Aberrant cortex contractions impact mammalian oocyte quality.

The cortex controls cell shape. In mouse oocytes, the cortex thickens in an Arp2/3-complex-dependent manner, ensuring chromosome positioning and segregation. Surprisingly, we identify that mouse oocytes lacking the Arp2/3 complex undergo cortical actin remodeling upon division, followed by cortical contractions that are unprecedented in mammalian oocytes. Using genetics, imaging, and machine learning, we show that these contractions stir the cytoplasm, resulting in impaired organelle organization and activity. Oocyte capacity to avoid polyspermy is impacted, leading to a reduced female fertility. We could diminish contractions and rescue cytoplasmic anomalies. Similar contractions were observed in human oocytes collected as byproducts during IVF (in vitro fertilization) procedures. These contractions correlate with increased cytoplasmic motion, but not with defects in spindle assembly or aneuploidy in mice or humans. Our study highlights a multiscale effect connecting cortical F-actin, contractions, and cytoplasmic organization and affecting oocyte quality, with implications for female fertility.

Meiosis: Actin and microtubule networks drive chromosome clustering in oocytes.

Actin and microtubule networks in human and porcine oocytes sequentially gather chromosomes in a cluster shortly after nuclear envelope breakdown to ensure their complete capture by the meiotic spindle.

Bypassing Mendel's First Law: Transmission Ratio Distortion in Mammals.

Mendel's law of segregation states that the two alleles at a diploid locus should be transmitted equally to the progeny. A genetic segregation distortion, also referred to as transmission ratio distortion (TRD), is a statistically significant deviation from this rule. TRD has been observed in several mammal species and may be due to different biological mechanisms occurring at diverse time points ranging from gamete formation to lethality at post-natal stages. In this review, we describe examples of TRD and their possible mechanisms in mammals based on current knowledge. We first focus on the differences between TRD in male and female gametogenesis in the house mouse, in which some of the most well studied TRD systems have been characterized. We then describe known TRD in other mammals, with a special focus on the farmed species and in the peculiar common shrew species. Finally, we discuss TRD in human diseases. Thus far, to our knowledge, this is the first time that such description is proposed. This review will help better comprehend the processes involved in TRD. A better understanding of these molecular mechanisms will imply a better comprehension of their impact on fertility and on genome evolution. In turn, this should allow for better genetic counseling and lead to better care for human families.

Protocol to measure cleavage efficiency of the meiotic cohesin subunit Rec8 by separase in mouse oocytes using a biosensor.

Here, we describe a biosensor to assess meiotic cohesin subunit Rec8 cleavage in mouse oocytes. We detail oocyte collection and microinjection of the mRNA expressing the biosensor. The biosensor is targeted to chromosomes and consists of two fluorophores flanking a Rec8 fragment containing separase cleavage sites. Cleavage leads to dissociation of one fluorophore from chromosomes, and the efficiency can be estimated by live imaging. We detail the use of this biosensor in mouse oocytes with or without Aurora B/C inhibitor. For complete details on the use and execution of this protocol, please refer to Nikalayevich et al. (2022).

An interpretable and versatile machine learning approach for oocyte phenotyping.

Meiotic maturation is a crucial step of oocyte formation, allowing its potential fertilization and embryo development. Elucidating this process is important for both fundamental research and assisted reproductive technology. However, few computational tools based on non-invasive measurements are available to characterize oocyte meiotic maturation. Here, we develop a computational framework to phenotype oocytes based on images acquired in transmitted light. We trained neural networks to segment the contour of oocytes and their zona pellucida using oocytes from diverse species. We defined a comprehensive set of morphological features to describe an oocyte. These steps were implemented in an open-source Fiji plugin. We present a feature-based machine learning pipeline to recognize oocyte populations and determine morphological differences between them. We first demonstrate its potential to screen oocytes from different strains and automatically identify their morphological characteristics. Its second application is to predict and characterize the maturation potential of oocytes. We identify the texture of the zona pellucida and cytoplasmic particle size as features to assess mouse oocyte maturation potential and tested whether these features were applicable to the developmental potential of human oocytes. This article has an associated First Person interview with the first author of the paper.

Aurora B/C-dependent phosphorylation promotes Rec8 cleavage in mammalian oocytes.

To generate haploid gametes, cohesin is removed in a stepwise manner from chromosome arms in meiosis I and the centromere region in meiosis II to segregate chromosomes and sister chromatids, respectively. Meiotic cohesin removal requires cleavage of the meiosis-specific kleisin subunit Rec8 by the protease separase.1,2 In yeast and C. elegans, Rec8 on chromosome arms has to be phosphorylated to be cleaved in meiosis I,3-7 whereas Rec8 at the centromere is protected from cleavage by the action of PP2A-B56.8-10 However, in mammalian meiosis, it is unknown whether Rec8 has to be equally phosphorylated for cleavage, and if so, the identity of the relevant kinase(s). This is due to technical challenges, as Rec8 is poorly conserved, preventing a direct translation of the knowledge gained from model systems such as yeast and C. elegans to mammals. Additionally, there is no turnover of Rec8 after cohesion establishment, preventing phosphomutant analysis of functional Rec8. To address the very basic question of whether Rec8 cleavage requires its phosphorylation in mammals, we adapted a biosensor that detects separase activity to study Rec8 cleavage in single mouse oocytes by live imaging. Crucially, through phosphomutant analysis, we identified phosphorylation sites in Rec8 promoting cleavage. We found that Rec8 cleavage depends on Aurora B/C kinase activities and identified an aminoacid residue that is phosphorylated in vivo. Accordingly, inhibition of Aurora B/C kinases during meiotic maturation impairs endogenous Rec8 phosphorylation and chromosome segregation.

Selfish centromeres, selfless heterochromatin.

Centromeres are specialized regions on chromosomes recruiting a set of proteins required for faithful chromosome segregation. Differences in centromere strength can potentially bias chromosome segregation toward one of the daughter cells during division. Kumon et al. propose a new model of evolutionary impact on the balance of centromere strength.

Artificially decreasing cortical tension generates aneuploidy in mouse oocytes.

Human and mouse oocytes' developmental potential can be predicted by their mechanical properties. Their development into blastocysts requires a specific stiffness window. In this study, we combine live-cell and computational imaging, laser ablation, and biophysical measurements to investigate how deregulation of cortex tension in the oocyte contributes to early developmental failure. We focus on extra-soft cells, the most common defect in a natural population. Using two independent tools to artificially decrease cortical tension, we show that chromosome alignment is impaired in extra-soft mouse oocytes, despite normal spindle morphogenesis and dynamics, inducing aneuploidy. The main cause is a cytoplasmic increase in myosin-II activity that could sterically hinder chromosome capture. We describe here an original mode of generation of aneuploidies that could be very common in oocytes and could contribute to the high aneuploidy rate observed during female meiosis, a leading cause of infertility and congenital disorders.

Detection of Separase Activity Using a Cleavage Sensor in Live Mouse Oocytes.

Separase proteolytically removes cohesin complexes from sister chromatid arms in meiosis I, which is essential for chromosome segregation. Regulation of separase activity is essential for proper cell cycle progression and correct chromosome segregation. Onset of endogenous separase activity has not yet been observed in live oocytes.We describe here a method for detecting separase activity in mouse oocytes in vivo. This method utilizes a previously described cleavage sensor made up of H2B-mCherry fused with Scc1(107-268 aa)-YFP. The cleavage sensor is loaded on the chromosomes through its H2B-tag, and the signal from both mCherry and YFP is visible. Upon separase activation the Scc1 fragment is cleaved and YFP dissociates from the chromosomes. The change in the ratio between mCherry and YFP fluorescence intensity is a readout of separase activity.

The NuRD nucleosome remodelling complex and NHK-1 kinase are required for chromosome condensation in oocytes.

Chromosome condensation during cell division is one of the most dramatic events in the cell cycle. Condensin and topoisomerase II are the most studied factors in chromosome condensation. However, their inactivation leads to only mild defects and little is known about the roles of other factors. Here, we took advantage of Drosophilaoocytes to elucidate the roles of potential condensation factors by performing RNA interference (RNAi). Consistent with previous studies, depletion of condensin I subunits or topoisomerase II in oocytes only mildly affected chromosome condensation. In contrast, we found severe undercondensation of chromosomes after depletion of the Mi-2-containing NuRD nucleosome remodelling complex or the protein kinase NHK-1 (also known as Ballchen in Drosophila). The further phenotypic analysis suggests that Mi-2 and NHK-1 are involved in different pathways of chromosome condensation. We show that the main role of NHK-1 in chromosome condensation is to phosphorylate Barrier-to-autointegration factor (BAF) and suppress its activity in linking chromosomes to nuclear envelope proteins. We further show that NHK-1 is important for chromosome condensation during mitosis as well as in oocytes.