bims-cebooc 2024-07-14 papers

bims-cebooc

Biomed News

on Cell biology of oocytes

Issue of 2024–07–14
ten papers selected by
Gabriele Zaffagnini, Centre for Genomic Regulation

Oocyte biology: Perhaps chromosomal glue can be reapplied.
High-sensitivity whole-mount in situ Hybridization of Mouse Oocytes and Embryos Visualizes the Super-resolution Structures and Distributions of mRNA Molecules.
The formation of aggregated chromatin/chromosomes in mouse oocytes treated with high concentration of IBMX as a model for a chromosome transfer in human.
Caenorhabditis elegans germ granules accumulate hundreds of low translation mRNAs with no systematic preference for germ cell fate regulators.
Formation and resolution of meiotic chromosome entanglements and interlocks.
Single-cell sequencing reveals the transcriptional alternations of 17β-estradiol suppressing primordial follicle formation in neonatal mouse ovaries.
Germ granule compartments coordinate specialized small RNA production.
Human DDX6 regulates translation and decay of inefficiently translated mRNAs.
Cohesin composition and dosage independently affect early development in zebrafish.
Roles of N6-methyladenosine writers, readers and erasers in the mammalian germline.

Curr Biol. 2024 Jul 08. pii: S0960-9822(24)00692-4. [Epub ahead of print]34(13): R628-R630

Oocyte biology: Perhaps chromosomal glue can be reapplied.

JoAnne Engebrecht.

Meiotic cohesion loss underlies elevated rates of infertility and chromosome abnormalities in children of older women. A new study shows that cohesins are turned over throughout meiotic prophase, suggesting that cohesion loss is likely not solely due to early establishment of cohesion.

DOI: https://doi.org/10.1016/j.cub.2024.05.049
Biol Proced Online. 2024 Jul 10. 26(1): 23

High-sensitivity whole-mount in situ Hybridization of Mouse Oocytes and Embryos Visualizes the Super-resolution Structures and Distributions of mRNA Molecules.

Takahiro Sanada, Tomoya Kotani.

  Mammalian oocytes accumulate more than ten thousand mRNAs, of which three to four thousand mRNAs are translationally repressed. The timings and sites of translational activation of these dormant mRNAs are crucial for promoting oocyte maturation and embryonic development. How these mRNAs are accumulated and distributed in oocytes is therefore a fundamental issue to be explored. A method that enables visualization of mRNA molecules with high resolution in a simple manner would be valuable for understanding how oocytes accumulate and regulate the dormant mRNAs. We have developed a highly sensitive whole-mount in situ hybridization method using in vitro-synthesized RNA probes and the tyramide signal amplification (TSA) system optimized for mouse oocytes and embryos. By using this method, Pou5f1/Oct4, Emi2, and cyclin B1 mRNAs were detected in immature oocytes and 2-cell stage embryos. Confocal microscopy showed that these mRNAs formed granular structures in the oocyte cytoplasm. The structures of Pou5f1/Oct4 and cyclin B1 mRNAs persisted in 2-cell stage embryos. Pou5f1/Oct4 RNA granules exhibited a solid-like property in immature oocytes and became liquid-like droplets in 2-cell stage embryos. Double-staining of cyclin B1 mRNA with Emi2 or Pou5f1/Oct4 mRNA revealed that these mRNAs were distributed as different RNA granules without overlapping each other and that the size of cyclin B1 RNA granules tended to be larger than that of Emi2 RNA granules. The structures and distribution patterns of these mRNAs were further analyzed by N-SIM super-resolution microscopy. This analysis revealed that the large-sized RNA granules consist of many small-sized granules, suggesting the accumulation and regulation of dormant mRNAs as basal-sized RNA granules. The method established in this study can easily visualize the structure and distribution of mRNAs accumulated in mammalian oocytes and embryos with high sensitivity and super-resolution. This method is useful for investigating the cellular and molecular mechanisms of translational control of mRNAs by which maturation and early developmental processes are promoted.

Keywords:  Embryo; Mammal; Maternal mRNA; Oocyte; Super-resolution microscopy; in situ hybridization

DOI:  https://doi.org/10.1186/s12575-024-00250-5
Syst Biol Reprod Med. 2024 Dec;70(1): 195-203

The formation of aggregated chromatin/chromosomes in mouse oocytes treated with high concentration of IBMX as a model for a chromosome transfer in human.

Wei Xiao, Sakura Akao, Ryota Okamoto, Junko Otsuki.

  The presence of cyclic adenosine monophosphate (cAMP) has been considered to be a fundamental factor in ensuring meiotic arrest prior to ovulation. cAMP is regarded as a key molecule in the regulation of oocyte maturation. However, it has been reported that increased levels of intracellular cAMP can result in abnormal cytokinesis, with some MI oocytes leading to symmetrically cleaved 2-cell MII oocytes. Consequently, we aimed to investigate the effects of elevated intracellular cAMP levels on abnormal cytokinesis and oocyte maturation during the meiosis of mouse oocytes. This study found that a high concentration of isobutylmethylxanthine (IBMX) also caused chromatin/chromosomes aggregation (AC) after the first meiosis. The rates of AC increased the greater the concentration of IBMX. In addition, AC formation was found to be reversible, showing that the re-formation of the spindle chromosome complex was possible after the IBMX was removed. In human oocytes, the chromosomes aggregate after the germinal vesicle breakdown and following the first and second polar body extrusions (the AC phase), while mouse oocytes do not have this AC phase. The results of our current study may indicate that the AC phase in human oocytes could be related to elevated levels of intracytoplasmic cAMP.

Keywords:  Cyclic adenosine monophosphate (cAMP); IBMX; chromatin; chromosome aggregation; meiosis

DOI:  https://doi.org/10.1080/19396368.2024.2368116
Development. 2024 Jul 01. pii: dev202575. [Epub ahead of print]151(13):

Caenorhabditis elegans germ granules accumulate hundreds of low translation mRNAs with no systematic preference for germ cell fate regulators.

Alyshia Scholl, Yihong Liu, Geraldine Seydoux.

  In animals with germ plasm, embryonic germline precursors inherit germ granules, condensates proposed to regulate mRNAs coding for germ cell fate determinants. In Caenorhabditis elegans, mRNAs are recruited to germ granules by MEG-3, a sequence non-specific RNA-binding protein that forms stabilizing interfacial clusters on germ granules. Using fluorescence in situ hybridization, we confirmed that 441 MEG-3-bound transcripts are distributed in a pattern consistent with enrichment in germ granules. Thirteen are related to transcripts reported in germ granules in Drosophila or Nasonia. The majority, however, are low-translation maternal transcripts required for embryogenesis that are not maintained preferentially in the nascent germline. Granule enrichment raises the concentration of certain transcripts in germ plasm but is not essential to regulate mRNA translation or stability. Our findings suggest that only a minority of germ granule-associated transcripts contribute to germ cell fate in C. elegans and that the vast majority function as non-specific scaffolds for MEG-3.

Keywords:   C. elegans; Germ granules; Germ plasm; Germline; MEG-3; Nanos

DOI:  https://doi.org/10.1242/dev.202575
J Cell Sci. 2024 Jul 01. pii: jcs262004. [Epub ahead of print]137(13):

Formation and resolution of meiotic chromosome entanglements and interlocks.

Iván Olaya, Sean M Burgess, Ofer Rog.

  Interactions between parental chromosomes during the formation of gametes can lead to entanglements, entrapments and interlocks between unrelated chromosomes. If unresolved, these topological constraints can lead to misregulation of exchanges between chromosomes and to chromosome mis-segregation. Interestingly, these configurations are largely resolved by the time parental chromosomes are aligned during pachytene. In this Review, we highlight the inevitability of topologically complex configurations and discuss possible mechanisms to resolve them. We focus on the dynamic nature of a conserved chromosomal interface - the synaptonemal complex - and the chromosome movements that accompany meiosis as potential mechanisms to resolve topological constraints. We highlight the advantages of the nematode Caenorhabditis elegans for understanding biophysical features of the chromosome axis and synaptonemal complex that could contribute to mechanisms underlying interlock resolution. In addition, we highlight advantages of using the zebrafish, Danio rerio, as a model to understand how entanglements and interlocks are avoided and resolved.

Keywords:   Caenorhabditis elegans ; Entanglement; Interlock; Meiosis; Synaptonemal complex; Zebrafish

DOI:  https://doi.org/10.1242/jcs.262004
Cell Prolif. 2024 Jul 10. e13713

Single-cell sequencing reveals the transcriptional alternations of 17β-estradiol suppressing primordial follicle formation in neonatal mouse ovaries.

Yutong Yan, Hui Zhang, Rui Xu, Linglin Luo, Lu Yin, Hao Wu, Yiqian Zhang, Chan Li, Sihai Lu, Yaju Tang, Xiaoe Zhao, Menghao Pan, Qiang Wei, Sha Peng, Baohua Ma.

Estrogen has been implicated in multiple biological processes, but the variation underlying estrogen-mediated primordial follicle (PF) formation remains unclear. Here, we show that 17β-estradiol (E2) treatment of neonatal mice led to the inhibition of PF formation and cell proliferation. Single-cell RNA sequencing (scRNA-seq) revealed that E2 treatment caused significant changes in the transcriptome of oocytes and somatic cells. E2 treatment disrupted the synchronised development of oocytes, pre-granulosa (PG) cells and stromal cells. Mechanistically, E2 treatment disrupted several signalling pathways critical to PF formation, especially down-regulating the Kitl and Smad1/3/4/5/7 expression, reducing the frequency and number of cell communication. In addition, E2 treatment influenced key gene expression, mitochondrial function of oocytes, the recruitment and maintenance of PG cells, the cell proliferation of somatic cells, as well as disordered the ovarian microenvironment. This study not only revealed insights into the regulatory role of estrogen during PF formation, but also filled in knowledge of dramatic changes in perinatal hormones, which are critical for the physiological significance of understanding hormone changes and reproductive protection.

DOI: https://doi.org/10.1111/cpr.13713
Nat Commun. 2024 Jul 10. 15(1): 5799

Germ granule compartments coordinate specialized small RNA production.

Xiangyang Chen, Ke Wang, Farees Ud Din Mufti, Demin Xu, Chengming Zhu, Xinya Huang, Chenming Zeng, Qile Jin, Xiaona Huang, Yong-Hong Yan, Meng-Qiu Dong, Xuezhu Feng, Yunyu Shi, Scott Kennedy, Shouhong Guang.

Germ granules are biomolecular condensates present in most animal germ cells. One function of germ granules is to help maintain germ cell totipotency by organizing mRNA regulatory machinery, including small RNA-based gene regulatory pathways. The C. elegans germ granule is compartmentalized into multiple subcompartments whose biological functions are largely unknown. Here, we identify an uncharted subcompartment of the C. elegans germ granule, which we term the E granule. The E granule is nonrandomly positioned within the germ granule. We identify five proteins that localize to the E granule, including the RNA-dependent RNA polymerase (RdRP) EGO-1, the Dicer-related helicase DRH-3, the Tudor domain-containing protein EKL-1, and two intrinsically disordered proteins, EGC-1 and ELLI-1. Localization of EGO-1 to the E granule enables synthesis of a specialized class of 22G RNAs, which derive exclusively from 5' regions of a subset of germline-expressed mRNAs. Defects in E granule assembly elicit disordered production of endogenous siRNAs, which disturbs fertility and the RNAi response. Our results define a distinct subcompartment of the C. elegans germ granule and suggest that one function of germ granule compartmentalization is to facilitate the localized production of specialized classes of small regulatory RNAs.

DOI: https://doi.org/10.1038/s41467-024-50027-3
Elife. 2024 Jul 11. pii: RP92426. [Epub ahead of print]13

Human DDX6 regulates translation and decay of inefficiently translated mRNAs.

Ramona Weber, Chung-Te Chang.

  Recent findings indicate that the translation elongation rate influences mRNA stability. One of the factors that has been implicated in this link between mRNA decay and translation speed is the yeast DEAD-box helicase Dhh1p. Here, we demonstrated that the human ortholog of Dhh1p, DDX6, triggers the deadenylation-dependent decay of inefficiently translated mRNAs in human cells. DDX6 interacts with the ribosome through the Phe-Asp-Phe (FDF) motif in its RecA2 domain. Furthermore, RecA2-mediated interactions and ATPase activity are both required for DDX6 to destabilize inefficiently translated mRNAs. Using ribosome profiling and RNA sequencing, we identified two classes of endogenous mRNAs that are regulated in a DDX6-dependent manner. The identified targets are either translationally regulated or regulated at the steady-state-level and either exhibit signatures of poor overall translation or of locally reduced ribosome translocation rates. Transferring the identified sequence stretches into a reporter mRNA caused translation- and DDX6-dependent degradation of the reporter mRNA. In summary, these results identify DDX6 as a crucial regulator of mRNA translation and decay triggered by slow ribosome movement and provide insights into the mechanism by which DDX6 destabilizes inefficiently translated mRNAs.

Keywords:  CCR4-NOT complex; DDX6; biochemistry; chemical biology; chromosomes; codon optimization; gene expression; human; mRNA decay; ribosome stalling

DOI:  https://doi.org/10.7554/eLife.92426
Development. 2024 Jul 08. pii: dev.202593. [Epub ahead of print]

Cohesin composition and dosage independently affect early development in zebrafish.

Anastasia A Labudina, Michael Meier, Gregory Gimenez, David Tatarakis, Sarada Ketharnathan, Bridget Mackie, Thomas F Schilling, Jisha Antony, Julia A Horsfield.

  Cohesin, a chromatin-associated protein complex with four core subunits (Smc1a, Smc3, Rad21 and either Stag1 or 2), has a central role in cell proliferation and gene expression in metazoans. Human developmental disorders termed "cohesinopathies" are characterised by germline mutations in cohesin or its regulators that do not entirely eliminate cohesin function. However, it is not clear if mutations in individual cohesin subunits have independent developmental consequences. Here we show that zebrafish rad21 or stag2b mutants independently influence embryonic tailbud development. Both mutants have altered mesoderm induction, but only homozygous or heterozygous rad21 mutation affects cell cycle gene expression. stag2b mutants have narrower notochords and reduced Wnt signaling in neuromesodermal progenitors as revealed by single cell RNA-sequencing. Stimulation of Wnt signaling rescues transcription and morphology in stag2b, but not rad21 mutants. Our results suggest that mutations altering the quantity versus composition of cohesin have independent developmental consequences, with implications for the understanding and management of cohesinopathies.

Keywords:  Cell fate; Cohesin; Rad21; Stag2; Zebrafish

DOI:  https://doi.org/10.1242/dev.202593
Curr Opin Genet Dev. 2024 Jul 08. pii: S0959-437X(24)00073-X. [Epub ahead of print]87 102224

Roles of N6-methyladenosine writers, readers and erasers in the mammalian germline.

Graeme R Wells, Ramesh S Pillai.

N6-methyladenosine (m6A) is the most abundant internal modification of mRNAs in eukaryotes. Numerous studies have shown that m6A plays key roles in many biological and pathophysiological processes, including fertility. The factors involved in m6A-dependent mRNA regulation include writers, which deposit the m6A mark, erasers, which remove it, and readers, which bind to m6A-modified transcripts and mediate the regulation of mRNA fate. Many of these proteins are highly expressed in the germ cells of mammals, and some have been linked to fertility disorders in human patients. In this review, we summarise recent findings on the important roles played by proteins involved in m6A biology in mammalian gametogenesis and fertility. Continued study of the m6A pathway in the mammalian germline will shed further light on the importance of epitranscriptomics in reproduction and may lead to effective treatment of human fertility disorders.

DOI: https://doi.org/10.1016/j.gde.2024.102224