Trafficking of Crumbs3 during cytokinesis is crucial for lumen formation.
Although lumen generation has been extensively studied through so-called cyst-formation assays in Madin-Darby canine kidney (MDCK) cells, an underlying mechanism that leads to the initial appearance of a solitary lumen remains elusive. Lumen formation is thought to take place at early stages in aggregates containing only a few cells. Evolutionarily conserved polarity protein complexes, namely the Crumbs, Par, and Scribble complexes, establish apicobasal polarity in epithelial cells, and interference with their function impairs the regulated formation of solitary epithelial lumina. Here, we demonstrate that MDCK cells form solitary lumina during their first cell division. Before mitosis, Crumbs3a becomes internalized and concentrated in Rab11-positive recycling endosomes. These compartments become partitioned in both daughter cells and are delivered to the site of cytokinesis, thus forming the first apical membrane, which will eventually form a lumen. Endosome trafficking in this context appears to depend on the mitotic spindle apparatus and midzone microtubules. Furthermore, we show that this early lumen formation is regulated by the apical polarity complexes because Crumbs3 assists in the recruitment of aPKC to the forming apical membrane and interference with their function can lead to the formation of a no-lumen or multiple-lumen phenotype at the two-cell stage.
Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, MI 48109, USA.
Marc A
MA
Schlüter
Catherine S
CS
Pfarr
Jay
J
Pieczynski
Eileen L
EL
Whiteman
Toby W
TW
Hurd
Shuling
S
Fan
Chia-Jen
CJ
Liu
Ben
B
Margolis
D000818:Animals; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D016764:Cell Polarity; D048749:Cytokinesis / Q000502:physiology*; D003599:Cytoskeleton / Q000378:metabolism; D004285:Dogs; D004705:Endocytosis / Q000502:physiology; D011992:Endosomes / Q000378:metabolism; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D008870:Microtubules / Q000378:metabolism; D009024:Morphogenesis; D046912:Multiprotein Complexes / Q000378:metabolism; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D011493:Protein Kinase C / Q000378:metabolism; D021381:Protein Transport / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D020691:rab GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism; D000099193:rab11 GTP-Binding Proteins
D018797:Cell Cycle Proteins; D008565:Membrane Proteins; D046912:Multiprotein Complexes; D020033:Protein Isoforms; D011493:Protein Kinase C; D011993:Recombinant Fusion Proteins; D020691:rab GTP-Binding Proteins; D000099193:rab11 GTP-Binding Proteins; C115015:PKC-3 protein