The microtubule end-binding protein EB2 is a central regulator of microtubule reorganisation in apico-basal epithelial differentiation

Abstract

Microtubule end-binding (EB) proteins influence microtubule dynamic instability, a process essential for microtubule reorganisation during apico-basal epithelial differentiation. Here we establish for the first time that EB2, but not EB1, expression is critical for initial microtubule reorganisation during apico-basal epithelial differentiation, and that EB2 downregulation promotes bundle formation. EB2 siRNA knockdown during early stages of apico-basal differentiation prevented microtubule reorganisation, while its downregulation at later stages promoted microtubule stability and bundle formation. Interestingly, while EB1 is not essential for microtubule reorganisation its knockdown prevented apico-basal bundle formation and epithelial elongation. EB2 siRNA depletion in undifferentiated epithelial cells induced formation of straight, less dynamic microtubules with EB1 and ACF7 lattice association and co-alignment with actin filaments, a phenotype that could be rescued by formin inhibition. Importantly, in situ inner ear and intestinal crypt epithelial tissue revealed direct correlations between low level of EB2 expression and presence of apico-basal microtubule bundles, which were absent where EB2 was elevated. EB2 is evidently important for initial microtubule reorganisation during epithelial polarisation, while its downregulation facilitates EB1/ACF7 microtubule lattice association, microtubule-actin filament co-alignment and bundle formation. The spatiotemporal expression of EB2 thus dramatically influences microtubule organisation, EB1/ACF7 deployment and epithelial differentiation.