Rho1 regulates adherens junction remodeling by promoting recycling endosome formation through activation of myosin II.
Bottom Line: We demonstrate that Rho1 also influences AJ remodeling by regulating the formation of DE-cadherin-containing, Rab11-positive recycling endosomes in Drosophila postmitotic pupal eye epithelia.This effect of Rho1 is mediated through Rok-dependent, but not MLCK-dependent, stimulation of myosin II activity yet independent of its effects upon actin remodeling.This work identifies spatially distinct functions for Rho1 in the regulation of DE-cadherin-containing vesicular trafficking during AJ remodeling in live epithelia.
Affiliation: ICCE Institute, Washington University School of Medicine, St. Louis, MO 63110 Department of Cell Biology and Physiology, Washington University School of Medicine, St. Louis, MO 63110 Department of Developmental Biology, Washington University School of Medicine, St. Louis, MO 63110.Show MeSH
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Mentions: The altered staining pattern of Rab11 in the AJ region of Rho1 LoF clones was not due to increased apical size of Rho1 LoF clones, as depleting Rok, a major downstream effector of Rho1 that mediates Rho's effects upon actomyosin contractility, also resulted in PECs with increased apical cell size (Supplemental Table S2; Warner and Longmore, 2009a), yet the Rab11 staining pattern was unchanged from that for wild-type cells (Figure 5Aâ€²â€²); nor was it simply the result of disrupting AJs, as clonal loss of DE-cadherin through the homozygous clonal expression of its LoF allele ShgR69 did not reduce Rab11 staining (Supplemental Figure S3, Gâ€“Gâ€²â€²). In embryos lacking one genomic copy of Rho1, the level of Rab11 protein was reduced by 25% compared with wild-type embryos (Figure 2G), suggesting that the altered Rab11 staining pattern at the AJ region of Rho1-deficient PECs may, in part, be due to decreased Rab11 protein level. The lethal effects of Rho172F homozygotes and ubiquitously targeted Rho1 RNAi precluded analysis of complete lack of Rho1 upon Rab11 protein levels in vivo. In further controls, the staining pattern of early endosomes (Rab5) and Golgi (Lva and dGM130) were unaffected by Rho1 loss (Figure 2, Fâ€“Fâ€²â€², and Supplemental Figure S4, D and E). Although Rab11 was sufficient to rescue the AJ defect between Rho1-deficient PECs (see later discussion), we were unable to determine whether Rab11 alone was necessary for AJ maintenance due to lethality of Rab11 depletion, even when the caspase inhibitor p35 was concurrently expressed (Supplemental Figure S3, Aâ€“Aâ€²â€²). We also tested whether removing a genomic copy of Rab11, through the use of the strongest allele Rab11EP3017, in a heterozygous Rho172F background resulted in AJ disruptions. AJs remained intact in the sensitized background, in which one genomic copy of Rho1 and Rab11 remained (Supplemental Figure S3, Jâ€“Jâ€²â€²â€²). In controls, heterozygous expressions of Rho172F and Rab11EP3017 individually in the whole animal were viable, and pupal eye PECs did not exhibit any AJ defect (Supplemental Figure S3, Jâ€² and Jâ€²â€²).
Affiliation: ICCE Institute, Washington University School of Medicine, St. Louis, MO 63110 Department of Cell Biology and Physiology, Washington University School of Medicine, St. Louis, MO 63110 Department of Developmental Biology, Washington University School of Medicine, St. Louis, MO 63110.