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MIT 7 72 - Coordinated cell-shape changes control epithelial movement

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DEVELOPMENT2671RESEARCH ARTICLEINTRODUCTIONContraction of an epithelial sheet to close an opening is a commonprocess in animal development and tissue repair, and requiresextensive modification of cell shape at the advancing epithelialmargin (Martin and Parkhurst, 2004). Many of the cellular andmolecular mechanisms controlling this event have been identifiedby analysis of Drosophila dorsal closure (reviewed by Harden, 2002;Jacinto et al., 2002b). Dorsal closure is one of the last morphogeneticmovements during Drosophila embryogenesis and occurs as theopposing fronts of the lateral epidermis converge dorsally, displacingthe underlying amnioserosa (Martinez-Arias, 1993). A central aspectof this process is the uniform elongation and constriction of marginalepidermal cells. This transforms the margin from a loose, scallopededge into a taut row of cells and probably ensures even epithelialadvance (Jacinto et al., 2002a; Young et al., 1993). The observedaccumulation of actin and myosin 2 in these cells suggests that thetransformation of the margin is achieved by a contractile actomyosin‘cable’. However, to what extent this represents a generalmechanism controlling epithelial movement is unclear.Similar to the Drosophila epidermis, the outer epithelium of thezebrafish embryo, the enveloping layer (EVL), is a simpleepithelium that spreads over the spherical yolk cell together with theunderlying deep cells, to ultimately seal the embryo on the vegetalside (Kane and Adams, 2002; Kimmel et al., 1995) (for illustration,see Fig. 1L). During EVL epiboly, the length of the advancingmargin must be reduced as the tissue closes around the vegetal yolkblastopore. Analysis of the related teleost Fundulus heteroclitus hasshown that this involves the constriction of marginal EVL cells(Keller and Trinkaus, 1987), similar to Drosophila dorsal closure.However, the mechanistic basis for EVL cell shape change remainsto be shown.Insights come from studies of both Fundulus and zebrafish, andsuggest a potential role of the underlying yolk syncytial layer (YSL)during EVL epiboly. The YSL constitutes the surface of the yolk celldirectly beneath the blastoderm and undergoes epibolysimultaneously with the EVL. In Fundulus, the leading edges ofmarginal EVL cells were found to be stably attached to the YSLwhile undergoing constriction. Furthermore, a dense network ofmicrofilaments was detected in the YSL along the EVL margin inFundulus (Betchaku and Trinkaus, 1978; Keller and Trinkaus,1987). A similar accumulation of actin within the YSL has recentlyalso been reported in zebrafish, and was implicated in EVL epiboly(Cheng et al., 2004). This suggests that the YSL may controlchanges in marginal EVL cell shape via an actin-dependentmechanism. However, YSL-specific analysis supporting a role foractin during EVL epiboly has so far been missing. Furthermore, themolecular mechanisms controlling actin function in the YSL are stillunknown.In this study, we show that localized recruitment of actin andmyosin 2 within the YSL correlates with EVL cell-shape change.Furthermore, we show that this process is dependent on Msn1, azebrafish ortholog of the Drosophila Ste20-like kinase Misshapen.Similarly, we show that Drosophila Misshapen is required foractin/myosin 2-based cell constriction at the epidermal marginduring dorsal closure. These findings point to a conservedmechanism of actin/myosin 2 recruitment and cell constrictionduring epithelial morphogenesis in zebrafish and Drosophila.MATERIALS AND METHODSZebrafish embryo maintenanceAll embryos were obtained from zebrafish AB, TL and Wik wild-type lines,grown at 31°C, and manipulated in E3 zebrafish embryo medium or Danieaubuffer.Drosophila stocks and genetic crossesWild-type embryos were from the Oregon R strain. UAS lines wereexpressed using the Gal4 system (Brand and Perrimon, 1993). The Gal4lines e22c-Gal4, LE-Gal4, en-Gal4 (Brand and Perrimon, 1993), and c381-Gal4 (AS-Gal4) and flies carrying the GFP balancer TM3, gal4-twi, UAS-2xEGFP (TTG) were kindly provided by the Bloomington stock centre.msn172was kindly provided by J. Treisman. Homozygous msn172embryosCoordinated cell-shape changes control epithelial movementin zebrafish and DrosophilaMathias Köppen1, Beatriz García Fernández2, Lara Carvalho1, Antonio Jacinto2and Carl-Philipp Heisenberg1,*Epithelial morphogenesis depends on coordinated changes in cell shape, a process that is still poorly understood. During zebrafishepiboly and Drosophila dorsal closure, cell-shape changes at the epithelial margin are of critical importance. Here evidence isprovided for a conserved mechanism of local actin and myosin 2 recruitment during theses events. It was found that during epibolyof the zebrafish embryo, the movement of the outer epithelium (enveloping layer) over the yolk cell surface involves theconstriction of marginal cells. This process depends on the recruitment of actin and myosin 2 within the yolk cytoplasm along themargin of the enveloping layer. Actin and myosin 2 recruitment within the yolk cytoplasm requires the Ste20-like kinase Msn1, anorthologue of Drosophila Misshapen. Similarly, in Drosophila, actin and myosin 2 localization and cell constriction at the margin ofthe epidermis mediate dorsal closure and are controlled by Misshapen. Thus, this study has characterized a conserved mechanismunderlying coordinated cell-shape changes during epithelial morphogenesis.KEY WORDS: Zebrafish epiboly, Drosophila dorsal closure, Cell shape, Misshapen, Actin, Myosin 2Development 133, 2671-2681 (2006) doi:10.1242/dev.024391Max Planck Institute of Molecular Cell Biology and Genetics, Pfotenhauerstr.108,01307 Dresden, Germany. 2Instituto de Medicina Molecular, Edificio Egas Moniz, Av.Profesor Egas Moniz, 1649-028 Lisbon, Portugal.*Author for correspondence (e-mail: [email protected])Accepted 9 May 2006DEVELOPMENT2672expressing GFP-Actin in the engrailed domains were generated by crossingUAS-GFP-Actin/Cyo; msn172/TTG flies to en-Gal4; msn172/TTG flies. Astrain carrying a UAS-DN-msn construct was a gift from Yong Rao (Houallaet al., 2005). Homozygous UAS-DN-msn flies were crossed with e22c-Gal4,LE-Gal4 or AS-Gal4 flies to express DN-msn in the whole epidermis,marginal cells of the epidermis, or the amnioserosa, respectively.Identification of msn genes and phylogenetic analysismisshapen-type genes were identified in the zebrafish genome using BLASTanalysis. cDNA sequences were obtained from PubMed.


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