Rho-GTPase stabilizes microtubules that are oriented towards the leading edge in serum-starved 3T3 fibroblasts through an unknown mechanism. We used a Rho-effector domain screen to identify mDia as a downstream Rho effector involved in microtubule stabilization. Constitutively active mDia or activation of endogenous mDia with the mDia-autoinhibitory domain stimulated the formation of stable microtubules that were capped and oriented towards the wound edge. mDia co-localized with stable microtubules when overexpressed and associated with microtubules in vitro. Rho kinase was not necessary for the formation of stable microtubules. Our results show that mDia is sufficient to generate and orient stable microtubules, and indicate that Dia-related formins are part of a conserved pathway that regulates the dynamics of microtubule ends.

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The asymmetric distribution of stable, posttranslationally modified microtubules (MTs) contributes to the polarization of many cell types, yet the factors controlling the formation of these MTs are not known. We have found that lysophosphatidic acid (LPA) is a major serum factor responsible for rapidly generating stable, detyrosinated (Glu) MTs in serum-starved 3T3 cells. Using C3 toxin and val14 rho we showed that rho was both necessary and sufficient for the induction of Glu MTs by LPA and serum. Unlike previously described factors that induce MT stability, rho induced the stabilization of only a subset of the MTs and, in wound-edge cells, these stable MTs were appropriately oriented toward the leading edge of the cell. LPA had little effect on individual parameters of MT dynamics, but did induce long states of pause in a subset of MTs near the edge of the cell. Rho stimulation of MT stability was independent of actin stress fiber formation. These results identify rho as a novel regulator of the MT cytoskeleton that selectively stabilizes MTs during cell polarization by acting as a switch between dynamic and stable states of MTs rather than as a modulator of MT assembly and disassembly.

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