The biosynthetic-secretory pathway, supplemented by recycling routes, specifies epithelial membrane polarity

Abstract

In prevailing epithelial polarity models, membrane-based polarity cues (e.g., the partitioning-defective PARs) position apicobasal cellular membrane domains. Intracellular vesicular trafficking expands these domains by sorting apicobasal cargo towards them. How the polarity cues are polarized and how sorting confers long-range vesicle directionality is still unclear. Here, a systems-based approach using two-tiered C. elegans genomics-genetics screens identifies trafficking molecules that are not implicated in apical sorting yet polarize apical membrane and PAR complex components. Live tracking of polarized membrane biogenesis suggests that the biosynthetic-secretory pathway, linked to recycling routes, is asymmetrically oriented towards the apical domain during its biosynthesis, upstream of PARs and independent of polarized target domains. This mode of membrane polarization could offer solutions to questions of current models of polarity and polarized trafficking.

A transcytotic actin shift polarizes vesicle trajectories and partitions apicobasal epithelial membrane domains

“…We conclude that bcAMs and actin are required to specify the position of apical PARs on not yet polarized membranes of still dividing and migrating intestinal cells and on already polarized but still expanding membranes of postmitotic cells within the fixed epithelium. These findings further support the proposed noncanonical function of bcAMs and actin in membrane polarity via directional trafficking that likewise operates upstream of PARs in intestinal polarity 7 . Our findings also reveal that branched chain actin dynamics functions upstream of the to date earliest polarity cue in the establishment of epithelial polarity in the C. elegans intestine.…”

“…To start mapping vesicle trajectories through C. elegans intestinal development, we directed expression of the ubiquitous RAB-11 (marking endosomes with presumed apical directionality) and RAB-10 (marking endosomes with presumed basolateral directionality to the intestine by elt-2 to visually resolve their subcellular positions in both pre- and post-intercalation intestinal cells ( 27 ; Methods). We found that RAB-11+ post-Golgi vesicles, previously identified as apical polarity cues 7 , indeed exhibit a bcAM/actin synchronous positional shift from the cytoplasm to the apical domain in the developing wild-type intestine, consistent with a directional change of presumed apical vesicle trajectories during membrane polarity establishment (Fig.6b). Based on this finding we next examined whether the positioning of these and other, not yet well mapped, vesicle populations was dependent on bcAMs/actin during de novo polarized membrane biogenesis.…”

“…Polarity conversion was defined as the basolateral displacement of apical membrane components and polarity cues in any or all the 20 cells of the single-layered C. elegans intestine, with and without subsequent ectopic basolateral lumen formation (transformation of the basolateral membrane into a junction-bounded apical membrane with microvilli 5 . This full membrane polarity change can be tracked by the apical identity marker ERM-1 whose recruitment to the membrane indicates successful apical polarization 7 . Among the identified molecules, 3 were the key components of the branched-chain actin machinery (here referred to as branched-chain actin modulators/bcAMs; Fig.1): UNC-60/cofilin (actin-depolymerizing), ARX-2/ARP-2/3 (actin nucleating) and CAP-1 (actin capping).…”

“…We conclude that UNC-60, ARX-2, and CAP-1, like the previously identified apical polarity cues, are required to specify the position of the apical domain in the C. elegans intestine. The close similarity of the bcAM- and trafficking-dependent polarity phenotype made actin a candidate component of the proposed cytoskeletal structure that might direct vesicle trajectories to the membrane to asymmetrically insert the nascent apical domain (see Introduction, 7 .…”

“…(a) The proposed apicobasal polarity model (see text; 7 : the anterograde biosynthetic-secretory pathway is constrained towards the expanding apical domain and partitions apical and basolateral membrane domains; apical PARs and junction components are concomitantly or subsequently recruited to the apical domain, with apicolateral junction assembly securing the expanding apical domain (not shown; mode of PAR delivery is not determined). Side view of intestinal cell is shown, dashed box contains dynamic components during polarity establishment.…”