Biology subjects

Pereira, A. J.

Publications and source records attributed to Pereira, A. J..

6 recordsLinked to original sources

bioRxiv2026

Microtubule occupancy at kinetochores links checkpoint silencing with mitotic memory

The spindle assembly checkpoint (SAC) promotes faithful chromosome segregation by delaying mitosis until all kinetochores attach to spindle microtubules. Paradoxically, a p53-dependent memory mechanism--the mitotic stopwatch" --blocks daughter cell proliferation after unusually prolonged mitoses. How the SAC coordinates with the mitotic stopwatch remains unknown. Here, we found that microtubule occupancy at kinetochores is a cornerstone linking SAC silencing with mitotic memory. By combining live-cell with super-resolution microscopy, FRAP, laser microsurgery, biochemistry and molecular perturbations in Indian muntjac fibroblasts, we show that SAC silencing at kinetochores is gradual, non-uniform and confined to highly-localized microtubule attachments. Augmin promotes timely SAC silencing with high microtubule occupancy at kinetochores, whereas MPS1/CDK1 inhibition bypasses this requirement. Conversely, low microtubule occupancy delays SAC silencing, increases segregation errors and blocks daughter cell proliferation due to mitotic stopwatch surveillance. Thus, timely SAC silencing with high microtubule occupancy avoids "bad memories" of mitosis to allow daughter cell proliferation.

cell biology↗

bioRxiv2024

Circling back: Vespula spp. re-invasion after knockout from volcanic ash fall in Patagonia

O_LIEffective management of invasive species requires understanding re-invasion dynamics, to prevent control failures, refine methodologies, and enhance protocols. Re-invasion is the process by which eradicated or naturally disappeared exotic species establish themselves again, in previously occupied areas. Re-invasion often bypasses initial invasion phases such as transport and introduction. During re-establishment, changes in the invader, the invaded environment, and species interactions can influence the success of invasion. C_LIO_LIWe investigated re-invasion patterns of Vespula germanica and Vespula vulgaris wasps in NW Patagonia (Argentina) following a significant natural disturbance: the 2011 Puyehue-Cordon Caulle volcanic eruption, which caused unprecedented arthropod defaunation. By establishing 23 sampling sites in heavily affected regions, we conducted sampling events over three years (2012-2014). C_LIO_LIOur findings show that re-invasion of both Vespula spp. occurs fast, with minimal interspecific competition among them observed. The proximity to urban centers, acting as refuges, further facilitated their establishment. C_LIO_LIThis unique case study highlights the adaptability of these invasive wasps in the face of extreme environmental disruptions. By describing the importance of considering reinvasion routes and refuges, this research may help develop more effective management strategies to control Vespula populations. C_LI

ecology↗

bioRxiv2024

Bridging the resolution-sectioning gap in stimulated emission depletion microscopy: theory and experiment

Microscopes generally exhibit superior performance in 2D compared to 3D. Fluorescence super-resolution imaging often intensifies this discrepancy, such as with the gold-standard vortex-based stimulated emission depletion (STED) microscope, which narrows the point-spread function only laterally. In this study, we developed a semi-analytical theory based on the Nijboer-Zernike expansion, conducted simulations and performed experiments to establish the merits of the alternative bivortex-based STED. We find that this mode reduces the axial-lateral resolution mismatch and effectively emulates noisier multi-beam approaches by providing access to point-spread function geometries that are strictly forbidden to the two conventional single-beam modes, 2D-STED and z-STED. Notably, theory and experiment indicate that, besides filling the gap, bivortex STED delivers a higher signal-to-background than the two modes it bridges.

biophysics↗

bioRxiv2021

A mitotic screen in Indian muntjac cells unveils how Augmin drives kinetochore fiber maturation

Chromosome segregation in mammals relies on the maturation of a thick bundle of kinetochore-attached microtubules known as k-fibers. How k-fibers mature from initial kinetochore-microtubule attachments remains a fundamental question. By combining functional perturbations in Indian muntjac, a placental mammal with the lowest known chromosome number (n=3) and distinctively large kinetochores, with fixed- and subsecond live-cell super-resolution CH-STED nanoscopy, we uncovered the mechanism by which Augmin mediates k-fiber maturation. Augmin promoted kinetochore microtubule turnover by sustaining microtubule formation from kinetochores and poleward flux, regardless of pioneer centrosomal microtubules. Tracking of microtubule growth events within individual k-fibers revealed a wide angular dispersion, consistent with Augmin-mediated branched microtubule nucleation. Augmin depletion reduced the frequency of microtubule growth events within k-fibers and hampered normal repair after acute k-fiber injury by laser microsurgery. Our work underscores the contribution of microtubule formation from kinetochores and Augmin-mediated microtubule amplification for k-fiber maturation and spindle assembly in mammals.

cell biology↗

bioRxiv2019

Microtubule tyrosination/detyrosination specifies a mitotic error code

Incorrect kinetochore-microtubule attachments during mitosis can lead to chromosomal instability, a hallmark of human cancers. Mitotic error correction relies on the kinesin-13 MCAK, a microtubule depolymerase whose activity in vitro is suppressed by -tubulin detyrosination - a post-translational modification enriched on long-lived microtubules. However, whether and how MCAK activity required for mitotic error correction is regulated by microtubule tyrosination/detyrosination remains unknown. Here we found that microtubule detyrosination accumulates on correct, more stable, kinetochore-microtubule attachments, whereas constitutively high microtubule detyrosination near kinetochores compromised efficient error correction. Rescue experiments suggest that mitotic errors due to excessive microtubule detyrosination result from suppression of MCAK activity, without globally affecting kinetochore microtubule half-life. Importantly, MCAK centromeric activity was required and sufficient to rescue mitotic errors due to excessive microtubule detyrosination. Thus, microtubules are not just passive elements during mitotic error correction, and their tyrosination/detyrosination works as a mitotic error code that allows centromeric MCAK to discriminate correct and incorrect kinetochore-microtubule attachments, thereby promoting mitotic fidelity.

cell biology↗

bioRxiv2018

Coherent-hybrid STED: a tunable photo-physical pinhole for super-resolution imaging at high contrast

Resolution in microscopy is not limited by diffraction as long as a nonlinear sample response is exploited. In a paradigmatic example, stimulated-emission depletion (STED) fluorescence microscopy fundamentally breaks the diffraction limit by using a structured optical pattern to saturate depletion on a previously excited sample area. Two-dimensional (2D) STED, the canonical low-noise STED mode, structures the STED beam by using a vortex phase mask, achieving a significant lateral resolution improvement over confocal fluorescence microscopy. However, axial resolution and optical sectioning remain bound to diffraction. Here we use a tunable coherent-hybrid (CH) beam to improve optical sectioning, markedly reducing background fluorescence. CH-STED, which inherits the 2D-STED immunity to spherical aberration, diversifies the depletion strategy, allowing an optimal balance between two key metrics (lateral resolution and background suppression) to be found. CH-STED is used to perform high-contrast imaging of complex biological structures, such as the mitotic spindle and the neuron cell body.

biophysics↗