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Ectopic expression screen identifies genes affecting Drosophila mesoderm development including the HSPG Trol.

Trisnadi N, Stathopoulos A - G3 (Bethesda) (2014)

Bottom Line: These include the FGF ligand Pyramus, α-integrins, E-cadherin, Cueball, EGFR, JAK/STAT signaling components, as well as the heparan sulfate proteoglycan (HSPG) Terribly reduced optic lobes (Trol).Our data support the view that both HSPGs function to support FGF-dependent processes in the early embryo as they share phenotypes with FGF mutants: Trol in terms of effects on mesoderm migration and caudal visceral mesoderm (CVM) migration and Sdc in terms of dorsal mesoderm specification.The differential roles uncovered for these two HSPGs suggest that HSPG cofactor choice may modify FGF-signaling outputs.

View Article: PubMed Central - PubMed

Affiliation: Division of Biology and Biological Engineering, California Institute of Technology, 1200 East California Boulevard, MC 114-96, Pasadena, California 91125.

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Embryos obtained from sdc, but not trol, mutant germline clones exhibit defects in Eve specification. (A) Schematic cross-section of the ventral half of an embryo at stage 11 during Eve specification. Mesoderm cells that reach the dorsal regions of the embryo (arrows) are able to receive signals, including FGF, from the ectoderm (dark orange) and undergo cell differentiation (dark blue). (B–G) Dorsal somatic mesoderm cell differentiation at stage 11 embryos is marked by Eve expression. (B) Wild-type whole embryo stained using anti-Eve antibody includes box showing region of magnification in subsequent panels. (C) Wild-type embryos have 10 clusters of three Eve+ cells. (D) trol germline clones show a normal number of Eve+ cells. (E) Embryos overexpressing trol in the ectoderm occasionally have clusters with four Eve+ cells, as indicated by the arrow. (F) sdc germline clones are missing clusters (arrows) and/or have reduced number of Eve+ cells within a cluster (arrowhead). (G) Embryos with sdc overexpressed in the ectoderm have multiple clusters with five or more Eve+ cells (arrows).
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fig6: Embryos obtained from sdc, but not trol, mutant germline clones exhibit defects in Eve specification. (A) Schematic cross-section of the ventral half of an embryo at stage 11 during Eve specification. Mesoderm cells that reach the dorsal regions of the embryo (arrows) are able to receive signals, including FGF, from the ectoderm (dark orange) and undergo cell differentiation (dark blue). (B–G) Dorsal somatic mesoderm cell differentiation at stage 11 embryos is marked by Eve expression. (B) Wild-type whole embryo stained using anti-Eve antibody includes box showing region of magnification in subsequent panels. (C) Wild-type embryos have 10 clusters of three Eve+ cells. (D) trol germline clones show a normal number of Eve+ cells. (E) Embryos overexpressing trol in the ectoderm occasionally have clusters with four Eve+ cells, as indicated by the arrow. (F) sdc germline clones are missing clusters (arrows) and/or have reduced number of Eve+ cells within a cluster (arrowhead). (G) Embryos with sdc overexpressed in the ectoderm have multiple clusters with five or more Eve+ cells (arrows).

Mentions: Pericardial and dorsal somatic muscle cells derived from the dorsal mesoderm are known to express Even-skipped (Eve) (Frasch 1999) and require proper migration of the mesoderm at an earlier stage prior to their specification. Once mesoderm cells migrate to dorsal regions of the ectoderm (Figure 2A arrowhead; Figure 6A arrow), they are induced by signals originating from the ectoderm to express Eve within 10 clusters of three cells each, spanning the trunk of the embryo (Figure 6, B and C). These differentiation cues include FGF, Wg, and Dpp—all of which have the ability to cooperate with HSPGs in the context of receptor activation (Lin 2004). In both trol and sdc mutant embryos, mesoderm cells reach the dorsal ectoderm as a result of their migration despite their nonmonolayered spreading (Figure 4H and Figure 5F). We examined if Trol is required for the subsequent patterning of dorsal somatic lineages, but no measurable defect in Eve-specification was observed in embryos derived from trol germline clones (Figure 6D). Previous studies, however, have shown that sdc zygotic mutants, in contrast, do exhibit defects in Eve induction and linked this to effects on FGF signaling through genetic interaction assay (Knox et al. 2011). Likewise, we found that in embryos obtained from sdc germline clone, a significant reduction of Eve+ cells was observed (Figure 6F). These results reinforce the view that Sdc is required to support the role of FGF in differentiation of dorsal somatic mesoderm lineages. Overexpression of either Trol or Sdc in the ectoderm results in increased Eve+ cell number. However, overexpressing Sdc had a stronger phenotype than Trol with multiple clusters containing five or more cells (compare Figure 6E with Figure 6G). Although Trol is not required to support differentiation of dorsal somatic lineages, it can potentially substitute for Sdc when ectopically expressed (compare Figure 6D with Figure 6E).


Ectopic expression screen identifies genes affecting Drosophila mesoderm development including the HSPG Trol.

Trisnadi N, Stathopoulos A - G3 (Bethesda) (2014)

Embryos obtained from sdc, but not trol, mutant germline clones exhibit defects in Eve specification. (A) Schematic cross-section of the ventral half of an embryo at stage 11 during Eve specification. Mesoderm cells that reach the dorsal regions of the embryo (arrows) are able to receive signals, including FGF, from the ectoderm (dark orange) and undergo cell differentiation (dark blue). (B–G) Dorsal somatic mesoderm cell differentiation at stage 11 embryos is marked by Eve expression. (B) Wild-type whole embryo stained using anti-Eve antibody includes box showing region of magnification in subsequent panels. (C) Wild-type embryos have 10 clusters of three Eve+ cells. (D) trol germline clones show a normal number of Eve+ cells. (E) Embryos overexpressing trol in the ectoderm occasionally have clusters with four Eve+ cells, as indicated by the arrow. (F) sdc germline clones are missing clusters (arrows) and/or have reduced number of Eve+ cells within a cluster (arrowhead). (G) Embryos with sdc overexpressed in the ectoderm have multiple clusters with five or more Eve+ cells (arrows).
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fig6: Embryos obtained from sdc, but not trol, mutant germline clones exhibit defects in Eve specification. (A) Schematic cross-section of the ventral half of an embryo at stage 11 during Eve specification. Mesoderm cells that reach the dorsal regions of the embryo (arrows) are able to receive signals, including FGF, from the ectoderm (dark orange) and undergo cell differentiation (dark blue). (B–G) Dorsal somatic mesoderm cell differentiation at stage 11 embryos is marked by Eve expression. (B) Wild-type whole embryo stained using anti-Eve antibody includes box showing region of magnification in subsequent panels. (C) Wild-type embryos have 10 clusters of three Eve+ cells. (D) trol germline clones show a normal number of Eve+ cells. (E) Embryos overexpressing trol in the ectoderm occasionally have clusters with four Eve+ cells, as indicated by the arrow. (F) sdc germline clones are missing clusters (arrows) and/or have reduced number of Eve+ cells within a cluster (arrowhead). (G) Embryos with sdc overexpressed in the ectoderm have multiple clusters with five or more Eve+ cells (arrows).
Mentions: Pericardial and dorsal somatic muscle cells derived from the dorsal mesoderm are known to express Even-skipped (Eve) (Frasch 1999) and require proper migration of the mesoderm at an earlier stage prior to their specification. Once mesoderm cells migrate to dorsal regions of the ectoderm (Figure 2A arrowhead; Figure 6A arrow), they are induced by signals originating from the ectoderm to express Eve within 10 clusters of three cells each, spanning the trunk of the embryo (Figure 6, B and C). These differentiation cues include FGF, Wg, and Dpp—all of which have the ability to cooperate with HSPGs in the context of receptor activation (Lin 2004). In both trol and sdc mutant embryos, mesoderm cells reach the dorsal ectoderm as a result of their migration despite their nonmonolayered spreading (Figure 4H and Figure 5F). We examined if Trol is required for the subsequent patterning of dorsal somatic lineages, but no measurable defect in Eve-specification was observed in embryos derived from trol germline clones (Figure 6D). Previous studies, however, have shown that sdc zygotic mutants, in contrast, do exhibit defects in Eve induction and linked this to effects on FGF signaling through genetic interaction assay (Knox et al. 2011). Likewise, we found that in embryos obtained from sdc germline clone, a significant reduction of Eve+ cells was observed (Figure 6F). These results reinforce the view that Sdc is required to support the role of FGF in differentiation of dorsal somatic mesoderm lineages. Overexpression of either Trol or Sdc in the ectoderm results in increased Eve+ cell number. However, overexpressing Sdc had a stronger phenotype than Trol with multiple clusters containing five or more cells (compare Figure 6E with Figure 6G). Although Trol is not required to support differentiation of dorsal somatic lineages, it can potentially substitute for Sdc when ectopically expressed (compare Figure 6D with Figure 6E).

Bottom Line: These include the FGF ligand Pyramus, α-integrins, E-cadherin, Cueball, EGFR, JAK/STAT signaling components, as well as the heparan sulfate proteoglycan (HSPG) Terribly reduced optic lobes (Trol).Our data support the view that both HSPGs function to support FGF-dependent processes in the early embryo as they share phenotypes with FGF mutants: Trol in terms of effects on mesoderm migration and caudal visceral mesoderm (CVM) migration and Sdc in terms of dorsal mesoderm specification.The differential roles uncovered for these two HSPGs suggest that HSPG cofactor choice may modify FGF-signaling outputs.

View Article: PubMed Central - PubMed

Affiliation: Division of Biology and Biological Engineering, California Institute of Technology, 1200 East California Boulevard, MC 114-96, Pasadena, California 91125.

Show MeSH
Related in: MedlinePlus