Search PubMed⌕ Search

Biomedical subjects

Antonio Jacinto

Publications and source records attributed to Antonio Jacinto.

8 recordsLinked to original sources
PubMed2005

The small GTPase Rac plays multiple roles in epithelial sheet fusion--dynamic studies of Drosophila dorsal closure.

The coordinated migration and fusion of epithelial sheets is a crucial morphogenetic tool used on numerous occasions during the normal development of an embryo and re-activated as part of the wound healing response. Drosophila dorsal closure, whereby a hole in the embryonic epithelium is zipped closed late in embryogenesis, serves as an excellent, genetically tractable model for epithelial migration. Using live confocal imaging, we have dissected multiple roles for the small GTPase Rac in this process. We show that constitutive activation of Rac1 leads to excessive assembly of lamellipodia and precocious halting of epithelial sweeping, possibly through premature activation of contact-inhibition machinery. Conversely, blocking Rac activity, either by loss-of-function mutations or expression of dominant negative Rac1, disables the assembly of both actin cable and protrusions by epithelial cells. Movies of mutant embryos show that continued contraction of the amnioserosa is sufficient to draw the epithelial edges towards one another, allowing the zipper machinery to bypass non-functioning regions of leading edge. In addition to illustrating the key role of Rac in organization of leading edge actin, loss-of-function mutants also provide substantive proof that Rac acts upstream in the Jun N-terminal kinase (JNK) cascade to direct epithelial cell shape changes during dorsal closure.

Animals↗
PubMed2005

Live imaging of wound inflammation in Drosophila embryos reveals key roles for small GTPases during in vivo cell migration.

Aa robust inflammatory response to tissue damage and infection is conserved across almost all animal phyla. Neutrophils and macrophages, or their equivalents, are drawn to the wound site where they engulf cell and matrix debris and release signals that direct components of the repair process. This orchestrated cell migration is clinically important, and yet, to date, leukocyte chemotaxis has largely been studied in vitro. Here, we describe a genetically tractable in vivo wound model of inflammation in the Drosophila melanogaster embryo that is amenable to cinemicroscopy. For the first time, we are able to examine the roles of Rho-family small GTPases during inflammation in vivo and show that Rac-mediated lamellae are essential for hemocyte motility and Rho signaling is necessary for cells to retract from sites of matrix- and cell-cell contacts. Cdc42 is necessary for maintaining cellular polarity and yet, despite in vitro evidence, is dispensable for sensing and crawling toward wound cues.

Animals↗
PubMed2005

Imaging cell movement during dorsal closure in Drosophila embryos.

During Drosophila embryogenesis, many cells and tissues undergo complex morphogenetic movements, such as ventral furrow formation, germ band extension and retraction, and dorsal closure. The best way to study and understand the cell behaviors during such tissue movements is to image them live using time course analysis. The Drosophila embryo lends itself perfectly to live imaging for several reasons: powerful genetics allow transgenic embryos expressing green fluorescent protein-fusion proteins to be quickly and easily generated and the expression domains of these reporters can be efficiently controlled using the GAL4-upstream activator sequence system. Embryos will survive for several hours mounted on a slide in a gas-permeable oil, such as Voltalef or Halocarbon, during which time they will undergo normal development and can be easily imaged using a confocal laser scanning microscope. Here, we describe in detail a protocol for live imaging of Drosophila embryos which, in our hands, is routinely used to study dorsal closure but is suitable for the live study of any process involving cell motility, be it a coordinated tissue movement such as dorsal closure or the movement of individual cells such as hemocytes within the embryo.

Animals↗
PubMed2002

Dynamic analysis of actin cable function during Drosophila dorsal closure.

Throughout development, a series of epithelial movements and fusions occur that collectively shape the embryo. They are dependent on coordinated reorganizations and contractions of the actin cytoskeleton within defined populations of epithelial cells. One paradigm morphogenetic movement, dorsal closure in the Drosophila embryo, involves closure of a dorsal epithelial hole by sweeping of epithelium from the two sides of the embryo over the exposed extraembryonic amnioserosa to form a seam where the two epithelial edges fuse together. The front row cells exhibit a thick actin cable at their leading edge. Here, we test the function of this cable by live analysis of GFP-actin-expressing embryos in which the cable is disrupted by modulating Rho1 signaling or by loss of non-muscle myosin (Zipper) function. We show that the cable serves a dual role during dorsal closure. It is contractile and thus can operate as a "purse string," but it also restricts forward movement of the leading edge and excess activity of filopodia/lamellipodia. Stripes of epithelium in which cable assembly is disrupted gain a migrational advantage over their wild-type neighbors, suggesting that the cable acts to restrain front row cells, thus maintaining a taut, free edge for efficient zippering together of the epithelial sheets.

Actins↗
PubMed2002

Dynamic analysis of dorsal closure in Drosophila: from genetics to cell biology.

Throughout development a series of epithelial bendings, sweepings, and fusions occur that collectively give shape to the embryo. These morphogenetic movements are driven by coordinated assembly and contraction of the actomyosin cytoskeleton in restricted populations of epithelial cells. One well-studied example of such a morphogenetic episode is dorsal closure in Drosophila embryogenesis. This process is tractable at a genetic level and has recently become the focus of live cell biology analysis because of the availability of flies expressing GFP-fusion proteins. This marriage of genetics and cell biology is very powerful and is allowing the dissection of fundamental signaling mechanisms that regulate the cytoskeletal reorganizations and contractions underlying coordinated tissue movements in the embryo.

Actins↗
PubMed2002

Wound healing recapitulates morphogenesis in Drosophila embryos.

The capacity to repair a wound is a fundamental survival mechanism that is activated at any site of damage throughout embryonic and adult life. To study the cell biology and genetics of this process, we have developed a wounding model in Drosophila melanogaster embryos that allows live imaging of rearrangements and changes in cell shape, and of the cytoskeletal machinery that draws closed an in vivo wound. Using embryos expressing green fluorescent protein (GFP) fusion proteins, we show that two cytoskeletal-dependent elements -- an actin cable and dynamic filopodial/lamellipodial protrusions -- are expressed by epithelial cells at the wound edge and are pivotal for repair. Modulating the activities of the small GTPases Rho and Cdc42 demonstrates that these actin-dependent elements have differing cellular functions, but that either alone can drive wound closure. The actin cable operates as a 'purse-string' to draw the hole closed, whereas filopodia are essential for the final 'knitting' together of epithelial cells at the end of repair. Our data suggest a more complex model for epithelial repair than previously envisaged and highlight remarkable similarities with the well-characterized morphogenetic movement of dorsal closure in Drosophila.

Actins↗
PubMed2002

Planar polarity and actin dynamics in the epidermis of Drosophila.

Dorsal closure is a morphogenetic process involving the coordinated convergence of two epithelial sheets to enclose the Drosophila melanogaster embryo. Specialized populations of cells at the edges of each epithelial sheet, the dorsal-most epidermal cells, emit actin-based processes that are essential for the proper enclosure of the embryo. Here we show that actin dynamics at the leading edge is preceded by a planar polarization of the dorsal-most epidermal cells associated with a reorganization of the cytoskeleton. An important consequence of this planar polarization is the formation of actin-nucleating centres at the leading edge, which are important in the dynamics of actin. We show that Wingless (Wg) signalling and Jun amino-terminal kinase (JNK) signalling have overlapping but different roles in these events.

Actins↗