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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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Related in: MedlinePlus

Pbl is required for the morphogenesis of the dorsal epithelium.(A–C) Cuticle preparations of embryos. (A) Wild-type cuticle. (B) Cuticle of an embryo injected with dsRNA for pbl. (C) Cuticle of a homozygous pbl3 mutant embryo. (D and E) Frames from movies of embryos expressing ZCL0423 protein trap- EGFP fusion protein. Embryos are shown in dorsal view, scale bars represent 50 µm. (D) Wild type embryo. (E) pbl3/pbl3 mutant embryo.
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pone-0022229-g006: Pbl is required for the morphogenesis of the dorsal epithelium.(A–C) Cuticle preparations of embryos. (A) Wild-type cuticle. (B) Cuticle of an embryo injected with dsRNA for pbl. (C) Cuticle of a homozygous pbl3 mutant embryo. (D and E) Frames from movies of embryos expressing ZCL0423 protein trap- EGFP fusion protein. Embryos are shown in dorsal view, scale bars represent 50 µm. (D) Wild type embryo. (E) pbl3/pbl3 mutant embryo.

Mentions: Pbl, one of the genes exhibiting thorax and dorsal closure defects, was further characterized. We investigated whether the RNAi phenotype is similar to the phenotypes of the loss-of-function pbl mutants by comparing the cuticles of embryos homozygous for the strong hypomorphic pbl3 allele and the cuticles of pbl dsRNA-treated embryos. Cuticle abnormalities were detected in the pbl mutant embryos identical to the embryos silenced for pbl confirming the accuracy of our screening method. In most of the embryos (88%, n = 24) injected with pbl dsRNA, a serious disturbance of the epithelial matching was observed. Instead of the segmentally repeated rows of cuticle hairs present in wild-type embryos, the pbl-silenced embryos displayed disorganized rows of hairs meeting at one point around the dorsal midline (Figure 6A–C). We detected identical segmental misalignments in all of the homozygous pbl3 mutant embryos. In addition, live imaging of pbl mutants expressing EGFP specifically in the DME cells revealed defects of closure dynamics identical to the pbl-silenced embryos (Figure 6D,E). In pbl mutants, quantitative parameters of the closure were similar to that of pbl-silenced embryos indicating further that the RNAi phenotype precisely phenocopies the loss-of-function phenotype of the pbl gene (Figure 4, Movie S4, 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)

Pbl is required for the morphogenesis of the dorsal epithelium.(A–C) Cuticle preparations of embryos. (A) Wild-type cuticle. (B) Cuticle of an embryo injected with dsRNA for pbl. (C) Cuticle of a homozygous pbl3 mutant embryo. (D and E) Frames from movies of embryos expressing ZCL0423 protein trap- EGFP fusion protein. Embryos are shown in dorsal view, scale bars represent 50 µm. (D) Wild type embryo. (E) pbl3/pbl3 mutant embryo.
© Copyright Policy
Related In: Results  -  Collection

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

pone-0022229-g006: Pbl is required for the morphogenesis of the dorsal epithelium.(A–C) Cuticle preparations of embryos. (A) Wild-type cuticle. (B) Cuticle of an embryo injected with dsRNA for pbl. (C) Cuticle of a homozygous pbl3 mutant embryo. (D and E) Frames from movies of embryos expressing ZCL0423 protein trap- EGFP fusion protein. Embryos are shown in dorsal view, scale bars represent 50 µm. (D) Wild type embryo. (E) pbl3/pbl3 mutant embryo.
Mentions: Pbl, one of the genes exhibiting thorax and dorsal closure defects, was further characterized. We investigated whether the RNAi phenotype is similar to the phenotypes of the loss-of-function pbl mutants by comparing the cuticles of embryos homozygous for the strong hypomorphic pbl3 allele and the cuticles of pbl dsRNA-treated embryos. Cuticle abnormalities were detected in the pbl mutant embryos identical to the embryos silenced for pbl confirming the accuracy of our screening method. In most of the embryos (88%, n = 24) injected with pbl dsRNA, a serious disturbance of the epithelial matching was observed. Instead of the segmentally repeated rows of cuticle hairs present in wild-type embryos, the pbl-silenced embryos displayed disorganized rows of hairs meeting at one point around the dorsal midline (Figure 6A–C). We detected identical segmental misalignments in all of the homozygous pbl3 mutant embryos. In addition, live imaging of pbl mutants expressing EGFP specifically in the DME cells revealed defects of closure dynamics identical to the pbl-silenced embryos (Figure 6D,E). In pbl mutants, quantitative parameters of the closure were similar to that of pbl-silenced embryos indicating further that the RNAi phenotype precisely phenocopies the loss-of-function phenotype of the pbl gene (Figure 4, Movie S4, 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