Rac1 modulation of the apical domain is negatively regulated by beta(Heavy)-spectrin

Epithelial polarity and morphogenesis require the careful coordination of signaling and cytoskeletal elements. In this paper, we describe multiple genetic interactions between the apical cytoskeletal protein beta(H) and Rac1 signaling in Drosophila: activation of Rac1 signaling by expression of the exchange factor Trio, is strongly enhanced by reducing bH levels, and such reductions in beta(H) levels alone are shown to cause an increase in GTP-Rac1 levels. In contrast, co-expression of a C-terminal fragment of beta(H) (beta H33) suppresses the Trio expression phenotype. In addition, sustained expression of beta H33 alone in the eye induces a strong dominant phenotype that is similar to the expression of dominant negative Rac1(N17), and this phenotype is also suppressed by the co-expression of Trio or by knockdown of Rac-GAP50C. We further demonstrate that a loss-of-function allele in pak, a Rac1 effector and negative regulator of beta(H)' dominantly suppresses larval lethality arising loss-of-function karst (beta(H)) alleles. Furthermore, expression of constitutively active Pak(myr) in the larval salivary gland induces expansion of the apical membrane and destabilization of the apical polarity determinants Crumbs and aPKC. These effects resemble a Rac1 activation phenotype and are suppressed by beta H33. Together, our data suggest that apical proteins including beta(H) are negatively regulated by Rac1 activation, but that Rac1 signaling is also suppressed by beta(H) through its C-terminal domain. Such a system would be bistable with either Rac1 or beta(H) predominant. We suggest a model for apical domain maintenance wherein Rac1 down-regulation of beta(H) (via Pak) is opposed by beta(H)-mediated down-regulation of Rac1 signaling. (C) 2010 Elsevier Ireland Ltd. All rights reserved.