Molecular mechanisms which cause skeletal abnormalities in the jaw and gills of larvae

T. Suzuki and T. Kurokawa (Nutrition Div)

   To reveal what factors induce skeletal malformation in larvae, we analyzed the phenotypes of jaw anomalies experimentally by treatment with retinoic acid. Accumulation of these kinds of data will make it possible to predict the cause of skeletal abnormalities based on their phenotype.

  Larvae reproduced for aquaculture often have skeletal abnormalities in the jaw, vertebrae and fins, which reduce not only the survival ratio but also the commercial value. Even though skeletal abnormalities are attributable to several factors such as excess vitamin A and high temperature, there is little information on the mechanisms how these factors cause abnormalities. So, we have been trying to reveal the molecular mechanisms that cause skeletal abnormalities.
  In most teleosts including flounder, the jaw and gill skeletal structure develop in the early larval stage before first feeding, whereas the vertebral skeletal structure forms in the late larval stage. Therefore, the jaw and gill skeletal development is particularly susceptible to the effects of maternal factors, such as vitamin A and its active metabolite retinoic acid (RA) stocked in the yolk. RA is a key molecule which controls embryogenesis by initiating mRNA expression of transcriptional factors and signal proteins, such as Hox and sonic hedgehog (shh) which regulate the differentiation of skeletal precursor cells. In addition, excess RA is known to exert teratogenic effects on skeleton. To reveal the teratogenic effect of RA on the jaw and gill skeletal structure, we examined whether RA affects Hox and shh expressions and what kind of abnormality is induced in the jaw and gill by RA. When gastrulating embryos were incubated with RA, the anterior border of the Hoxd-4 expression domain at both central nervous system and pharyngeal area shifted anteriorly (Fig. 1A, B). In such larvae, severe malformation such as fusion of skeletal elements occurred in the lower jaw (Fig. 2A, B). When flounder were exposed to RA in the late embryonic stage, the shh expression in the jaw and gills was severely suppressed. The jaw and gill skeletal structure grew downwards in larvae of which the shh expression was affected by RA. Thus, different phenotypes of skeletal abnormalities are caused by RA at each developmental stage. We are further focusing on the mechanisms which cause malformation in the vertebrae and fins.
1) Suzuki, T., Oohara, I., Kurokawa, T. (1999) Retinoic acid given at late embryonic stage suppresses sonic hedgehog and Hoxd-4 expression in the pharyngeal area and induces skeletal malformation in flounder (Paralichthys olivaceus) embryos. Develop. Growth Differ. 41, 143-152.
2) Suzuki, T., Srivastava, A. S., Kurokawa T. (2000) Experimental induction of jaw, gill and pectoral fin malformations in Japanese flounder, Paralichthys olivaceus, larvae. Aquaculture, 185, 175-187.
3) Suzuki, T., Kurokawa T., Srivastava, A. S. (2001) Induction of bent cartilaginous skeletons and undulating notochord in flounder embryos by disulfiram and a, a-dipyridyl. Zool. Sci., 18, 345-351.

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