Cellularization in Drosophila melanogaster is disrupted by the inhibition of Rho activity and the activation of Cdc42 function

被引:58
作者
Crawford, JM
Harden, N
Leung, T
Lim, L
Kiehart, DP
机构
[1] Duke Univ, Med Ctr, Dept Cell Biol, Durham, NC 27710 USA
[2] Natl Univ Singapore, Glaxo IMCB Grp, Inst Mol & Cell Biol, Singapore 117609, Singapore
[3] Inst Neurol, Dept Neurochem, London WC1N 1PJ, England
关键词
D O I
10.1006/dbio.1998.9061
中图分类号
Q [生物科学];
学科分类号
07 ; 0710 ; 09 ;
摘要
Regulation of cytoskeletal dynamics is essential for cell shape change and morphogenesis. Drosophila melanogaster embryos offer a well-defined system for observing alterations in the cytoskeleton during the process of cellularization, a specialized form of cytokinesis. During cellularization, the actomyosin cytoskeleton forms a hexagonal array and drives invagination of the plasma membrane between the nuclei located at the cortex of the syncytial blastoderm. Rho, Rac, and Cdc42 proteins are members of the Rho subfamily of Ras-related G proteins that are involved in the formation and maintenance of the actin cytoskeleton throughout phylogeny and in D. melanogaster. To investigate how Rho subfamily activity affects the cytoskeleton during cellularization stages, embryos were microinjected with C3 exoenzyme from Clostridium botulinum or with wild-type, constitutively active, or dominant negative versions of Rho, Rac, and Cdc42 proteins. C3 exoenzyme ADP-ribosylates and inactivates Rho with high specificity, whereas constitutively active dominant mutations remain in the activated GTP-bound state to activate downstream effecters. Dominant negative mutations likely inhibit endogenous small G protein activity by sequestering exchange factors. Of the 10 agents microinjected, C3 exoenzyme, constitutively active Cdc42, and dominant negative Rho have a specific and indistinguishable effect: the actomyosin cytoskeleton is disrupted, cellularization halts, and embryogenesis arrests. Time-lapse video records of DIC imaged embryos show that nuclei in injected regions move away from the cortex of the embryo, thereby phenocopying injections of cytochalasin or antimyosin. Rhodamine phalloidin staining reveals that the actin-based hexagonal array normally seen during cellularization is disrupted in a dose-dependent fashion. Additionally, DNA stain reveals that nuclei in the microinjected embryos aggregate in regions that correspond to actin disruption. These embryos halt in cellularization and do not proceed to gastrulation. We conclude that Rho activity and Cdc42 regulation are required for cytoskeletal function in actomyosin-driven furrow canal formation and nuclear positioning. (C) 1998 Academic Press.
引用
收藏
页码:151 / 164
页数:14
相关论文
共 78 条
  • [11] CDC42 AND RAC1 CONTROL DIFFERENT ACTIN-DEPENDENT PROCESSES IN THE DROSOPHILA WING DISC EPITHELIUM
    EATON, S
    AUVINEN, P
    LUO, LQ
    JAN, YN
    SIMONS, K
    [J]. JOURNAL OF CELL BIOLOGY, 1995, 131 (01) : 151 - 164
  • [12] Edwards KA, 1996, DEVELOPMENT, V122, P1499
  • [13] DOMINANT INHIBITORY MUTATIONS IN THE MG-2+-BINDING SITE OF RASH PREVENT ITS ACTIVATION BY GTP
    FARNSWORTH, CL
    FEIG, LA
    [J]. MOLECULAR AND CELLULAR BIOLOGY, 1991, 11 (10) : 4822 - 4829
  • [14] Fehon RG, 1997, GENETICS, V146, P245
  • [15] INHIBITION OF NIH-3T3 CELL-PROLIFERATION BY A MUTANT RAS PROTEIN WITH PREFERENTIAL AFFINITY FOR GDP
    FEIG, LA
    COOPER, GM
    [J]. MOLECULAR AND CELLULAR BIOLOGY, 1988, 8 (08) : 3235 - 3243
  • [16] FOE VE, 1983, J CELL SCI, V61, P31
  • [17] FOE VE, 1993, DEV DROSOPHILA MELAN
  • [18] Rho GTPases and the actin cytoskeleton
    Hall, A
    [J]. SCIENCE, 1998, 279 (5350) : 509 - 514
  • [19] HARDEN N, 1995, DEVELOPMENT, V121, P903
  • [20] CHARACTERIZATION OF RHO GTPASE FAMILY HOMOLOGS IN DROSOPHILA-MELANOGASTER - OVEREXPRESSING RHO1 IN RETINAL CELLS CAUSES A LATE DEVELOPMENTAL DEFECT
    HARIHARAN, IK
    HU, KQ
    ASHA, H
    QUINTANILLA, A
    EZZELL, RM
    SETTLEMAN, J
    [J]. EMBO JOURNAL, 1995, 14 (02) : 292 - 302