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A functional genomic screen combined with time-lapse microscopy uncovers a novel set of genes involved in dorsal closure of Drosophila embryos.

Jankovics F, Henn L, Bujna Á, Vilmos P, Kiss N, Erdélyi M - PLoS ONE (2011)

Bottom Line: Morphogenesis, the establishment of the animal body, requires the coordinated rearrangement of cells and tissues regulated by a very strictly-determined genetic program.We show that pbl regulates actin accumulation and protrusion dynamics in the leading edge of the migrating epithelial cells.Finally, we provide evidence that pbl is involved in closure of the adult thorax, suggesting its general requirement in epithelial closure processes.

View Article: PubMed Central - PubMed

Affiliation: Institute of Genetics, Biological Research Center of the Hungarian Academy of Sciences, Szeged, Hungary. jankovic@brc.hu

ABSTRACT
Morphogenesis, the establishment of the animal body, requires the coordinated rearrangement of cells and tissues regulated by a very strictly-determined genetic program. Dorsal closure of the epithelium in the Drosophila melanogaster embryo is one of the best models for such a complex morphogenetic event. To explore the genetic regulation of dorsal closure, we carried out a large-scale RNA interference-based screen in combination with in vivo time-lapse microscopy and identified several genes essential for the closure or affecting its dynamics. One of the novel dorsal closure genes, the small GTPase activator pebble (pbl), was selected for detailed analysis. We show that pbl regulates actin accumulation and protrusion dynamics in the leading edge of the migrating epithelial cells. In addition, pbl affects dorsal closure dynamics by regulating head involution, a morphogenetic process mechanically coupled with dorsal closure. Finally, we provide evidence that pbl is involved in closure of the adult thorax, suggesting its general requirement in epithelial closure processes.

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Quantification of abnormal dorsal closure dynamics.(A and B) Graphs showing closure kinetics of the dorsal hole in a buffer-injected control embryo, in a homozygous pbl3 mutant embryo and embryos silenced for pbl and Arf51F. For each category, data of individual representative embryos are shown. (A) “Width” represents the maximal distance between zippering ends. (B) “Height” represents the maximal distance between the converging epithelial layers. Velocity of the epithelial sheet translocation (v), the rate constant of zippering (kz) and the fractional contribution of zippering (fz) to the velocity of the closure were calculated as described [22].
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pone-0022229-g004: Quantification of abnormal dorsal closure dynamics.(A and B) Graphs showing closure kinetics of the dorsal hole in a buffer-injected control embryo, in a homozygous pbl3 mutant embryo and embryos silenced for pbl and Arf51F. For each category, data of individual representative embryos are shown. (A) “Width” represents the maximal distance between zippering ends. (B) “Height” represents the maximal distance between the converging epithelial layers. Velocity of the epithelial sheet translocation (v), the rate constant of zippering (kz) and the fractional contribution of zippering (fz) to the velocity of the closure were calculated as described [22].

Mentions: Silencing of Arf51F also induced abnormal closure dynamics. Arf51F encodes for a conserved member of the Arf family of small GTPases regulating membrane trafficking. However, the mammalian homolog of Arf51F (Arf6) has been implicated in the regulation of subcortical actin remodeling, cell adhesion dynamics and cell migration [35]–[36]. Null mutants of Arf51F are viable but they exhibit defects in cytokinesis in the male germ line and in the brain [37]. In addition to these phenotypes, live imaging of the embryonic morphogenesis also revealed a requirement for Arf51F in dorsal closure. In Arf51F-silenced embryos, the convergence of the lateral epithelial sheets took place normally, while zippering was inefficient in both anterior and posterior ends of the dorsal hole (Figure 3, Movie S3). As a consequence, the dorsal opening became abnormally narrow. The abnormal dynamics phenotype was characterized in a quantitative manner using a mathematical model of dorsal closure [22]. In the movies, quantitative features (height and width) of the dorsal opening were measured and the velocity of the epithelial sheet translocation (v), as well as the fractional contribution of zippering (fz) to the velocity of the closure were calculated. Silencing of Arf51F resulted in a decrease of fz suggesting that Arf51F function is essential for efficient zippering (Figure 4, Table S2).


A functional genomic screen combined with time-lapse microscopy uncovers a novel set of genes involved in dorsal closure of Drosophila embryos.

Jankovics F, Henn L, Bujna Á, Vilmos P, Kiss N, Erdélyi M - PLoS ONE (2011)

Quantification of abnormal dorsal closure dynamics.(A and B) Graphs showing closure kinetics of the dorsal hole in a buffer-injected control embryo, in a homozygous pbl3 mutant embryo and embryos silenced for pbl and Arf51F. For each category, data of individual representative embryos are shown. (A) “Width” represents the maximal distance between zippering ends. (B) “Height” represents the maximal distance between the converging epithelial layers. Velocity of the epithelial sheet translocation (v), the rate constant of zippering (kz) and the fractional contribution of zippering (fz) to the velocity of the closure were calculated as described [22].
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3140500&req=5

pone-0022229-g004: Quantification of abnormal dorsal closure dynamics.(A and B) Graphs showing closure kinetics of the dorsal hole in a buffer-injected control embryo, in a homozygous pbl3 mutant embryo and embryos silenced for pbl and Arf51F. For each category, data of individual representative embryos are shown. (A) “Width” represents the maximal distance between zippering ends. (B) “Height” represents the maximal distance between the converging epithelial layers. Velocity of the epithelial sheet translocation (v), the rate constant of zippering (kz) and the fractional contribution of zippering (fz) to the velocity of the closure were calculated as described [22].
Mentions: Silencing of Arf51F also induced abnormal closure dynamics. Arf51F encodes for a conserved member of the Arf family of small GTPases regulating membrane trafficking. However, the mammalian homolog of Arf51F (Arf6) has been implicated in the regulation of subcortical actin remodeling, cell adhesion dynamics and cell migration [35]–[36]. Null mutants of Arf51F are viable but they exhibit defects in cytokinesis in the male germ line and in the brain [37]. In addition to these phenotypes, live imaging of the embryonic morphogenesis also revealed a requirement for Arf51F in dorsal closure. In Arf51F-silenced embryos, the convergence of the lateral epithelial sheets took place normally, while zippering was inefficient in both anterior and posterior ends of the dorsal hole (Figure 3, Movie S3). As a consequence, the dorsal opening became abnormally narrow. The abnormal dynamics phenotype was characterized in a quantitative manner using a mathematical model of dorsal closure [22]. In the movies, quantitative features (height and width) of the dorsal opening were measured and the velocity of the epithelial sheet translocation (v), as well as the fractional contribution of zippering (fz) to the velocity of the closure were calculated. Silencing of Arf51F resulted in a decrease of fz suggesting that Arf51F function is essential for efficient zippering (Figure 4, Table S2).

Bottom Line: Morphogenesis, the establishment of the animal body, requires the coordinated rearrangement of cells and tissues regulated by a very strictly-determined genetic program.We show that pbl regulates actin accumulation and protrusion dynamics in the leading edge of the migrating epithelial cells.Finally, we provide evidence that pbl is involved in closure of the adult thorax, suggesting its general requirement in epithelial closure processes.

View Article: PubMed Central - PubMed

Affiliation: Institute of Genetics, Biological Research Center of the Hungarian Academy of Sciences, Szeged, Hungary. jankovic@brc.hu

ABSTRACT
Morphogenesis, the establishment of the animal body, requires the coordinated rearrangement of cells and tissues regulated by a very strictly-determined genetic program. Dorsal closure of the epithelium in the Drosophila melanogaster embryo is one of the best models for such a complex morphogenetic event. To explore the genetic regulation of dorsal closure, we carried out a large-scale RNA interference-based screen in combination with in vivo time-lapse microscopy and identified several genes essential for the closure or affecting its dynamics. One of the novel dorsal closure genes, the small GTPase activator pebble (pbl), was selected for detailed analysis. We show that pbl regulates actin accumulation and protrusion dynamics in the leading edge of the migrating epithelial cells. In addition, pbl affects dorsal closure dynamics by regulating head involution, a morphogenetic process mechanically coupled with dorsal closure. Finally, we provide evidence that pbl is involved in closure of the adult thorax, suggesting its general requirement in epithelial closure processes.

Show MeSH
Related in: MedlinePlus