CYP306A1, a cytochrome P450 enzyme, is essential for ecdysteroid biosynthesis in the prothoracic glands of Bombyx and Drosophila
CYP306A1, a cytochrome P450 enzyme, is essential for ecdysteroid biosynthesis in the prothoracic glands of Bombyx and Drosophila
复制标题
DOI：
10.1074/jbc.m404514200
复制
发表时间：
2004-08-20
影响因子：
4.8
通讯作者：
Kataoka, H
中科院分区：
文献类型：
--
作者：
Niwa, R;Matsuda, T;Kataoka, H
蜕皮甾体介导昆虫的多种发育和生理事件。在昆虫胚后发育过程中，蜕皮激素由前胸腺合成。虽然许多研究已经揭示了蜕皮甾体生物合成酶的生化和生理特性，但这些酶的大部分分子身份尚未阐明。在这里，我们描述了一个未知的细胞色素P450基因，命名为Cyp306a1，这是必不可少的蜕皮甾体生物合成的PG的家蚕家蚕和果蝇黑腹果蝇。使用微阵列技术分析PG细胞中的基因表达谱在发育过程中，我们确定了两个PG特异性P450基因的时间表达模式与蜕皮激素滴度的变化在发展过程中。氨基酸序列分析表明，其中一个P450基因属于CYP306A1亚家族。果蝇Cyp306a1同源物的时空表达模式与果蝇Cyp306a1同源物的时空表达模式基本相同。我们还发现果蝇Cyp306a1在幻影（phm）突变体中被破坏，也被称为万圣节突变体。phm的形态缺陷和蜕皮激素诱导基因表达的降低表明该突变体不能产生高滴度的蜕皮激素。最后，我们证明了转染Cyp306a1的S2细胞通过碳25羟基化将酮二醇转化为酮三醇。这些结果有力地表明，CYP306A1作为一个碳25羟化酶的功能，并在昆虫发育过程中蜕皮类固醇的生物合成中发挥重要作用。
Ecdysteroids mediate a wide variety of developmental and physiological events in insects. In the postembryonic development of insects, ecdysone is synthesized in the prothoracic gland (PG). Although many studies have revealed the biochemical and physiological properties of the enzymes for ecdysteroid biosynthesis, most of the molecular identities of these enzymes have not been elucidated. Here we describe an uncharacterized cytochrome P450 gene, designated Cyp306a1, that is essential for ecdysteroid biosynthesis in the PGs of the silkworm Bombyx mori and fruit fly Drosophila melanogaster. Using the microarray technique for analyzing gene expression profiles in PG cells during Bombyx development, we identified two PG-specific P450 genes whose temporal expression patterns are correlated with changes in ecdysteroid titer during development. Amino acid sequence analysis showed that one of the Bombyx P450 genes belongs to the CYP306A1 subfamily. The temporal and spatial expression pattern of the Drosophila Cyp306a1 homolog is essentially the same as that of Bombyx Cyp306a1. We also found that Drosophila Cyp306a1 is disrupted in the phantom (phm) mutant, known also as the Halloween mutant. The morphological defects and decreased expression of ecdysone-inducible genes in phm suggest that this mutant cannot produce a high titer of ecdysone. Finally we demonstrate that S2 cells transfected with Cyp306a1 convert ketodiol to ketotriol via carbon 25 hydroxylation. These results strongly suggest that CYP306A1 functions as a carbon 25 hydroxylase and has an essential role in ecdysteroid biosynthesis during insect development.