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Def6 is required for convergent extension movements during zebrafish gastrulation downstream of Wnt5b signaling.

Goudevenou K, Martin P, Yeh YJ, Jones P, Sablitzky F - PLoS ONE (2011)

Bottom Line: Wnt signaling results in downstream activation of Rho GTPases that in turn regulate actin cytoskeleton rearrangements essential for co-ordinated CE cell movement.Here we show that def6, a novel GEF, regulates CE cell movement during zebrafish gastrulation.In addition, by knocking down both def6 and Wnt11, we show that def6 synergises with the Wnt11 signaling pathway.

View Article: PubMed Central - PubMed

Affiliation: School of Biology, Queen's Medical Centre, The University of Nottingham, Nottingham, United Kingdom.

ABSTRACT
During gastrulation, convergent extension (CE) cell movements are regulated through the non-canonical Wnt signaling pathway. Wnt signaling results in downstream activation of Rho GTPases that in turn regulate actin cytoskeleton rearrangements essential for co-ordinated CE cell movement. Rho GTPases are bi-molecular switches that are inactive in their GDP-bound stage but can be activated to bind GTP through guanine nucleotide exchange factors (GEFs). Here we show that def6, a novel GEF, regulates CE cell movement during zebrafish gastrulation. Def6 morphants exhibit broadened and shortened body axis with normal cell fate specification, reminiscent of the zebrafish mutants silberblick and pipetail that lack Wnt11 or Wnt5b, respectively. Indeed, def6 morphants phenocopy Wnt5b mutants and ectopic overexpression of def6 essentially rescues Wnt5b morphants, indicating a novel role for def6 as a central GEF downstream of Wnt5b signaling. In addition, by knocking down both def6 and Wnt11, we show that def6 synergises with the Wnt11 signaling pathway.

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Zebrafish def6 is dynamically expressed during development.(A) Schematic representation of the def6 protein domain arrangement consisting of an N-terminal putative Ca2+-binding EF-hand domain followed by a def6/swap-70 homology (DSH), a pleckstrin-homology (PH) domain and a C-terminal Dbl-homology-like (DHL) domain. (B) Sequence identity and similarity of the various def6 domains between different species as indicated. (C-E) Def6 is ubiquitously expressed at the (C) 8-cell stage, (D) 40% epiboly and (E) 80% epiboly stages. (F) At 24hpf, def6 transcripts are found in the developing brain, somite boundaries and tail. (G) The expression pattern of def6 becomes more restricted at 3dpf where mRNA can be detected in the pharyngeal arches, medial and pectorial fins and anterior and posterior neuromasts of the lateral line.
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pone-0026548-g001: Zebrafish def6 is dynamically expressed during development.(A) Schematic representation of the def6 protein domain arrangement consisting of an N-terminal putative Ca2+-binding EF-hand domain followed by a def6/swap-70 homology (DSH), a pleckstrin-homology (PH) domain and a C-terminal Dbl-homology-like (DHL) domain. (B) Sequence identity and similarity of the various def6 domains between different species as indicated. (C-E) Def6 is ubiquitously expressed at the (C) 8-cell stage, (D) 40% epiboly and (E) 80% epiboly stages. (F) At 24hpf, def6 transcripts are found in the developing brain, somite boundaries and tail. (G) The expression pattern of def6 becomes more restricted at 3dpf where mRNA can be detected in the pharyngeal arches, medial and pectorial fins and anterior and posterior neuromasts of the lateral line.

Mentions: In order to address def6 function in zebrafish development, we first determined the zebrafish orthologue of def6. Database searches of the Ensembl genome database revealed that the zebrafish genome contains five genes related to def6 and its homologue swap-70 within a predicted family of proteins; ENS00000002981. Putative amino acid sequences of these genes were compared with sequences of mouse and human proteins to establish phylogenetic relationships. A neighbour-joining tree generated using Jalview showed the def6- and swap-70-related proteins clustered in three separate groups identifying the hypothetical protein encoded by the zgc:63721 gene as the closest zebrafish orthologue of human and mouse def6 (Shuen, et al., in preparation). N-terminal, PH and DH-like domains of zebrafish def6 (indicated in Figure 1A) exhibit 63%–72% identity and 78%–84% homology with human and mouse def6 (Figure 1B).


Def6 is required for convergent extension movements during zebrafish gastrulation downstream of Wnt5b signaling.

Goudevenou K, Martin P, Yeh YJ, Jones P, Sablitzky F - PLoS ONE (2011)

Zebrafish def6 is dynamically expressed during development.(A) Schematic representation of the def6 protein domain arrangement consisting of an N-terminal putative Ca2+-binding EF-hand domain followed by a def6/swap-70 homology (DSH), a pleckstrin-homology (PH) domain and a C-terminal Dbl-homology-like (DHL) domain. (B) Sequence identity and similarity of the various def6 domains between different species as indicated. (C-E) Def6 is ubiquitously expressed at the (C) 8-cell stage, (D) 40% epiboly and (E) 80% epiboly stages. (F) At 24hpf, def6 transcripts are found in the developing brain, somite boundaries and tail. (G) The expression pattern of def6 becomes more restricted at 3dpf where mRNA can be detected in the pharyngeal arches, medial and pectorial fins and anterior and posterior neuromasts of the lateral line.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3198796&req=5

pone-0026548-g001: Zebrafish def6 is dynamically expressed during development.(A) Schematic representation of the def6 protein domain arrangement consisting of an N-terminal putative Ca2+-binding EF-hand domain followed by a def6/swap-70 homology (DSH), a pleckstrin-homology (PH) domain and a C-terminal Dbl-homology-like (DHL) domain. (B) Sequence identity and similarity of the various def6 domains between different species as indicated. (C-E) Def6 is ubiquitously expressed at the (C) 8-cell stage, (D) 40% epiboly and (E) 80% epiboly stages. (F) At 24hpf, def6 transcripts are found in the developing brain, somite boundaries and tail. (G) The expression pattern of def6 becomes more restricted at 3dpf where mRNA can be detected in the pharyngeal arches, medial and pectorial fins and anterior and posterior neuromasts of the lateral line.
Mentions: In order to address def6 function in zebrafish development, we first determined the zebrafish orthologue of def6. Database searches of the Ensembl genome database revealed that the zebrafish genome contains five genes related to def6 and its homologue swap-70 within a predicted family of proteins; ENS00000002981. Putative amino acid sequences of these genes were compared with sequences of mouse and human proteins to establish phylogenetic relationships. A neighbour-joining tree generated using Jalview showed the def6- and swap-70-related proteins clustered in three separate groups identifying the hypothetical protein encoded by the zgc:63721 gene as the closest zebrafish orthologue of human and mouse def6 (Shuen, et al., in preparation). N-terminal, PH and DH-like domains of zebrafish def6 (indicated in Figure 1A) exhibit 63%–72% identity and 78%–84% homology with human and mouse def6 (Figure 1B).

Bottom Line: Wnt signaling results in downstream activation of Rho GTPases that in turn regulate actin cytoskeleton rearrangements essential for co-ordinated CE cell movement.Here we show that def6, a novel GEF, regulates CE cell movement during zebrafish gastrulation.In addition, by knocking down both def6 and Wnt11, we show that def6 synergises with the Wnt11 signaling pathway.

View Article: PubMed Central - PubMed

Affiliation: School of Biology, Queen's Medical Centre, The University of Nottingham, Nottingham, United Kingdom.

ABSTRACT
During gastrulation, convergent extension (CE) cell movements are regulated through the non-canonical Wnt signaling pathway. Wnt signaling results in downstream activation of Rho GTPases that in turn regulate actin cytoskeleton rearrangements essential for co-ordinated CE cell movement. Rho GTPases are bi-molecular switches that are inactive in their GDP-bound stage but can be activated to bind GTP through guanine nucleotide exchange factors (GEFs). Here we show that def6, a novel GEF, regulates CE cell movement during zebrafish gastrulation. Def6 morphants exhibit broadened and shortened body axis with normal cell fate specification, reminiscent of the zebrafish mutants silberblick and pipetail that lack Wnt11 or Wnt5b, respectively. Indeed, def6 morphants phenocopy Wnt5b mutants and ectopic overexpression of def6 essentially rescues Wnt5b morphants, indicating a novel role for def6 as a central GEF downstream of Wnt5b signaling. In addition, by knocking down both def6 and Wnt11, we show that def6 synergises with the Wnt11 signaling pathway.

Show MeSH
Related in: MedlinePlus