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Control of brain patterning by Engrailed paracrine transfer: a new function of the Pbx interaction domain.

Rampon C, Gauron C, Lin T, Meda F, Dupont E, Cosson A, Ipendey E, Frerot A, Aujard I, Le Saux T, Bensimon D, Jullien L, Volovitch M, Vriz S, Joliot A - Development (2015)

Bottom Line: In light of recent reports on the paracrine activity of homeoproteins, including Engrailed, we asked whether Engrailed intercellular transfer was also involved in brain patterning and boundary formation.Both zebrafish Eng2a and Eng2b are competent for intercellular transfer in vivo, but only extracellular endogenous Eng2b, and not Eng2a, participates in DMB positioning.In addition, disruption of the Pbx-interacting motif in Engrailed, known to strongly reduce the gain-of-function phenotype, also downregulates Engrailed transfer, thus revealing an unsuspected participation of the Pbx interaction domain in this pathway.

View Article: PubMed Central - PubMed

Affiliation: Université Paris Diderot, Sorbonne Paris Cité, Paris 75205, Cedex 13, France Center for Interdisciplinary Research in Biology (CIRB) - CNRS UMR 7241, INSERM U1050, Labex MemoLife, PSL Research University, Collège de France, Paris F-75005, France.

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Relative size of the anterior pax6-positive domain and pax6-negative domain. Sizes of the anterior pax6 expression domain and of the pax6-negative domain were measured on flat-mount embryos stained for pax6 expression by in situ hybridisation at 1 dpf. Green and blue squares, En2 mRNA was injected at the one-cell stage and CYC was added (blue) or not (green) at 50% epiboly. Green triangles (anti-myc control) and red squares (anti-Engrailed), antibodies were injected at the blastula stage to block Engrailed paracrine activity.
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DEV114181F5: Relative size of the anterior pax6-positive domain and pax6-negative domain. Sizes of the anterior pax6 expression domain and of the pax6-negative domain were measured on flat-mount embryos stained for pax6 expression by in situ hybridisation at 1 dpf. Green and blue squares, En2 mRNA was injected at the one-cell stage and CYC was added (blue) or not (green) at 50% epiboly. Green triangles (anti-myc control) and red squares (anti-Engrailed), antibodies were injected at the blastula stage to block Engrailed paracrine activity.

Mentions: It has been proposed previously that Engrailed gain of function induced a transformation of forebrain tissue identity towards a mesencephalic fate rather than an intrinsic shortening of the diencephalon (Scholpp et al., 2003). We indeed observed that activation of En2 at the appropriate time did not affect apoptosis during development (supplementary material Fig. S7), supporting an identity switch between the two adjacent structures. To confirm this hypothesis, we compared the relative size of the anterior pax6-positive (diencephalic) and adjacent pax6-negative (mesencephalic) domains in all control and treated animals (injected with 4D9 antibody) previously analysed in Fig. 4. Mesencephalon shortening induced by blocking En2 transfer with 4D9 (red squares) was correlated with a corresponding enlargement of the diencephalon (Fig. 5). To our surprise, the reverse effect in gain-of-function experiments was not as clear, the size of the mesencephalon being not statistically affected by En2 ectopic expression, despite the significant shortening of the diencephalon (blue squares) (Fig. 5).Fig. 5.


Control of brain patterning by Engrailed paracrine transfer: a new function of the Pbx interaction domain.

Rampon C, Gauron C, Lin T, Meda F, Dupont E, Cosson A, Ipendey E, Frerot A, Aujard I, Le Saux T, Bensimon D, Jullien L, Volovitch M, Vriz S, Joliot A - Development (2015)

Relative size of the anterior pax6-positive domain and pax6-negative domain. Sizes of the anterior pax6 expression domain and of the pax6-negative domain were measured on flat-mount embryos stained for pax6 expression by in situ hybridisation at 1 dpf. Green and blue squares, En2 mRNA was injected at the one-cell stage and CYC was added (blue) or not (green) at 50% epiboly. Green triangles (anti-myc control) and red squares (anti-Engrailed), antibodies were injected at the blastula stage to block Engrailed paracrine activity.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4440920&req=5

DEV114181F5: Relative size of the anterior pax6-positive domain and pax6-negative domain. Sizes of the anterior pax6 expression domain and of the pax6-negative domain were measured on flat-mount embryos stained for pax6 expression by in situ hybridisation at 1 dpf. Green and blue squares, En2 mRNA was injected at the one-cell stage and CYC was added (blue) or not (green) at 50% epiboly. Green triangles (anti-myc control) and red squares (anti-Engrailed), antibodies were injected at the blastula stage to block Engrailed paracrine activity.
Mentions: It has been proposed previously that Engrailed gain of function induced a transformation of forebrain tissue identity towards a mesencephalic fate rather than an intrinsic shortening of the diencephalon (Scholpp et al., 2003). We indeed observed that activation of En2 at the appropriate time did not affect apoptosis during development (supplementary material Fig. S7), supporting an identity switch between the two adjacent structures. To confirm this hypothesis, we compared the relative size of the anterior pax6-positive (diencephalic) and adjacent pax6-negative (mesencephalic) domains in all control and treated animals (injected with 4D9 antibody) previously analysed in Fig. 4. Mesencephalon shortening induced by blocking En2 transfer with 4D9 (red squares) was correlated with a corresponding enlargement of the diencephalon (Fig. 5). To our surprise, the reverse effect in gain-of-function experiments was not as clear, the size of the mesencephalon being not statistically affected by En2 ectopic expression, despite the significant shortening of the diencephalon (blue squares) (Fig. 5).Fig. 5.

Bottom Line: In light of recent reports on the paracrine activity of homeoproteins, including Engrailed, we asked whether Engrailed intercellular transfer was also involved in brain patterning and boundary formation.Both zebrafish Eng2a and Eng2b are competent for intercellular transfer in vivo, but only extracellular endogenous Eng2b, and not Eng2a, participates in DMB positioning.In addition, disruption of the Pbx-interacting motif in Engrailed, known to strongly reduce the gain-of-function phenotype, also downregulates Engrailed transfer, thus revealing an unsuspected participation of the Pbx interaction domain in this pathway.

View Article: PubMed Central - PubMed

Affiliation: Université Paris Diderot, Sorbonne Paris Cité, Paris 75205, Cedex 13, France Center for Interdisciplinary Research in Biology (CIRB) - CNRS UMR 7241, INSERM U1050, Labex MemoLife, PSL Research University, Collège de France, Paris F-75005, France.

Show MeSH