Mechanical regulation of cell division orientation: investigating the role of nuclear mitotic apparatus protein

Abstract

Cells within an organism experience a variety of mechanical forces from their surrounding tissue environment. Reading and responding to these forces is crucial to shape and maintain tissues, with errors in this process contributing to failures in embryogenesis and diseases such as cancer. Forces are known to regulate cell division orientation, which is determined by the position of the mitotic spindle. For example, stretching a tissue causes divisions to align with the stretch axis. In order for external forces to regulate division orientation, mechanical cues must be relayed to the mitotic spindle. Spindle-associated cortical proteins are likely key to this process as they may sense forces acting on the cell cortex and transduce it into a spindle orientation response. One such candidate, nuclear mitotic apparatus protein (NuMA) has been implicated in orienting the spindle according to force although the mechanistic details remain uncharacterised, especially in a tissue context. Therefore, we utilised the Xenopus laevis animal cap tissue, to which tensile forces can be applied externally, to understand the role of NuMA in mechanosensitive spindle orientation. Using GFP-tagged NuMA, we show that cortical localisation of GFP-NuMA is dynamic and sensitive to uniaxial stretch, with recruitment to the polar cortices earlier during mitosis in stretched tissues. Using morpholino-targeted knockdown of endogenous Xenopus NuMA in early embryos, we also show that a reduction in NuMA levels disrupts the ability of cells to orient divisions along the axis of tissue stretch and cell shape. Interestingly, our data suggest that mechanosensitive spindle orientation through NuMA is an effect of direct force sensing rather than sensing changes in cell shape. Furthermore, by comparing two different tissue stretch regimes, we demonstrate that NuMA responds specifically to anisotropic tension to orient cell divisions in stretched Xenopus tissues. Overall, our results indicate that NuMA is vital to orient a mitotic spindle according to external forces.

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