Biology subjects

Chenevert, J.

Publications and source records attributed to Chenevert, J..

9 recordsLinked to original sources

bioRxiv2026

Spatiotemporal regulation of LGN/NuMA and astral microtubules generate mirror symmetric spindle rotations

The positioning of the mitotic spindle controls the size, content, and position of daughter cells within embryos and tissues. A major spindle positioning mechanism is cortical pulling whereby the membrane-bound complex LGN/NuMA/Dynein captures astral microtubules and pulls centrosomes toward the cell cortex. Cytoplasmic dynein and astral growth tend to counteract cortical pulling and position spindles at the cell center. It remains unclear how these opposing forces cooperate. Here we examine the ascidian embryo, where spindles of two germ line cells rotate toward one another causing divisions which are both mirror symmetric and unequal. We find that this spindle behavior is governed by transient enrichment of LGN and NuMA and enhanced cortical pulling at the shared cell contact. Inhibition of the LGN/NuMA complex disrupts spindle alignment, unequal cleavage, and mirror symmetry. Temporal analysis shows that cortical pulling force initates at anaphase when there is a sharp increase in astral microtubule length. These results point towards two phases of spindle positioning forces, with cytoplasmic pulling and cortical pulling operating sequentially during early and late mitosis.

cell biology↗

bioRxiv2026

The extraordinary robustness of mitotic spindle assembly to microtubule dynamics plasticity revealed by end-binding proteins and tubulin tagging

Microtubules are dynamic, conserved cytoskeletal filaments that are essential for all eukaryotic cells. Microtubule dynamic properties are primarily characterized by measuring how fast they grow and shrink (growth and shrinkage rates) and how often they switch between assembly and disassembly (catastrophe and rescue frequencies). These four parameters measured for individual filaments can inform on the behavior of the entire microtubule network at the cell level. By comparing microtubule dynamics in Caenorhabditis elegans one-cell embryos using different genetically-encoded fluorescent probes, we observed an unexpected high variability in these parameters. Microtubule dynamics parameters were consistently higher in C. elegans strains expressing a fluorescently labelled microtubule end-binding protein than in strains relying on tubulin labelling, with microtubule growth rates differing by nearly a factor of two between the two conditions. This discrepancy was not limited to C. elegans, as we observed a similar effect in embryos of the tunicate Phallusia mammillata. Despite this, spindle size and assembly timing were only mildly affected. However, embryos expressing labelled end-binding protein exhibited higher frequency of mitotic defects and perturbed embryonic development upon exposure to various stresses such as elevated temperature or a compromised spindle assembly checkpoint. Thus, our work reveals both the remarkable robustness of mitotic spindle assembly in response to extreme microtubule dynamics plasticity, and the requirement for strict control of microtubule dynamics across successive early embryonic divisions. Our findings should also serve as a cautionary note when using tagged end-binding proteins to measure microtubule dynamics.

cell biology↗

bioRxiv2025

Embryonic development of the Mediterranean starfish Hacelia attenuata

Starfish play essential ecological roles as predators and ecosystem regulators, however detailed developmental descriptions exist for only a handful of species, none of which come from the Mediterranean. In this study we provide the first full account of the development of the Mediterranean starfish Hacelia attenuata, from oocyte maturation through embryogenesis and larval formation, showing that its development largely follows the canonical pattern known from other asteroids. Oocytes resume meiosis when exposed to 1-methyladenine, exhibiting conserved features such as formation of a nuclear actin shell for chromosome gathering prior to meiotic spindle assembly. Embryogenesis then proceeds through equal radial cleavages to form a ciliated blastula, followed by gastrulation via invagination and mesenchymal cell delamination. The larvae develop through typical bipinnaria and brachiolaria stages, displaying characteristic feeding structures and attachment organs. Importantly, developmental rate increases substantially at higher temperatures, consistent with the species distribution in warm Mediterranean waters. Together, these findings position H. attenuata as a promising new model for studying thermal adaptation, environmental resilience and conserved developmental mechanisms in starfish.

developmental biology↗

bioRxiv2025

Tug-of-war between cortical and cytoplasmic forces shaping planar of 4-cell stage embryos

Early embryonic cleavages often follow conserved geometric rules, resulting in species-specific cleavage patterns. How these rules are mechanistically implemented, however, varies widely across species and remains poorly understood. Here, using quantitative 3D live imaging, mechanical and biochemical manipulations, we dissect the mechanisms governing centrosomal complex (CC) migration and spindle orientation in ascidian 2-cell stage embryos, which generate the characteristic planar, square 4-cell stage. We show that following the first mitotic division, the CCs in each blastomere form at variable orientations relative to the mother spindle and progressively achieve parallel and coplanar alignment during interphase of the 2-cell stage. Final CC orientation is established before nuclear envelope breakdown, contrasting with the dynamic spindle reorientation during mitosis reported in other organisms. Our analyses reveal that CC rotation is driven by forces acting on long astral microtubules and guided by an anisotropic, endoplasmic reticulum (ER)-rich domain that surrounds the CCs. This ER domain is associated with a dense astral microtubule network, enabling efficient length-dependent microtubule pulling that can orient the CCs even in the absence of cell shape cues. In parallel, dynein-mediated cortical pulling refines CCs tilt and maintains coplanarity. Together, these findings uncover a cell-cycle-regulated cooperation between ER-mediated cytoplasmic forces and anisotropic cortical forces that ensures robust planar cleavage in ascidian early embryos.

developmental biology↗

bioRxiv2023

Reduction of cortical pulling at mitotic entry facilitates aster centration

Although it has been studied for more than a century, the question of how one cell divides into two equal parts is still not fully resolved. Zygotes have provided much of the mechanistic insight into how the mitotic apparatus finds the center of the cell since the centrally-located mitotic apparatus is created from a large sperm aster that forms at the cortex and thus far from the zygote center. Here we show that in ascidians, the sperm aster extends throughout the cytoplasm during interphase yet remains located near the cortex and does not migrate towards the zygote center. It is only at mitotic entry, when the sperm aster has duplicated and the mitotic apparatus is being assembled, that most of the migration and centration occurs. This temporal pattern of centration behavior is mirrored by primate zygotes (including human). The current mechanisms of aster centration include cytoplasmic pulling that scale with microtubule (MT) length, MT pushing against the proximal cortex or MT-based cortical pulling. However, it is not yet known whether and how these 3 mechanisms are coordinated to prevent aster migration during interphase and trigger migration at mitotic entry. By monitoring quantitatively all three mechanisms (cytoplasmic pulling, pushing and cortical pulling) we have discovered that cortical pulling is switched off as the zygote enters mitosis while both cytoplasmic pulling and proximal cortical pushing remain active. Physical simulations could recapitulate both the static and migratory aspects of sperm aster and mitotic apparatus behavior. We therefore surmise that the reduction in cortical pulling at mitotic entry represents a switch that allows proximal cortical pushing forces and cytoplasmic pulling forces to center the nascent mitotic apparatus. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=156 SRC="FIGDIR/small/533625v1_ufig1.gif" ALT="Figure 1"> View larger version (35K): org.highwire.dtl.DTLVardef@135189dorg.highwire.dtl.DTLVardef@e2ef1dorg.highwire.dtl.DTLVardef@c070f7org.highwire.dtl.DTLVardef@38832f_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LISperm aster/mitotic apparatus centration occurs at entry into first mitosis C_LIO_LIMT-based cortical pulling is active during interphase and switched off at mitotic entry C_LIO_LILoss of cortical pulling at mitosis entry facilitates centration of the aster C_LIO_LIMT-based cytoplasmic pulling is active during both interphase and mitosis C_LIO_LIAgent-based simulations advocate the need for cytoplasmic pulling, a switch in cortical pulling and a minor role of pushing for aster centration at mitotic entry. C_LI

cell biology↗

bioRxiv2022

Modulation of Spindle Assembly Checkpoint efficiency by cell fate in a chordate embryo

The spindle assembly checkpoint (SAC) is a surveillance system which preserves genome integrity by delaying anaphase onset until all chromosomes are correctly attached to spindle microtubules. Recruitment of SAC proteins to unattached kinetochores generates an inhibitory signal that prolongs mitotic duration. Chordate embryos are atypical in that spindle defects do not delay mitotic progression during early development, implying that either the SAC is inactive or the cell-cycle target machinery unresponsive. Here we show that in embryos of the chordate Phallusia mammillata the SAC delays mitotic progression from the 8th cleavage divisions. Unattached kinetochores are not recognized by the SAC machinery until the 7th cell cycle when the SAC is acquired. Following acquisition, SAC strength, manifest as the degree of mitotic lengthening induced by spindle perturbations, is specific to different cell types and is modulated by cell size showing similarity to SAC control in early Caenorhabditis elegans embryos. We conclude that SAC acquisition is a process likely specific to chordate embryos, while modulation of SAC efficiency in SAC proficient stages depends on cell fate and cell size similarly to non-chordate embryos.

cell biology↗

bioRxiv2019

The spindle assembly checkpoint functions during early development in non-chordate embryos.

In eukaryotic cells, a spindle assembly checkpoint (SAC) ensures accurate chromosome segregation. This control mechanism monitors proper attachment of chromosomes to spindle microtubules and delays mitotic progression if connections are erroneous or absent. The SAC operates in all eukaryotic cells tested so far, but is thought to be relaxed during early embryonic development in animals. Here, we evaluate the checkpoint response to lack of kinetochore-spindle microtubule interactions in early embryos of diverse animal species from the main metazoan groups. Our analysis shows that there are two classes of embryos, either proficient or deficient for SAC activation during cleavage. Sea urchins, mussels and jellyfish embryos show a prolonged mitotic block in the absence of spindle microtubules from the first cleavage division, while ascidian and amphioxus embryos, like those of Xenopus and zebrafish, continue mitotic cycling without delay. SAC competence during early development shows no correlation with cell size, chromosome number or kinetochore to cell volume ratio, ruling out the hypothesis that lack of checkpoint activity in early embryos is due to the large egg volume. Our results instead indicate that there is no inherent incompatibility between SAC activity and large fast-dividing embryonic cells. We suggest that SAC proficiency is the default situation of metazoan embryos, and that SAC activity is specifically silenced in chordate species with fast dividing embryos.

cell biology↗

bioRxiv2017

Role of midbody remnant in meiosis II creating tethered polar bodies

Polar body (PB) formation is an extreme form of unequal cell division that occurs in oocytes due to the eccentric position of the small meiotic spindle near the oocyte cortex. Prior to PB formation, a chromatin-centered process causes the cortex overlying the meiotic chromosomes to become polarized. This polarized cortical subdomain marks the site where a cortical protrusion or outpocket forms at the oocyte surface creating the future PBs. We observed that PB1 becomes tethered to the egg via PB2, indicating that the site of PB1 cytokinesis directed the precise site for PB2 emission. We therefore studied whether the midbody remnant left behind following PB1 emission was involved, together with the egg chromatin, in defining the precise cortical site for PB2 emission. During outpocketing of PB2 in ascidians, we discovered that a small corps around 1m in diameter protruded from the center of the cortical outpocket that will form the future PB2, which we call the \"polar corps\". During emission of PB2, this small polar corps became localized between PB2 and PB1 and appeared to link PB2 to PB1. We tested the hypothesis that this small polar corps on the surface of the forming PB2 was the midbody remnant from the previous round of PB1 cytokinesis. We had previously discovered that Plk1::Ven labeled midbody remnants in ascidian embryos. We therefore used Plk1::Ven to follow the dynamics of the PB1 midbody remnant during meiosis II. Plk1::Ven strongly labeled the small polar corps that formed on the surface of the cortical outpocket that created PB2. Following emission of PB2, this polar corps was rich in Plk1::Ven and linked PB2 to PB1. By labelling actin (with LifeAct::mCherry/GFP or TRITC-Phalloidin) we also demonstrated that actin accumulates at the midbody remnant and also forms a cortical cap around the midbody remnant in meiosis II that prefigured the precise site of cortical outpocketing during PB2 emission. Phalloidin staining of actin and immunolabelling of anti-phospho aPKC during meiosis II in eggs that had PB1 removed showed that the midbody remnant remained within the egg following emission of PB1. Dynamic imaging of microtubules labelled with Ens::3GFP, MAP7::GFP or EB3::3GFP showed that one pole of the second meiotic spindle was located near the midbody remnant while the other pole rotated away from the cortex during outpocketing. Finally, we report that failure of the second meiotic spindle to rotate can lead to the formation of two cortical outpockets at anaphase II, one above each set of chromatids. It is not known whether the midbody remnant of PB1 is involved in directing the precise location of PB2 in other species as in ascidians. However, a review of the literature indicates that PB1 is tethered to the egg surface via PB2 in a number of species including members of the cnidarians, lophotrochozoa and echinoids, suggesting that the midbody remnant formed during PB1 emission may be involved in directing the precise site of PB2 emission throughout the invertebrates.

developmental biology↗

bioRxiv2017

Kif2 microtubule depolymerase is required for unequal cell division and localizes to a subdomain of cortical endoplasmic reticulum

Unequal cell division (UCD) is a fundamental process responsible for creating sibling cell size asymmetry; however, how microtubules are specifically depolymerized on one aster of the mitotic spindle creating a smaller sibling cell during UCD has remained elusive. Using invertebrate chordate embryos (ascidian) that possess a large cortical structure (CAB) that causes UCD, we identified a microtubule depolymerase (Kif2) involved in creating cell size asymmetry. Kif2 localizes to the cortical subdomain of endoplasmic reticulum in the CAB. During three successive UCDs, Kif2 protein accumulates at the CAB during interphase and is delocalized from the CAB in mid mitosis. Rapid imaging of microtubule dynamics at the cortex revealed that microtubules do not penetrate the CAB during interphase. Inhibition of Kif2 function prevents the development of mitotic aster asymmetry and centrosome movement towards the CAB thereby blocking UCD, whereas locally increasing microtubule depolymerization causes exaggerated asymmetric spindle positioning. This study provides insights into the fundamental process of UCD and for the first time shows that a microtubule depolymerase is localized to a cortical site controlling UCD.

cell biology↗