TL;DR
This review summarizes studies investigating the roles of actin, myosin, and kinesin in mouse and human oocyte maturation and describes mechanisms underlying the involvement of contractile proteins in oocyte maturation.
NIMA-related kinase 2 (NEK2) is a serine/threonine kinase that plays crucial roles in cellular events such as centrosome separation, cell cycle progression, spindle assembly checkpoint (SAC) regulation, and microtubule stabilization. However, its regulatory function during oocyte meiotic maturation remains unclear. In this study, NEK2 activity was inhibited using the specific inhibitor MBM-55 to evaluate its role in oocyte meiotic maturation and early embryonic development. NEK2 depletion impaired multiple indicators associated with oocyte maturation and decreased the maturation rate. Mechanistic analysis suggested that NEK2 inhibition altered spindle organization and chromosome alignment, which may be associated with impaired kinetochore-microtubule (K-MT) interactions and altered SAC-associated signaling, collectively contributing to MI-stage arrest and increased aneuploidy. In addition, NEK2 deficiency impaired the actin network, reducing spindle migration, preventing the efficient repair of accumulated DNA damage and ultimately leading to apoptosis. NEK2 inhibition in oocytes reduced subsequent embryonic developmental competence, leading to a lower blastocyst formation rate. These findings demonstrate that NEK2 is a critical regulator of porcine oocyte meiotic maturation through its roles in SAC regulation, K-MT attachment, cytoskeletal dynamics, and the DNA damage response.
The ER is a complex network of membranes that inhabits much of the cytoplasm of cells – however, this network undergoes a massive condensation and rapid remodeling during cell division. In Drosophila cleavage divisions, this results in a tight association of the ER with centrosomes and mitotic spindle poles. Previous work has shown that this relationship between the ER and centrosomes must be finely tuned to enable successful spindle elongation, and that overaccumulation of the ER in these stages can result in failed centrosome maturation. During interphase, the ER exists in tubular and sheet-like arrangements, with a variety of “shaping” proteins enforcing these topologies. Here, we examine the contributions of these ER shaping proteins to the rapid changes that occur during cleavage mitoses in the Drosophila embryo. A screen of ER shaping proteins revealed that disruption of Reep-family proteins leads to mitotic failures at characteristic cleavage stages. Compromising ReepA, the Drosophila ortholog of the Reep1-4 subfamily, had a lesser impact on early embryonic mitoses. However, ReepB (the ortholog of the Reep5-6 subfamily) disruption, significantly affects ER mitotic coat morphologies, resulting in a ‘frilled ER’ phenotype and a reduction of ER adherence to the spindle space accompanied by division failures. Overexpressing ReepA does not rescue ReepB mitotic or ER morphology defects and instead introduces local condensates of abnormal ER structures. These data suggest that dedicated Reep proteins guide ER mitotic properties at specific early developmental stages. Using a cell-based in vitro analysis of Drosophila Reeps, we identify differential “tubulating” properties of ReepA and ReepB. Together these data suggest that the minutes-scale ER remodeling required for early mitoses is governed by shaping proteins, and that ReepB family members are especially important in some of the most rapid cleavage divisions that occur in early embryo.
Proper regulation of contraction and relaxation in biological tubes is essential for organismal function. In C. elegans, the spermatheca, composed of smooth muscle-like cells, undergoes repeated stretching and contraction as oocytes pass through. Here we describe PES-8, a previously uncharacterized protein, as a regulator of spermathecal contractility. PES-8 contains a predicted extracellular zona pellucida-like domain and an unstructured cytoplasmic tail, suggesting dual roles in extracellular and cytoplasmic signaling. PES-8 localizes to the plasma membrane of the spermatheca, the spermathecal-uterine valve, and uterus. Functional analysis shows that PES-8 is essential for spermathecal function; its loss disrupts actomyosin fiber alignment, FLN-1/filamin localization, apical junction organization, and Ca²⁺ signaling, preventing oocyte transit. These findings identify PES-8 as a key regulator of cytoskeletal organization and calcium-mediated contractility in the C. elegans spermatheca. [Media: see text] [Media: see text] [Media: see text] [Media: see text] [Media: see text] [Media: see text] [Media: see text] [Media: see text] [Media: see text] [Media: see text].
Myosin is a superfamily of motor proteins that has attracted extensive research interest for many years due to its ability to efficiently convert chemical energy into mechanical energy during muscle contraction and various other movements in eukaryotic cells. Non-muscle myosin II (NMII) consists of three members, NMIIA, NMIIB, and NMIIC, which are positioned at the downstream end of numerous signaling pathways and plays a central role in cell adhesion, migration, and division. During cell division, NMII serves as the primary protein that generates mechanical force. It participates not only in cell rounding from prophase to metaphase but also interacts with actin to assemble a contractile ring during late stages, which then contracts to split the mother cell into two daughter cells. Loss of NMII function may result in cell division failure and consequently tetraploidy. Subsequent division of tetraploid cells can generate aneuploidy, which is closely associated with tumorigenesis as shown in many studies. In this review, we will discuss the roles of NMII throughout the entire cell division cycle.
Oocytes rely on a cohort of proteins whose sustained expression ensures normal meiotic progression and reproductive competence throughout an animal's reproductive life. Age-related declines in these proteins are a major cause of reduced oocyte quality and female fertility during reproductive aging. Here, we report that the cohesin regulatory protein PDS5B, a dynamically maintained factor in oocytes, declines with age and plays a noncanonical role in the spindle pole formation independent of its cohesion function during oocyte meiotic maturation. Specifically, we found that PDS5B was expressed throughout the oocyte meiosis and localized at the spindle poles at metaphase stages, while its protein abundance was reduced in aged oocytes, concomitant with decreased messenger ribonucleic acid (mRNA) levels and translational efficiency. Knockdown or heterozygous knockout of PDS5B caused spindle assembly defects, meiotic arrest, and aneuploidy in oocytes, ultimately leading to female subfertility. Mechanistically, immunoprecipitation/mass spectrometry analyses revealed that PDS5B recruited deubiquitinating enzyme USP9X to spindle poles to stabilize nuclear mitotic apparatus and promote proper spindle assembly. Moreover, expression of exogenous PDS5B in aged oocytes partially alleviated meiotic defects associated with advanced maternal age. Altogether, our findings uncover a unique spindle pole-specific function of PDS5B in oocytes and suggest that maintaining PDS5B levels may be a potential strategy to improve the quality of aged oocytes.
Proteasomes are fundamental for protein homeostasis and genome integrity and essential in spermatogenesis and fertilization. However, their presence, composition and role within the sperm nucleus are a subject of debate. Here we use in situ cryo-electron tomography in human sperm cells to elucidate the molecular architecture of nuclear proteasomes, which cluster in DNA-free, nuclear cavities within the sperm nucleus. We show that the main population of proteasomes consists of 20S core particles, with a smaller fraction of 20S capped by PA200 activator. Using single-particle cryo-electron microscopy of purified native human sperm proteasomes, we elucidate the features of the essential testis-specific subunit α4s, reporting the presence of a unique splice variant. We resolve a native peptide in the catalytic β2 subunit, providing insight into the proteolysis mechanism and PA200-mediated enhancement of trypsin activity. We show nuclear enrichment of proteasomes during sperm-cell differentiation in human testis tissue, with 20S and PA200 clustering following meiosis, at the spermatid stage. Our findings shed light on the organization and compositional diversity of nuclear proteasomes in human sperm cells, as well as their catalytic function.
