Mitotic spindle dynamics in stretched epithelial tissue in vivo and in silico

Abstract

Cell division is vital for the growth and homeostasis of tissues. The outcome of division, for example a contribution to tissue spreading or tissue stratification, depends upon its spatial orientation within the tissue. In turn, the external environment of the tissue feeds back to determine cell division orientation, with divisions commonly aligning with an axis of greatest tensile force. Division orientation is determined by the orientation of the mitotic spindle. From its assembly until chromosome segregation, the spindle dynamically rotates and explores the cell by the interaction of its astral microtubules with protein complexes at the cell periphery. The nuclear mitotic apparatus protein (NuMA) is one such element implicated in spindle positioning which is localised dynamically to the cell cortex during cell division and to the spindle poles during cell division. As such, NuMA has been highlighted as a key candidate in driving the orientation of division with external force. Recent unpublished work in the Woolner lab has revealed that NuMA localisation to the cortex is sensitive to tissue tension and is perturbed in cells experiencing an externally applied force, though the precise mechanism by which NuMA functions to orient divisions with external force remains unclear. To determine how mechanosensitive spindle orientation is regulated, we used a combination of biological and mathematical approaches to analyse dynamic movements of the mitotic spindle. We utilised the tightly adhered epithelial layer of the Xenopus laevis animal cap to study spindle movements in stretched and unstretched tissues to assess the impact of stretch on the spindle rotational and translational dynamics. We find that the mitotic spindles undergo oscillatory movements as they seek out their division axis. The period of these oscillations is insensitive to external forces but we see an affinity for oscillating which is higher in unstretched tissues rather than stretched tissues. Crucially, the period of oscillation is sensitive to the depletion of NuMA. We develop a model of spindle pole displacements due to cortical pulling forces and microtubule-based restoring forces, using stochastic simulations, Fokker-planck equations, ODEs and an algebraic formulation. By systematically reducing the mathematical system we highlight the key relationships between parameters which promote dynamic movements of the spindle pole. We also show that oscillations in the position of a single spindle pole may occur in a select region of parameter space. Our results suggest that depletion of NuMA reduces the restoring force acting on the spindle which allows the spindle to oscillate with a longer period at lower numbers of cortical pulling elements. We highlight new avenues to explore to determine the role of NuMA in mechanosensitive orientation, through the use of interdisciplinary techniques that allow us to vary properties of cortical force generators in silico.

Date of Award	1 Aug 2023
Original language	English
Awarding Institution	The University of Manchester
Supervisor	Oliver Jensen (Supervisor) & Sarah Woolner (Supervisor)