Biology subjects

Yaguchi, K.

Publications and source records attributed to Yaguchi, K..

13 recordsLinked to original sources

bioRxiv2026

Spindle pole proteins confine chromosomes to ensure their expulsion during female meiosis

Animal oocytes undergo highly asymmetric divisions to expel excess copies of their genome into compact cells called polar bodies. This requires tight clustering and cortical positioning of meiotic chromosomes, yet the mechanism remains incompletely understood. Using C. elegans oocytes, we found that the meiotic spindle pole protein ZYG-9/ch-TOG delocalizes from microtubules to spread along the surface of chromosomes and prevent their dispersal in meiotic anaphase. This effect was more pronounced in the absence of the spindle, where ZYG-9 formed into a micron-scale droplet that enveloped all chromosomes. Purified ZYG-9 was sufficient to bind DNA and coat reconstituted chromatin. Mutations that perturb ZYG-9-DNA binding impaired chromosome packaging into polar bodies, resulting in oocytes carrying extra chromosomes and reduced fertility. We propose that liquid-like assemblies of spindle pole proteins are repurposed as surface-acting glue to tightly package meiotic chromosomes into polar bodies, thus ensuring oocytes have the correct genome copy number.

cell biology↗

bioRxiv2025

TRIM37 recognizes a bipartite degron to ubiquitinate centrosome substrates

Dysregulation of the E3 ubiquitin ligase TRIM37 is associated with tumor formation and Mulibrey nanism, a recessive developmental syndrome. TRIM37 regulates steady-state levels of centrosome proteins and limits their ectopic assembly, but how it recognizes and ubiquitinates its substrates is poorly understood. We found that TRIM37 directly ubiquitinates the centrosome-forming protein Cep192 at 7 lysines clustered near its C-terminus. TRIM37 binds Cep192 at a C-terminal intrinsically disordered region followed by an ASH domain (IDR+ASH8). Mutation of the 7 lysines or the IDR+ASH8 domain increased Cep192 levels and stability in cells, indicating loss of TRIM37-based regulation. Fusing IDR+ASH8 to an unrelated protein (GFP-EB1) was sufficient to enable its degradation via TRIM37. Biochemical assays revealed that IDR+ASH8 is primarily monomeric and binds TRIM37 via two separate coiled-coil motifs with mid-nanomolar affinity. We propose that the IDR+ASH8 motif is a bipartite degron for TRIM37, enabling it to target centrosome proteins and adjust their levels.

cell biology↗

bioRxiv2024

eIF4ET regulates meiotic proteome levels to enable oocyte formation and storage

Animals store oocytes in a dormant state for weeks to decades before ovulation. The homeostatic programs oocytes use to endure long-term storage are poorly understood. Using female nematodes as a short-lived model, we found that oocyte formation and storage required IFET-1, the conserved eIF4E-Transporter protein (eIF4ET). IFET-1 co-assembled with CAR-1 (Lsm14) to form micron-scale condensates in stored oocytes, which dissipated after oocyte activation. Depletion of IFET-1 destabilized the stored oocyte proteome, leading to lower translation, a decline in microtubule maintenance proteins, and errors in microtubule organization and meiotic spindle assembly. Deleting individual domains within IFET-1 impaired oocyte storage without affecting oocyte formation. Thus, in addition to establishing a healthy oocyte reserve in young mothers, IFET-1 ensures that correct levels of cytoskeletal proteins are maintained as oocytes age. Human eIF4ET also localized to micron-scale puncta in dormant oocytes in a reproductively healthy patient. Our results clarify the role for eIF4ET in maintaining the oocyte reserve and further support eIF4ET dysfunction as an upstream cause of embryonic aneuploidy and age-related infertility.

developmental biology↗

bioRxiv2024

Flexible Adjustment of Oxytocin Neuron Activity in Mother Mice Revealed by Microendoscopy

Oxytocin (OT) neurons in the hypothalamic paraventricular nucleus (PVH) play an important role in a range of physiological and behavioral processes, including the initiation of milk ejection and the regulation of parental behaviors in mothers. However, their activity patterns at the single-cell level remain poorly understood. Using microendoscopic Ca2+ imaging in freely moving mother mice, we demonstrate highly correlated pulsatile activity among individual OT neurons during lactation. The number of OT neurons engaged in the pulsatile activity, along with the characteristics of individual waveforms, was dynamically modulated by lactation and weaning experiences. Notably, only [~]10% of the imaged OT neurons exhibited a significantly elevated response during pup retrieval, a hallmark of maternal behaviors, with a magnitude 18 times smaller than that observed during lactation. Collectively, these findings demonstrate the utility of microendoscopic imaging for PVH OT neurons and highlight the flexible adjustments of their individual activity patterns in freely behaving mother mice.

neuroscience↗

bioRxiv2024

Vasopressin-to-Oxytocin Receptor Crosstalk in the Preoptic Area Underlying Parental Behaviors in Male Mice

The transition to parenthood brings significant changes in behavior toward offspring. For instance, in anticipation of their offspring, male mice shift from infanticidal to caregiving behaviors. While the release of oxytocin from the paraventricular hypothalamus (PVH) plays a critical role in paternal caregiving, it does not fully account for the entire behavioral shift. The specific downstream neurons and signaling mechanisms involved in this process remain obscure. Here, we demonstrate that PVH vasopressin neurons also essentially contribute to a paternal behavioral shift. This vasopressin signal is partially transmitted through oxytocin receptors (OTRs) expressed in the anterior commissure and medial nuclei of the preoptic area. These OTR-expressing neurons receive inputs from both PVH oxytocin and vasopressin neurons and are responsible for expressing paternal caregiving behaviors. Collectively, this non-canonical vasopressin-to-OTR crosstalk within specific limbic circuits acts as a pivotal regulator of paternal behavioral changes in mice. HighlightsO_LIPVH vasopressin neurons are required for and can trigger paternal caregiving behaviors. C_LIO_LIVasopressin-induced paternal behaviors are mediated in part by OTRs in the preoptic area (POA). C_LIO_LIPOA OTR neurons receive inputs from both PVH oxytocin and vasopressin neurons. C_LIO_LIPOA OTR neurons play a critical and facilitative role in promoting paternal caregiving behaviors. C_LI

neuroscience↗

bioRxiv2024

Fragility of ER homeostatic regulation underlies haploid instability in human somatic cells

Mammalian somatic cells are generally unstable in the haploid state, resulting in haploid-to-diploid conversion within a short time frame. However, cellular and molecular principles that limit the sustainability of somatic haploidy remain unknown. In this study, we found the haploidy-linked vulnerability to ER stress as a critical cause of haploid intolerance in human somatic cells. Pharmacological induction of ER stress selectively induced apoptosis in haploid cells, facilitating the replacement of haploids by co-existing diploidized cells in a caspase-dependent manner. Biochemical analyses revealed that unfolded protein response (UPR) was activated with similar dynamics between haploids and diploids upon ER stress induction. However, haploids were less efficient in solving proteotoxic status, resulting in a bias toward a proapoptotic mode of UPR signaling. Artificial replenishment of chaperone function or inhibition of a UPR signal transducer ATF6 substantially alleviated the haploidy-linked upregulation of proapoptotic signaling and improved haploid cell retention under ER stress. These data demonstrate that the ER stress-driven haploid instability stems from inefficient proteostatic control that alters the functionality of UPR to cause apoptosis selectively in haploids. Interestingly, haploids suffered a higher level of protein aggregation even in unperturbed conditions, and the long-term stability of the haploid state was significantly improved by alleviating their natural proteotoxicity. Based on these results, we propose that the haploidy-specific vulnerability to ER stress creates a fundamental cause of haploid intolerance in mammalian somatic cells. Our findings provide new insight into the principle that places a stringent restriction on the evolution of animal life cycles.

biochemistry↗

bioRxiv2023

Multivalent coiled-coil interactions enable full-scale centrosome assembly and strength

During mitotic spindle assembly, microtubules generate tensile stresses on pericentriolar material (PCM), the outermost layer of centrosomes. The molecular interactions that enable PCM to assemble rapidly and resist external forces are unknown. Here we use cross-linking mass spectrometry to identify interactions underlying supramolecular assembly of SPD-5, the main PCM scaffold protein in C. elegans. Crosslinks map primarily to alpha helices within the phospho-regulated region (PReM), a long C-terminal coiled-coil, and a series of four N-terminal coiled-coils. PLK-1 phosphorylation of SPD-5 creates new homotypic contacts, including two between PReM and the CM2-like domain, and eliminates numerous contacts in disordered linker regions, thus favoring coiled-coil-specific interactions. Mutations within these interacting regions cause PCM assembly defects that are partly rescued by eliminating microtubule-mediated forces. Thus, PCM assembly and strength are interdependent. In vitro, self-assembly of SPD-5 scales with coiled-coil content, although there is a defined hierarchy of association. We propose that multivalent interactions among coiled-coil regions of SPD-5 build the PCM scaffold and contribute sufficient strength to resist microtubule-mediated forces.

cell biology↗

bioRxiv2022

Dynamic Modulation of Pulsatile Activities of Oxytocin Neurons in Lactating Wild-type Mice

Breastfeeding, which is essential for the survival of mammalian infants, is critically mediated by pulsatile secretion of the pituitary hormone oxytocin from the central oxytocin neurons located in the paraventricular and supraoptic hypothalamic nuclei of mothers. Despite its importance, the molecular and neural circuit mechanisms of the milk ejection reflex remain poorly understood, in part because a mouse model to study lactation was only recently established. In our previous study, we successfully introduced fiber photometry-based chronic imaging of the pulsatile activities of oxytocin neurons during lactation. However, the necessity of Cre recombinase-based double knock-in mice substantially compromised the use of various Cre-dependent neuroscience toolkits. To overcome this obstacle, we developed a simple Cre-free method for monitoring oxytocin neurons by an adeno-associated virus vector driving GCaMP6s under a 2.6 kb mouse oxytocin mini-promoter. Using this method, we monitored calcium ion transients of oxytocin neurons in the paraventricular nucleus in wild-type C57BL/6N and ICR mothers without genetic crossing. By combining this method with video recordings of mothers and pups, we found that the pulsatile activities of oxytocin neurons require physical mother-pup contact for the milk ejection reflex. Notably, the frequencies of photometric signals were dynamically modulated by mother-pup reunions after isolation and during natural weaning stages. Collectively, the present study illuminates the temporal dynamics of pulsatile activities of oxytocin neurons in wild-type mice and provides a tool to characterize maternal oxytocin functions.

neuroscience↗

bioRxiv2022

Tetraploidy-linked sensitization to CENP-E inhibition in human cells

Tetraploidy caused by whole-genome duplication is a hallmark of cancer cells, and tetraploidy-selective cell growth suppression is a potential strategy for targeted cancer therapy. However, how tetraploid cells differ from normal diploids in their sensitivity to anti-proliferative treatments remains largely unknown. In this study, we found that tetraploid cells are significantly more susceptible to inhibitors of a mitotic kinesin CENP-E than diploids. CENP-E inhibitor preferentially diminished the tetraploid cell population in diploid-tetraploid co-culture at optimum conditions. Live imaging revealed that tetraploidy-linked increase in unsolvable polar chromosome misalignment caused substantially longer mitotic delay in tetraploids than in diploids upon moderate CENP-E inhibition. This time gap of mitotic arrest resulted in cohesion fatigue and subsequent cell death, specifically in tetraploids, leading to tetraploidy-selective cell growth suppression. In contrast, the microtubule-stabilizing compound paclitaxel caused tetraploidy-selective growth suppression through the aggravation of spindle multipolarization. We also found that CENP-E inhibitor had superior generality to paclitaxel in its tetraploidy selectivity across a broader spectrum of cell lines. Our results highlight the unique properties of CENP-E inhibitors in tetraploidy-selective suppression, giving us clues on the further development of tetraploidy-targeting interventions in cancer.

cell biology↗

bioRxiv2022

Centrosome Loss And Cell Proliferation Defects Underlie Developmental Failure In Haploid Zebrafish Larvae

Haploid larvae in non-mammalian vertebrates are lethal with characteristic organ growth retardation collectively called "haploid syndrome." In contrast to mammals whose haploid intolerance is attributed to imprinting misregulation, the cellular principle of haploidy-linked defects in non-mammalian vertebrates remains unknown. Here, we investigated cellular defects that disrupt the ontogeny of gynogenetic haploid zebrafish larvae. Unlike diploid control, haploid larvae manifested unscheduled cell death at the organogenesis stage, attributed to haploidy-linked p53 upregulation. Moreover, we found that haploid larvae specifically suffered the gradual aggravation of mitotic spindle monopolarization during 1-3 days post fertilization, causing spindle assembly checkpoint-mediated mitotic arrest throughout the entire body. High-resolution imaging revealed that this mitotic defect accompanied the haploidy-linked centrosome loss occurring concomitantly with the gradual decrease in larval cell size. Either resolution of mitotic arrest or depletion of p53 partially improved organ growth in haploid larvae. Based on these results, we propose that haploidy-linked mitotic defects and cell death are parts of critical cellular causes shared among vertebrates that limit the larval growth in the haploid state, contributing to an evolutionary constraint on allowable ploidy status in the vertebrate life cycle.

developmental biology↗

bioRxiv2020

Mevalonate pathway-mediated ER homeostasis is required for haploid stability in human somatic cells

The somatic haploidy is unstable in diplontic animals, but cellular processes determining haploid stability remain elusive. Here, we found that inhibition of mevalonate pathway by pitavastatin, a widely used cholesterol-lowering drug, drastically destabilized the haploid state in HAP1 cells. Interestingly, cholesterol supplementation did not restore haploid stability in pitavastatin-treated cells, and cholesterol inhibitor U18666A did not phenocopy haploid destabilization. These results ruled out the involvement of cholesterol in haploid stability. Besides cholesterol perturbation, pitavastatin induced endoplasmic reticulum (ER) stress, the suppression of which by a chemical chaperon significantly restored haploid stability in pitavastatin-treated cells. Our data demonstrate the involvement of the mevalonate pathway in the stability of the haploid state in human somatic cells through managing ER stress, highlighting a novel link between ploidy and ER homeostatic control.

cell biology↗

bioRxiv2020

Uncoupling of DNA replication and centrosome duplication cycles is a primary cause of haploid instability in mammalian somatic cells

Mammalian haploid somatic cells are unstable and prone to diploidize, but the cause of haploid instability remains largely unknown. Previously, we found that mammalian haploid somatic cells suffer chronic centrosome loss stemming from the uncoupling of DNA replication and centrosome duplication cycles. However, the lack of methodology to restore the coupling between DNA replication and centrosome duplication has precluded us from investigating the potential contribution of the haploidy-linked centrosome loss to haploid instability. In this study, we developed an experimental method that allows the re-coupling of DNA and centrosome cycles through the chronic extension of the G1/S phase without compromising cell proliferation using thymidine treatment/release cycles. Chronic extension of G1/S restored normal mitotic centrosome number and mitotic control, substantially improving the stability of the haploid state in HAP1 cells. Stabilization of the haploid state was compromised when cdk2 was inhibited during the extended G1/S, or when early G1 was chronically extended instead of G1/S, showing that the coupling of DNA and centrosome cycles rather than a general extension of the cell cycle is required for haploid stability. Our data indicate the chronic centriole loss arising from the uncoupling of centrosome and DNA cycles as a direct cause of genome instability in haploid somatic cells, and also demonstrate the feasibility of modulation of haploid stability through artificial coordination between DNA and centrosome cycles in mammalian somatic cells.

cell biology↗

bioRxiv2017

Uncoordinated centrosome duplication cycle underlies the instability of non-diploid states in mammalian somatic cells

In animals, somatic cells are usually diploid and are unstable when haploid for unknown reasons. In this study, by comparing isogenic human cell lines with different ploidies, we found frequent centrosome loss specifically in the haploid state, which profoundly contributed to haploid instability through monopolar spindle formation and subsequent mitotic defects. We also found that efficiency of centriole licensing and duplication, but not that of DNA replication, changes proportionally to ploidy level, causing gradual loss or frequent overduplication of centrioles in haploid and tetraploid cells, respectively. Centriole licensing efficiency seemed to be modulated by astral microtubules, whose development scaled with ploidy level, and artificial enhancement of aster formation in haploid cells restored centriole licensing efficiency to diploid levels. Haploid-specific centrosome loss was also observed in parthenogenetic mouse embryos. We propose that incompatibility between the centrosome duplication and DNA replication cycles arising from different scaling properties of these bioprocesses upon ploidy changes, underlies the instability of non-diploid somatic cells in mammals.\n\nSummaryYaguchi et al. show that a delay or acceleration of centriole licensing compromises the control of centrosome number in haploid or tetraploid human cells, respectively, suggesting a cellular basis of the instability of non-diploid somatic cells in mammals.

cell biology↗