Pak1 control of E-cadherin endocytosis regulates salivary gland lumen size and shape

Abstract

SUMMARYGenerating and maintaining proper lumen size and shape in tubular organs is essential for organ function. Here, we demonstrate a novel role for p21-activated kinase 1 (Pak1) in defining the size and shape of the Drosophila embryonic salivary gland lumen by regulating the size and elongation of the apical domain of individual cells. Pak1 mediates these effects by decreasing and increasing E-cadherin levels at the adherens junctions and basolateral membrane, respectively, through Rab5-and Dynamindependent endocytosis. We also demonstrate that Cdc42 and Merlin act together with Pak1 to control lumen size. A role for Pak1 in E-cadherin endocytosis is supported by our studies of constitutively active Pak1, which induces the formation of multiple intercellular lumens in the salivary gland in a manner dependent on Rab5, Dynamin and Merlin. These studies demonstrate a novel and crucial role for Pak1 and E-cadherin endocytosis in determining lumen size and shape, and also identify a mechanism for multiple lumen formation, a poorly understood process that occurs in normal embryonic development and pathological conditions.

Rho GTPase controls Drosophila salivary gland lumen size through regulation of the actin cytoskeleton and Moesin

“…In wild-type glands, the lumen diameter in the proximal region of the gland gradually decreased between embryonic stages 11 and 12 as the gland turned and migrated posteriorly, whereas lumen diameter in the medial and distal regions did not change (Pirraglia et al, 2010) (supplementary material Fig. S1).…”

“…Apical domain elongation ratio of an individual gland cell was measured according to E-cad immunofluorescence staining of stage 12 embryos. Elongation ratio represents the ratio of a single measurement of the longest length of the apical domain oriented along the proximal-distal axis to a single measurement of the longest length of the apical domain along the dorsalventral axis (Pirraglia et al, 2010). Measurements of apical domain elongation ratio were performed with the eight most proximal cells in each gland.…”

“…In three-dimensional Caco-2 cell cultures, Cdc42 prevents multiple lumen formation by orienting cell divisions and directing apical membrane growth (Jaffe et al, 2008). We recently showed that in the Drosophila salivary gland, Cdc42 and the p21 activated kinase (Pak) 1 regulate gland lumen size (Pirraglia et al, 2010). In contrast to Cdc42 and Rac GTPases, the role of Rho in tube and lumen morphogenesis is poorly understood.…”

“…After salivary gland cells invaginate, they undergo a phase of robust apical surface membrane growth (Myat and Andrew, 2002) and the apical domain size of individual cells decreases and elongates to become anisotropic along the longest axis of the lumen (Pirraglia et al, 2010). Apical membrane growth is limited by the basic helix-loop-helix (bHLH) transcriptional repressor Hairy and its regulation of Huckebein (Hkb), an Sp1/Egr-like transcription factor, and by target genes klarsicht (klar), which encodes a KASH-domain-containing regulator of organelle and nuclear transport (Fischer-Vize and Mosley, 1994;Fischer et al, 2004;Guo et al, 2005;Mosley-Bishop et al, 1999) and crumbs (crb), which encodes an apical membrane protein that is necessary for the establishment and maintenance of apical-basal polarity (Myat and Andrew, 2002;Tepass and Knust, 1993;Tepass et al, 1990).…”

“…Based on mathematical modeling, it is thought that salivary gland lumens of rib mutant embryos fail to elongate because of increased apical surface stiffness and viscosity (Cheshire et al, 2008). We recently showed that apical domain elongation is regulated by Pak1 through differential localization of E-cadherin (E-cad; Shotgun -FlyBase) at the adherens junctions and at the basolateral membrane (Pirraglia et al, 2010). Here, we provide the first evidence that Rho1 controls lumen size in the Drosophila embryonic salivary gland through regulation of the actin cytoskeleton and Moesin.…”