Visualization of an endogenous retinoic acid gradient across embryonic development

Satoshi Shimozono, Tadahiro Iimura, Tetsuya Kitaguchi, Shin Ichi Higashijima, Atsushi Miyawaki

Research output: Contribution to journalArticle

119 Citations (Scopus)

Abstract

In vertebrate development, the body plan is determined by primordial morphogen gradients that suffuse the embryo. Retinoic acid (RA) is an important morphogen involved in patterning the anterior-posterior axis of structures, including the hindbrain and paraxial mesoderm. RA diffuses over long distances, and its activity is spatially restricted by synthesizing and degrading enzymes. However, gradients of endogenous morphogens in live embryos have not been directly observed; indeed, their existence, distribution and requirement for correct patterning remain controversial. Here we report a family of genetically encoded indicators for RA that we have termed GEPRAs (genetically encoded probes for RA). Using the principle of fluorescence resonance energy transfer we engineered the ligand-binding domains of RA receptors to incorporate cyan-emitting and yellow-emitting fluorescent proteins as fluorescence resonance energy transfer donor and acceptor, respectively, for the reliable detection of ambient free RA. We created three GEPRAs with different affinities for RA, enabling the quantitative measurement of physiological RA concentrations. Live imaging of zebrafish embryos at the gastrula and somitogenesis stages revealed a linear concentration gradient of endogenous RA in a two-tailed source-sink arrangement across the embryo. Modelling of the observed linear RA gradient suggests that the rate of RA diffusion exceeds the spatiotemporal dynamics of embryogenesis, resulting in stability to perturbation. Furthermore, we used GEPRAs in combination with genetic and pharmacological perturbations to resolve competing hypotheses on the structure of the RA gradient during hindbrain formation and somitogenesis. Live imaging of endogenous concentration gradients across embryonic development will allow the precise assignment of molecular mechanisms to developmental dynamics and will accelerate the application of approaches based on morphogen gradients to tissue engineering and regenerative medicine.

Original languageEnglish
Pages (from-to)363-366
Number of pages4
JournalNature
Volume496
Issue number7445
DOIs
Publication statusPublished - 2013 Apr 18
Externally publishedYes

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Tretinoin
Embryonic Development
Embryonic Structures
Rhombencephalon
Fluorescence Resonance Energy Transfer
Gastrula
Retinoic Acid Receptors
Regenerative Medicine
Mesoderm
Zebrafish
Tissue Engineering
Vertebrates
Pharmacology
Ligands

ASJC Scopus subject areas

  • General

Cite this

Shimozono, S., Iimura, T., Kitaguchi, T., Higashijima, S. I., & Miyawaki, A. (2013). Visualization of an endogenous retinoic acid gradient across embryonic development. Nature, 496(7445), 363-366. https://doi.org/10.1038/nature12037

Visualization of an endogenous retinoic acid gradient across embryonic development. / Shimozono, Satoshi; Iimura, Tadahiro; Kitaguchi, Tetsuya; Higashijima, Shin Ichi; Miyawaki, Atsushi.

In: Nature, Vol. 496, No. 7445, 18.04.2013, p. 363-366.

Research output: Contribution to journalArticle

Shimozono, S, Iimura, T, Kitaguchi, T, Higashijima, SI & Miyawaki, A 2013, 'Visualization of an endogenous retinoic acid gradient across embryonic development', Nature, vol. 496, no. 7445, pp. 363-366. https://doi.org/10.1038/nature12037
Shimozono S, Iimura T, Kitaguchi T, Higashijima SI, Miyawaki A. Visualization of an endogenous retinoic acid gradient across embryonic development. Nature. 2013 Apr 18;496(7445):363-366. https://doi.org/10.1038/nature12037
Shimozono, Satoshi ; Iimura, Tadahiro ; Kitaguchi, Tetsuya ; Higashijima, Shin Ichi ; Miyawaki, Atsushi. / Visualization of an endogenous retinoic acid gradient across embryonic development. In: Nature. 2013 ; Vol. 496, No. 7445. pp. 363-366.
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