TPs and Prms. By studying spermatogenesis in the absence of H2A.L.2, a spermatid-specific histone variant, we identify a new step essential for the "metamorphosis" of nucleosomes to nucleoprotamines. H2A.L.2 plays the role of a "Trojan horse" histone, which induces dramatic structural changes that enables the invasion of nucleosomes by TPs, themselves driving the processing and incorporation of Prms. H2A.L.2 incorporation initiates a series of rapidly changing transitional states that ultimately shift to a more stable state characterized by nucleoprotamine assembly and histone-TP Powered by Editorial Manager® and ProduXion Manager® from Aries Systems Corporationdisplacement, leading to the generation of mature spermatozoa. The textbook view of histone-to-nucleoprotein transition needs to be revisited to include a new step with H2A.L.2 assembly prior to the action of TPs and Prms.
2017-05-10 | GSE93251 | GEO
Project description:Spermatozoa have a unique genome organization: their chromatin is almost completely devoid of histones and is formed instead of protamines which confer a high level of compaction and preserve paternal genome integrity until fertilization. Histone-to-protamine transition takes place in spermatids and is indispensable for the production of functional sperm. Here we show that the H3K79-methyltransferase DOT1L controls spermatid chromatin remodelling and subsequent reorganization and compaction of spermatozoon genome. Using a mouse model in which Dot1l is knocked-out (KO) in postnatal male germ cells, we found that Dot1l-KO sperm chromatin is less compact and has an abnormal content, characterized by the presence of transition proteins, immature protamine 2 forms and a higher level of histones. Proteomics and transcriptomics analyses performed on spermatids reveal that Dot1l-KO modifies the chromatin prior to histone removal, and leads to the deregulation of genes involved in flagellum formation and apoptosis during spermatid differentiation. As a consequence of these chromatin and gene expression defects, Dot1l-KO spermatozoa have less compact heads and are less motile, which results in impaired fertility.
2023-03-30 | PXD030734 | Pride
Project description:Histone hyperacetylation is thought to drive the replacement of histones by transition proteins that occurs in elongating spermatids after a general shut-down of transcription. The molecular machineries underlying this histone hyperacetylation remain still undefined. Here we focused our attention on the role of Cbp and p300 in histone hyperacetylation and in the preceding late gene transcriptional activity in elongating spermatids. A strategy was designed to partially deplete Cbp and p300 in elongating spermatids. These cells progressed normally through spermiogenesis and showed normal histone hyperacetylation and removal. However, a genome-wide transcriptomic analysis, performed in the round and elongating spermatids, revealed the existence of a gene regulatory circuit encompassing genes presenting high expression levels in pre-meiotic cells, undergoing a repressed state in spermatocytes and early post-meiotic cells, but becoming reactivated in elongating spermatids, just prior to the global shut-down of transcription. Interestingly, this group of genes was over-represented within the genes affected by Cbp/p300 knock-down and were all involved in metabolic remodelling. This study revealed the occurrence of a tightly regulated Cbp/p300-dependent gene expression program that drives a specific metabolic state both in progenitor spermatogenic cells and in late transcriptionally active spermatids and confirmed a special link between Cpb/p300 and cell metabolism programming previously shown in somatic cells. Total RNA were obtained from post meiotic male germ cells (round spermatids and condensing spermatids) of adult mice either wild-type or with a double knock down of Cbp/P300 in post-meiotic cells. In each experiments, 6 replicates of each genotype and cell type were used.
2014-03-12 | E-GEOD-55767 | biostudies-arrayexpress
Project description:During spermatogenesis, intricate gene expression is coordinately regulated by epigenetic modifiers, which are required for differentiation of spermatogonial stem cells (SSCs) contained among undifferentiated spermatogonia. We previously found that KMT2B conveys H3K4me3 at bivalent and monovalent promoters in undifferentiated spermatogonia. Because these genes are expressed late in spermatogenesis or during embryogenesis, we expect that many of them are potentially programmed by KMT2B for future expression. Here, we show that one of the genes targeted by KMT2B, Tsga8, plays an essential role in spermatid morphogenesis. Loss of Tsga8 in mice leads to male infertility associated with abnormal chromosomal distribution in round spermatids, malformation of elongating spermatid heads and spermiation failure. Tsga8 depletion leads to dysregulation of thousands of genes, including the X chromosome genes that are reactivated in spermatids, and insufficient nuclear condensation accompanied by reductions of TNP1 and PRM1, key factors for histone-to-protamine transition. Intracytoplasmic sperm injection (ICSI) of spermatids rescued the infertility phenotype, suggesting competency of the spermatid genome for fertilization. Thus, Tsga8 is a KMT2B target that is vitally necessary for spermiogenesis and fertility.
2021-04-16 | GSE147805 | GEO
Project description:Protamines replace histones as the main nuclear protein in sperm cells having a crucial role in compacting the paternal genome. Human spermatozoa contain the protamine 1 (P1) and the family of protamine 2 (P2) proteins. Alterations in protamine PTMs or on the P1/P2 ratio could be associated with male infertility. Top-down proteomics enables large-scale analysis of intact proteforms derived from alternative splicing, missense or nonsense genetic variants or PTMs. In contrast to current gold standard techniques, top-down allows a more in-depth analysis of protamine PTMs and proteoforms, opening up new perspectives to unravel their impact on male fertility. We analyzed two normozoospermic semen samples by top-down and discussed in detail the difficulties we found in the data analysis and the suggested solutions, as this step is one of the current bottlenecks in top-down proteomics with currently available bioinformatic tools. Our strategy for the data analysis combines two different software, ProSight PD (PS) and TopPIC suite (TP), with a clustering algorithm to decipher protamine proteoforms. We identified up to 32 protamine proteoforms at different levels of characterization. This in-depth analysis of the protamine proteoform landscape of an individual boosts personalized diagnosis of male infertility.
2021-05-13 | PXD024405 | Pride