Fig. 10. Model of how IRSp53 controls nascent lumen formation in epithelial tubular tissues.
Control MDCK cells plated on Matrigel in Matrigel-containing media undergo symmetry-breaking events immediately after the first cell division, when after cytokinetic furrow ingression a midbody is formed4. Around the midbody, the apical membrane initiation site (AMIS) is assembled establishing the location of the nascent lumen. AMIS assembly requires the polarized transport of highly-charged and -glycosylated transmembrane proteins, such as Podocalyxin (PODXL), that from the outer cell face become endocytosed and traffic via vesicular recycling carriers to the newly formed opposing plasma membrane. At this site, repulsive charges contribute to generate interconnected mini-lumens (leftmost inset show a Correlative Light Electron Microscopy-CLEM-analysis of MDCK cells capture after the very first cell division) that will evolve to form a single apical lumen, typical of an epithelial cyst. IRSp53 is required both for directing the trafficking of PODXL and to ensure its targeting at the AMIS via stabilizing the GTPase RAB35 (not shown), and through its interaction with the actin capping protein Eps8 (not shown). At the AMIS, IRSp53 through its membrane deforming and curvature sensing I-BAR domain ensures the structural integrity, continuity and correct shape of the opposing plasma membrane at two-cells stage. Consistent with this, IRSp53 genetic loss leads to interruption of the continuity of the PM at the AMIS (top rightmost inset, yellow arrowhead indicate membrane discontinuities), with the formation of inter-cytoplasmic bridges, and occasionally ectopic lateral lumen (bottom inset, magenta line outlines the intervening PM and the extra lumen).