Apical targeting of Diaphanous mediates polarized secretion in tubular organs. Eyal D. Schejter, Tal Rousso, Rada Massarwa, Erez Geron, Ben-Zion Shilo. Dept Molecular Genetics, Weizmann Institute of Science, Rehovot, Israel.

   The epithelial cells comprising tubular organs are highly polarized, a feature that enables efficient execution of specialized functions, such as vectorial secretion and nutrient absorption. We have previously shown that secretion from these epithelial cells into the organ lumen utilizes Myosin V-mediated transport of secretory vesicles over an apical layer of microfilaments, generated by the formin Diaphanous (Dia). This secretory mechanism, which is common to all Drosophila embryonic tubular organs examined, appears to be evolutionarily-conserved, as we have recently demonstrated that it underlies apical secretion in the mouse exocrine pancreas and salivary glands. Apical restriction of Dia activity is the central feature ensuring formation of the critical microfilament array that promotes apical secretion in Drosophila tubular organs. In the current study we have examined the mechanistic basis for this polarization of Dia activity. We find that Dia apical targeting requires the coincident detection of both PIP2 and the GTPase Rho1 on the apical surface of the tubular organ cells. We show that PIP2 levels regulate Dia localization. An N-terminal basic domain of Dia is critical for this regulation, indicative of direct interaction between Dia and membranal PIP2. Our data indicate that apical enrichment of PIP2 is a common feature of tubular organs in the fly embryo and larva, attributed in part to apical restriction of the PIP5 kinase (PIP5K) Skittles. We further show that Rho1 facilitates Dia apical targeting, both by inducing the open, active conformation of Dia, thereby exposing N-terminal domains that are critical for localization, as well as by physically anchoring Dia to the apical surface. This anchoring appears to be essential for the Dia-PIP2 interaction, leading to synergistic relations between the two apical cues. The mechanism we describe relies on the utilization of inherent and global features of tubular organs, in order to achieve a distinct localization pattern of Dia, thereby enabling proper execution of oriented secretion in these organs.