Search Results (8)

In vitro maturation (IVM) of oocytes retrieved from ovum pick-up (OPU) or ovarian tissue (OT) is a standard approach for patients with specific conditions where prior hormonal stimulation is contraindicated. However, the developmental competence of oocytes matured in vitro is still inferior to that of oocytes matured in vivo. Capacitation IVM (CAPA-IVM) includes an extra step of pre-maturation culture (PMC) with c-type natriuretic peptide (CNP) as a meiotic arrestor to better synchronize cytoplasmic and nuclear maturity in oocytes by allowing the cytoplasm additional time to acquire essential components critical for optimal competency. This study aims to evaluate the effect of CAPA-IVM on equine oocyte quality and developmental competence. Immature cumulus–oocyte complexes (COCs) were retrieved from slaughterhouse ovaries and matured in vitro either in CAPA-IVM (short 6 h, long 24 h pre-maturation) or standard IVM. Mature oocytes from each group were analyzed for calcium-releasing potential (n = 52) and single-oocyte proteomics (n = 44), and embryo development (n = 229) was assessed after fertilization with piezo-drilled intracytoplasmic sperm injection (ICSI). Genetic analysis of developed blastocysts (n = 41) was performed to detect chromosomal aberrations. Our findings demonstrate that CAPA-IVM of equine COCs yields significantly higher maturation rates than controls. Moreover, short CAPA-IVM with six hours pre-maturation culture showed substantially higher embryo development potential than the control group (20/69 vs. 9/63, respectively). Genetic analysis revealed a high euploidy rate in equine blastocysts regardless of the maturation conditions. Live calcium imaging of the fertilized oocytes demonstrated that the majority of oocytes displayed non-continuous calcium oscillation patterns, irrespective of maturation conditions. Single-oocyte proteomics reveals a comparable proteomic landscape between mature oocytes subjected to short CAPA-IVM and standard IVM. However, we identified four enriched gene sets with positive enrichment scores after short CAPA-IVM, related to cytoskeleton regulation, ribosomal function, and cytosolic components. Our findings indicate that CAPA-IVM holds the potential to improve oocyte quality and competence in horses. However, further fine-tuning of culture conditions would benefit the effective use of these IVM systems. Moreover, given that the mare serves as an excellent model for human reproduction, the molecular trends identified in this study could provide valuable insights for advancing human artificial reproductive technologies. Full article

Chromosomal instability (CIN) is commonly seen in cancer cells, and related to tumor progression and poor prognosis. Among the causes of CIN, insufficient correction of erroneous kinetochore (KT)-microtubule (MT) attachments plays pivotal roles in various situations. In this review, we focused on the previously unappreciated role of chromosome oscillation in the correction of erroneous KT-MT attachments, and its relevance to the etiology of CIN. First, we provided an overview of the error correction mechanisms for KT-MT attachments, especially the role of Aurora kinases in error correction by phosphorylating Hec1, which connects MT to KT. Next, we explained chromosome oscillation and its underlying mechanisms. Then we introduced how chromosome oscillation is involved in the error correction of KT-MT attachments, based on recent findings. Chromosome oscillation has been shown to promote Hec1 phosphorylation by Aurora A which localizes to the spindle. Finally, we discussed the link between attenuated chromosome oscillation and CIN in cancer cells. This link underscores the role of chromosome dynamics in mitotic fidelity, and the mutual relationship between defective chromosome dynamics and CIN in cancer cells that can be a target for cancer therapy. Full article

Cell proliferation and sexual reproduction require the faithful segregation of chromosomes. Chromosome segregation is driven by the interaction of chromosomes with the spindle, and the attachment of chromosomes to the proper spindle poles is essential. Initial attachments are frequently erroneous due to the random nature of the attachment process; however, erroneous attachments are selectively eliminated. Proper attachment generates greater tension at the kinetochore than erroneous attachments, and it is thought that attachment selection is dependent on this tension. However, studies of meiotic chromosome segregation suggest that attachment elimination cannot be solely attributed to tension, and the precise mechanism of selective elimination of erroneous attachments remains unclear. During attachment elimination, chromosomes oscillate between the spindle poles. A recent study on meiotic chromosome segregation in fission yeast has suggested that attachment elimination is coupled to chromosome oscillation. In this review, the possible contribution of chromosome oscillation in the elimination of erroneous attachment is discussed in light of the recent finding. Full article

Kv10.1 (potassium voltage-gated channel subfamily H member 1, known as EAG1 or Ether-à-go-go 1), is a voltage-gated potassium channel, prevailingly expressed in the central nervous system. The aberrant expression of Kv10.1 is detected in over 70% of all human tumor tissues and correlates with poorer prognosis. In peripheral tissues, Kv10.1 is expressed almost exclusively during the G2/M phase of the cell cycle and regulates its progression—downregulation of Kv10.1 extends the duration of the G2/M phase both in cancer and healthy cells. Here, using biochemical and imaging techniques, such as live-cell measurements of microtubule growth and of cytosolic calcium, we elucidate the mechanisms of Kv10.1-mediated regulation at the G2/M phase. We show that Kv10.1 has a dual effect on mitotic microtubule dynamics. Through the functional interaction with ORAI1 (calcium release-activated calcium channel protein 1), it modulates cytosolic calcium oscillations, thereby changing microtubule behavior. The inhibition of either Kv10.1 or ORAI1 stabilizes the microtubules. In contrast, the knockdown of Kv10.1 increases the dynamicity of mitotic microtubules, resulting in a stronger spindle assembly checkpoint, greater mitotic spindle angle, and a decrease in lagging chromosomes. Understanding of Kv10.1-mediated modulation of the microtubule architecture will help to comprehend how cancer tissue benefits from the presence of Kv10.1, and thereby increase the efficacy and safety of Kv10.1-directed therapeutic strategies. Full article

Microtubules are hollow cylindrical polymers composed of the highly negatively-charged (~23e), high dipole moment (1750 D) protein α, β- tubulin. While the roles of microtubules in chromosomal segregation, macromolecular transport, and cell migration are relatively well-understood, studies on the electrical properties of microtubules have only recently gained strong interest. Here, we show that while microtubules at physiological concentrations increase solution capacitance, free tubulin has no appreciable effect. Further, we observed a decrease in electrical resistance of solution, with charge transport peaking between 20–60 Hz in the presence of microtubules, consistent with recent findings that microtubules exhibit electric oscillations at such low frequencies. We were able to quantify the capacitance and resistance of the microtubules (MT) network at physiological tubulin concentrations to be 1.27 × 10⁻⁵ F and 9.74 × 10⁴ Ω. Our results show that in addition to macromolecular transport, microtubules also act as charge storage devices through counterionic condensation across a broad frequency spectrum. We conclude with a hypothesis of an electrically tunable cytoskeleton where the dielectric properties of tubulin are polymerisation-state dependent. Full article

During the cell cycle, hundreds of proteins become phosphorylated and dephosphorylated, indicating that protein kinases and protein phosphatases play a central role in its regulation. It has been widely recognized that oscillation in cyclin-dependent kinase (CDK) activity promotes DNA replication, during S-phase, and chromosome segregation, during mitosis. Each CDK substrate phosphorylation status is defined by the balance between CDKs and CDK-counteracting phosphatases. In fission yeast and animal cells, PP2A/B55 is the main protein phosphatase that counteracts CDK activity. PP2A/B55 plays a key role in mitotic entry and mitotic exit, and it is regulated by the Greatwall-Endosulfine (ENSA) molecular switch that inactivates PP2A/B55 at the onset of mitosis, allowing maximal CDK activity at metaphase. The Greatwall-ENSA-PP2A/B55 pathway is highly conserved from yeast to animal cells. In yeasts, Greatwall is negatively regulated by nutrients through TORC1 and S6 kinase, and couples cell growth, regulated by TORC1, to cell cycle progression, driven by CDK activity. In animal cells, Greatwall is phosphorylated and activated by Cdk1 at G2/M, generating a bistable molecular switch that results in full activation of Cdk1/CyclinB. Here we review the current knowledge of the Greatwall-ENSA-PP2A/B55 pathway and discuss its role in cell cycle progression and as an integrator of nutritional cues. Full article

Environmental and occupational exposure to benzene from fuels is a major cause for concern for national and international authorities, as benzene is a known carcinogen in humans and there is no safe limit for exposure to carcinogens. The objective of this study was to evaluate the genotoxic effects of chronic occupational exposure to benzene among two groups of workers: filling station workers (Group I) and security guards working at vehicles entrances (Group II), both on the same busy highway in Rio de Janeiro, Brazil. Sociodemographic data on the workers were evaluated; the concentration of benzene/toluene (B/T) in atmospheric air and individual trans,trans-muconic acid (ttMA) and S-phenylmercapturic acid (S-PMA) were measured; oxidative stress was analyzed by catalase (CAT), glutathione S-transferase (GST), superoxide dismutase (SOD), thiol groups (THIOL) and malondialdehyde (MDA); genotoxicity was measured by metaphases with chromosomal abnormalities (MCA) and nuclear abnormalities, comet assay using the enzyme formamidopyrimidine DNA glycosylase (C-FPG), and methylation of repetitive element LINE-1, CDKN2B and KLF6 genes. Eighty-six workers participated: 51 from Group I and 35 from Group II. The B/T ratio was similar for both groups, but Group I had greater oscillation of benzene concentrations because of their work activities. No differences in ttMA and S-PMA, and no clinical changes were found between both groups, but linearity was observed between leukocyte count and ttMA; and 15% of workers had leukocyte counts less than 4.5 × 10⁹ cells L⁻¹, demanding close worker’s attention. No differences were observed between the two groups for THIOL, MDA, MCA, or nuclear abnormalities. A multiple linear relationship was obtained for the biomarkers MCA and C-FPG. A significant correlation was found between length of time in current job and the biomarkers C-FPG, MCA, GST, and MDA. Although both populations had chronic exposure to benzene, the filling station workers were exposed to higher concentrations of benzene during their work activities, indicating an increased risk of DNA damage. Full article

The cell cycle is a series of events by which cellular components are accurately segregated into daughter cells, principally controlled by the oscillating activities of cyclin-dependent kinases (CDKs) and their co-activators. In eukaryotes, DNA replication is confined to a discrete synthesis phase while chromosome segregation occurs during mitosis. During mitosis, the chromosomes are pulled into each of the two daughter cells by the coordination of spindle microtubules, kinetochores, centromeres, and chromatin. These four functional units tie chromosomes to the microtubules, send signals to the cells when the attachment is completed and the division can proceed, and withstand the force generated by pulling the chromosomes to either daughter cell. Protein ubiquitination is a post-translational modification that plays a central role in cellular homeostasis. E3 ubiquitin ligases mediate the transfer of ubiquitin to substrate proteins determining their fate. One of the largest subfamilies of E3 ubiquitin ligases is the family of the tripartite motif (TRIM) proteins, whose dysregulation is associated with a variety of cellular processes and directly involved in human diseases and cancer. In this review we summarize the current knowledge and emerging concepts about TRIMs and their contribution to the correct regulation of cell cycle, describing how TRIMs control the cell cycle transition phases and their involvement in the different functional units of the mitotic process, along with implications in cancer progression. Full article