Tuba, a Cdc42 GEF, is required for polarized spindle orientation during epithelial cyst formation.
The Cdc42 guanosine triphosphatase is essential for cell polarization in several organisms and in vitro for the organization of polarized epithelial cysts. A long-standing question concerns the identity of the guanine nucleotide exchange factor (GEF) that controls this process. Using Madin-Darby canine kidney cells grown in Matrigel, we screened 70 GEFs by RNA interference. Of these, six positives were identified that caused a multilumen phenotype, including Tuba, a Cdc42-specific GEF localized below the apical cortex. Loss of Tuba abolishes Cdc42 enrichment at the apical cortex. Normal lumen formation is rescued by human Tuba or active Cdc42 but not by a GEF-negative Tuba mutant. Silencing Cdc42 causes a similar phenotype, including multilumen formation and reduced atypical protein kinase C (aPKC) activity. Lumen disorganization after depletion of Tuba or Cdc42 or inhibition of aPKC is caused by defective spindle orientation. Together, our findings implicate Tuba as a key activator of the Cdc42 GTPase during epithelial ductal morphogenesis, which in turn activates apical aPKC to ensure that spindles orient parallel to the lateral plane.
Qin
Yi
Y
Center for Cell Signaling, University of Virginia School of Medicine, Charlottesville, VA 22908, USA.
Meisen
Walter H
WH
Hao
Yi
Y
Macara
Ian G
IG
D000818:Animals; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism*; D016764:Cell Polarity / Q000502:physiology*; D002478:Cells, Cultured; D004285:Dogs; D004847:Epithelial Cells / Q000166:cytology / Q000378:metabolism*; D020662:Guanine Nucleotide Exchange Factors / Q000235:genetics / Q000378:metabolism*; D034622:RNA Interference; D015398:Signal Transduction; D020764:cdc42 GTP-Binding Protein / Q000235:genetics / Q000378:metabolism*
D018797:Cell Cycle Proteins; D020662:Guanine Nucleotide Exchange Factors; D020764:cdc42 GTP-Binding Protein