Laboratoire de Physique Statistique, Ecole Normale Supérieure, UPMC Université Paris 06, Université Paris Diderot, CNRS UMR8550, 24 rue Lhomond, 75005 Paris, France.
Development. 2012 Sep;139(18):3355-62. doi: 10.1242/dev.077776. Epub 2012 Aug 8.
All-trans retinoic acid (RA) is a key player in many developmental pathways. Most methods used to study its effects in development involve continuous all-trans RA activation by incubation in a solution of all-trans RA or by implanting all-trans RA-soaked beads at desired locations in the embryo. Here we show that the UV-driven photo-isomerization of 13-cis RA to the trans-isomer (and vice versa) can be used to non-invasively and quantitatively control the concentration of all-trans RA in a developing embryo in time and space. This facilitates the global or local perturbation of developmental pathways with a pulse of all-trans RA of known concentration or its inactivation by UV illumination. In zebrafish embryos in which endogenous synthesis of all-trans RA is impaired, incubation for as little as 5 minutes in 1 nM all-trans RA (a pulse) or 5 nM 13-cis RA followed by 1-minute UV illumination is sufficient to rescue the development of the hindbrain if performed no later than bud stage. However, if subsequent to this all-trans RA pulse the embryo is illuminated (no later than bud stage) for 1 minute with UV light (to isomerize, i.e. deactivate, all-trans RA), the rescue of hindbrain development is impaired. This suggests that all-trans RA is sequestered in embryos that have been transiently exposed to it. Using 13-cis RA isomerization with UV light, we further show that local illumination at bud stage of the head region (but not the tail) is sufficient to rescue hindbrain formation in embryos whose all-trans RA synthetic pathway has been impaired.
全反式视黄酸（RA）是许多发育途径中的关键因子。大多数用于研究其在发育过程中作用的方法都涉及通过在全反式 RA 溶液中孵育或在胚胎中所需位置植入全反式 RA 浸泡珠来持续激活全反式 RA。在这里，我们展示了 13-顺式 RA 向反式异构体（反之亦然）的 UV 驱动光异构化可以用于非侵入性和定量地控制发育中的胚胎中全反式 RA 的浓度，从而在时间和空间上控制其浓度。这使得具有已知浓度的全反式 RA 的脉冲或通过 UV 照射使其失活的脉冲，能够全局或局部地干扰发育途径。在全反式 RA 内源性合成受损的斑马鱼胚胎中，在 1 nM 全反式 RA（脉冲）或 5 nM 13-顺式 RA 中孵育 5 分钟，然后进行 1 分钟的 UV 照射，足以在芽期之前挽救后脑的发育。然而，如果在此之后全反式 RA 脉冲，胚胎随后在芽期之前用紫外线照射 1 分钟（使其异构化，即失活，全反式 RA），则会损害后脑发育的挽救。这表明全反式 RA 被暂时暴露于其中的胚胎隔离。使用 13-顺式 RA 异构化和 UV 光，我们进一步表明，在头部区域的芽期（而不是尾部）进行局部照射足以挽救全反式 RA 合成途径受损的胚胎中的后脑形成。
