- Title
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Retinoic acid catabolizing enzyme CYP26C1 is a genetic modifier in SHOX deficiency
- Authors
- Montalbano, A., Juergensen, L., Roeth, R., Weiss, B., Fukami, M., Fricke-Otto, S., Binder, G., Ogata, T., Decker, E., Nuernberg, G., Hassel, D., Rappold, G.A.
- Source
- Full text @ EMBO Mol. Med.
Pattern of defects in zebrafish embryos injected with anti‐shox morpholino. Wild‐type embryos injected with control MO or with shox MO. A.Lateral views of the embryos at 55 hours post‐fertilization (hpf). B. Dorsal view and magnification on the lateral view of the embryos. Dotted line, pectoral fins. shox morphants show smaller fins compared to controls (n = 30 embryos). C. Expression of col2a1 at 55 hpf was examined by in situ hybridization in wild‐type embryos injected with control MO and with shox MO. Arrow and dotted line indicate the pectoral fin. D. Pectoral fin area was measured by ImageJ (n = 30 embryos). |
Pattern of defects in zebrafish embryos injected with anti‐cyp26c1 morpholino. Wild‐type embryos injected with control MO or with cyp26c1 MO. A, B (A) Lateral views of the embryos at 55 hours post‐fertilization (hpf). (B) Dorsal view and magnification on the lateral view of the embryos. Dotted line, pectoral fins; *, otic vesicle. cyp26c1 morphants show smaller fins compared to controls (n = 30 embryos). C, D Expression of col2a1 (C) and shox (D) at 55 hpf was examined by in situ hybridization in embryos injected with control MO or with cyp26c1 MO. (C) Dorsal view and magnification on the pectoral fins of col2a1 expression. Arrow and dotted line indicate the pectoral fin. (D) Dorsal view and magnification on the pectoral fins of shox expression. Arrow and dotted line indicate the pectoral fin. E. Pectoral fin area was measured by ImageJ (n = 30 embryos). F. Cignal‐RARE system luciferase assay to test cyp26c1 MO knockdown effect on RA acid levels in zebrafish embryos (n = 5 replicates). In each replicate, 20–30 embryos per condition were assayed. RLU, relative light units. G. Relative shox mRNA expression normalized to reference genes eef1a and b‐actin in zebrafish embryos injected with control MO or cyp26c1 MO (n = 4). RNA was extracted from 10–15 injected embryos at 55 hpf. H. Relative shox mRNA expression normalized to reference genes eef1a and b‐actin in zebrafish embryos treated with 100 nM ATRA (n = 4). Embryos were collected at 24 hpf and treated with mock control or ATRA. RNA was extracted from 10–15 embryos after 6‐h treatment. |
Co‐injection of titrated subphenotypic doses of anti‐shox and anti‐cyp26c1 morpholinos impairs limb development Wild‐type embryos injected with control MO, subphenotypic doses of shox MO, cyp26c1 MO, or a combination of shox + cyp26c1 MOs. A. Dorsal views of the embryos at 55 hours post‐fertilization (hpf). Dotted line, pectoral fins. shox + cyp26c1 double morphants show smaller fins compared to control and single MOs (n = 30 embryos). B, C Dorsal view and magnification on the pectoral fins of col2a1 expression at 55 hpf. Arrow and dotted line indicate the pectoral fin. D. Pectoral fin area was measured by ImageJ (n = 30 embryos). Data are shown as means ± SD. ***P = 0.0001 (one‐way ANOVA with Bonferroni's multiple comparison test). E. Relative expression of shox mRNA normalized to reference genes eef1a and b‐actin in zebrafish embryos injected with control MO, shox MO, cyp26c1 MO, or shox + cyp26c1 MOs (n = 4). RNA was extracted from 10–15 injected embryos at 55 hpf. Data are shown as means ± SD. **P = 0.0048 (Kruskal–Wallis with Dunn's multiple comparison test). |
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