PgmNr W4189: A vesicle-intrinsically regulated pathway for apical polarity.

Authors:
Nan Zhang 1,2 ; Hongjie Zhang 1,3 ; Liakot Khan 1 ; Gholamali Jafari 1 ; Yong Eun 1 ; Edward Membreno 1 ; Razan El-Daouk 1 ; Verena Gobel 1


Institutes
1) Mucosal Immunology and Biology Research Center, MGH/Harvard Medical School, Boston, MA, USA; 2) College of Life Sciences, Jilin University, Changchun, Jilin, China; 3) Faculty of Health Sciences/University of Macau, Taipa, Macau, China.


Keyword: Transportation of intracellular components

Abstract:

Polarized epithelial membrane domains are thought to be established by membrane- and junction-associated polarity cues (e.g. partitioning-defective PARs), with directional trafficking subsequently maintaining them.  Our prior work showed that glycosphingolipids (GSLs), clathrin and its AP-1 adaptor function as vesicle-associated apical polarity cues in C. elegans tubular epithelia.  Their loss causes intestinal polarity inversion with a full basolateral-to-apical membrane domain conversion and ectopic lateral lumen formation.  These and other findings suggested that trafficking itself may define polarized domain identities, in part by targeting polarity determinants (e.g. PARs), but possibly by targeting multiple membrane components.

Here, the GSL-loss-of-function intestinal polarity inversion phenotype was used to identify components and map trajectories of such a putative vesicle-intrinsically regulated apical polarity pathway.  We mined three of our unbiased and targeted multi- and unicellular tubulogenesis screens that had examined the asymmetrical placement and function of the apical/lumenal actin linker ERM-1 for genes with predicted roles in trafficking.  49 trafficking genes with a broad range of vesicle-associated functions were identified as required for apical membrane biogenesis, only few of which previously implicated in directional trafficking.  We examined the ability of these genes to modify polarity inversion in the GSL-biosynthetic 3-ketoacyl-CoA reductase deficient mutant let-767.

These genetic interaction screens identified eleven enhancers implicating secretory/biosynthetic trafficking routes in this pathway, and five suppressors implicating endocytic/recycling routes.  In an attempt to map polarized trajectories, three suppressors, DAB-1/Disabled, RAB-7, and VHA-6 (a V-ATPase component), were selected for further analysis.  Imaging and loss-of-function studies of expanding intestines revealed unexpected functions in apical and basolateral membrane biogenesis for these endo- and plasma-membrane-associated molecules and placed them on recycling routes.  They thereby position suppressors upstream of GSL sorting sites, while in turn suggesting that GSLs may sort beyond the Golgi on biosynthetic trajectories that receive input from recycling routes during plasma membrane biogenesis.

This study identifies multiple vesicle components for polarized trafficking, including novel vesicle-carrier-associated polarity cues, and proposes trajectories for a vesicle-intrinsically regulated trafficking pathway for apical polarity.