Essential Role of Duox in Stabilization of Drosophila Wing
Essential Role of Duox in Stabilization of Drosophila Wing
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DOI：
10.1074/jbc.m111.263178
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发表时间：
2011-09-23
影响因子：
4.8
通讯作者：
Kamei, Kaeko
中科院分区：
文献类型：
--
作者：
Nguyen Thi Tu Anh;Nishitani, Maiko;Kamei, Kaeko
NADPH氧化酶产生活性氧（ROS）。黑胃果蝇有两个NADPH氧化酶的同源物，dNox和dDuox，其在体内的功能尚不清楚。为了阐明这些功能，研究人员使用了两种独立的转基因蝇系，表达针对dDuox mRNA不同部分的dsRNA。在en-GAL4> UAS-dDuoxIR（976-1145）和en-GAL4> UAS-dDuoxIR（370-518）两种果蝇中，dDuox在翅盘后部区域被选择性地敲除，成年后的翅膀后腔室变得更苍白，更脆弱，与前腔相比翼静脉不清晰。对en-GAL4 b> UAS-dDuoxIR（976-1145）成虫翅膀进行荧光染色，发现后室ROS浓度明显低于前室。此外，在这些果蝇中，用抗裂解caspase-3抗体免疫染色检测到的翅膀成像盘后室的凋亡细胞数量多于前室。酪氨酸羟化酶或多巴脱羧酶的分别敲低显示后室翼叶颜色变浅，与敲低dduox基因的表型相似。与此同时，对成年果蝇翅膀中儿茶酚和二酪氨酸成分的分析证明，dDuox通过酪氨酸交联稳定翅膀角质层结构，并可能通过ROS产生硬化和黑化过程发挥重要作用。这些敲除dDuox的果蝇系将为进一步研究果蝇发育过程中dDuox的功能提供有用的工具。
NADPH oxidase produces reactive oxygen species (ROS). Drosophila melanogaster has two homologs of NADPH oxidase, dNox and dDuox, with functions that remain unclear in vivo. To clarify these functions, two independent transgenic fly lines expressing dsRNA targeted for different portions of dDuox mRNA were used. In both flies, en-GAL4> UAS-dDuoxIR(976-1145) and en-GAL4> UAS-dDuoxIR(370-518), in which dDuox was knocked down selectively in the posterior area of the wing disc, the posterior compartment of the adult wings became paler and more fragile with wing veins that were indistinct by comparison with the anterior one. Fluorescence staining of the en-GAL4> UAS-dDuoxIR(976-1145) adult wings revealed that the ROS concentration in the posterior compartment was significantly lower than that in the anterior compartment. Moreover, in these flies, the posterior compartment of the wing imaginal disc showed a greater number of apoptotic cells detected by immunostaining with anti-cleaved caspase-3 antibody than those in the anterior compartment. Respective knockdown of tyrosine hydroxylase or dopa-decarboxylase showed paler wing blades in the posterior compartment similar to the phenotype of dDuox-knockdown files. Along with this observation, analysis of the catecholic and dityrosine components in the wings of adult flies proved that dDuox plays important roles in the stabilization of the cuticle structure of the wings via tyrosine cross-linking, the sclerotization and melanization processes possibly through ROS production. These dDuox-knockdown fly lines would be useful tools for further studying dDuox functions during the development of Drosophila.