Dual role of Cdc42 in spindle orientation control of adherent cells.
The spindle orientation is regulated by the interaction of astral microtubules with the cell cortex. We have previously shown that spindles in nonpolarized adherent cells are oriented parallel to the substratum by an actin cytoskeleton- and phosphatidylinositol 3,4,5-triphosphate [PtdIns(3,4,5)P3]-dependent mechanism. Here, we show that Cdc42, a Rho family of small GTPases, has an essential role in this mechanism of spindle orientation by regulating both the actin cytoskeleton and PtdIns(3,4,5)P3. Knockdown of Cdc42 suppresses PI(3)K activity in M phase and induces spindle misorientation. Moreover, knockdown of Cdc42 disrupts the cortical actin structures in metaphase cells. Our results show that p21-activated kinase 2 (PAK2), a target of Cdc42 and/or Rac1, plays a key role in regulating actin reorganization and spindle orientation downstream from Cdc42. Surprisingly, PAK2 regulates spindle orientation in a kinase activity-independent manner. BetaPix, a guanine nucleotide exchange factor for Rac1 and Cdc42, is shown to mediate this kinase-independent function of PAK2. This study thus demonstrates that spindle orientation in adherent cells is regulated by two distinct pathways downstream from Cdc42 and uncovers a novel role of the Cdc42-PAK2-betaPix-actin pathway for this mechanism.
Mitsushima
Masaru
M
Department of Cell and Developmental Biology, Graduate School of Biostudies, Kyoto University, Sakyo-ku, Kyoto 606-8502, Japan.
Toyoshima
Fumiko
F
Nishida
Eisuke
E
D000199:Actins / Q000378:metabolism; D000818:Animals; D002448:Cell Adhesion / Q000502:physiology*; D002453:Cell Cycle / Q000502:physiology; D016764:Cell Polarity; D004789:Enzyme Activation; D020662:Guanine Nucleotide Exchange Factors / Q000235:genetics / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D064067:Rho Guanine Nucleotide Exchange Factors; D015398:Signal Transduction / Q000502:physiology; D008941:Spindle Apparatus / Q000378:metabolism*; D020764:cdc42 GTP-Binding Protein / Q000235:genetics / Q000378:metabolism*; D054462:p21-Activated Kinases / Q000235:genetics / Q000378:metabolism; D020830:rac1 GTP-Binding Protein / Q000235:genetics / Q000378:metabolism
D000199:Actins; D020662:Guanine Nucleotide Exchange Factors; D018129:Phosphatidylinositol Phosphates; D034741:RNA, Small Interfering; D011993:Recombinant Fusion Proteins; D064067:Rho Guanine Nucleotide Exchange Factors; C060974:phosphatidylinositol 3,4,5-triphosphate; D019869:Phosphatidylinositol 3-Kinases; D054462:p21-Activated Kinases; D020764:cdc42 GTP-Binding Protein; D020830:rac1 GTP-Binding Protein