Characterization of the carotenoid-binding protein of the Y-gene dominant mutants of Bombyx mori.
Characterization of the carotenoid-binding protein of the Y-gene dominant mutants of Bombyx mori.
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DOI：
10.1016/j.jinsphys.2004.02.006
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发表时间：
2004-04
影响因子：
2.2
通讯作者：
K. Tsuchida;Z. Jouni;Jennifer S Gardetto;Yutaka Kobayashi;H. Tabunoki;M. Azuma;H. Sugiyama;N. Takada;H. Maekawa;Y. Banno;H. Fujii;H. Iwano;M. Wells
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文献类型：
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作者：
K. Tsuchida;Z. Jouni;Jennifer S Gardetto;Yutaka Kobayashi;H. Tabunoki;M. Azuma;H. Sugiyama;N. Takada;H. Maekawa;Y. Banno;H. Fujii;H. Iwano;M. Wells
类胡萝卜素在昆虫中发挥着重要而多样的作用。最近，我们从野生型家蚕中纯化并部分鉴定了一种类胡萝卜素结合蛋白（CBP）。在本报告中，我们利用免疫印迹、ELISA和免疫细胞化学进一步表征和定位了CBP在野生型和四种与类胡萝卜素运输相关的天然突变体的幼虫中肠和蚕丝腺中的表达。CBP在整个第5体育场均有表达，以丝腺第4 ~ 5天和中肠第3 ~ 5天表达量最高。免疫印迹分析显示沿中肠中部存在CBP。显微免疫细胞化学表明，33 kDa的CBP在中肠上皮柱状细胞的刷状边缘均匀表达，说明其在帮助吸收膳食类胡萝卜素方面的功能。同样，CBP在丝腺中部远端膜高度表达，表明其在脂蛋白吸收类胡萝卜素中起作用。当用兔抗CBP抗体孵育4个突变体的中间丝腺和中间肠时，只有y基因显性突变体的蛋白与抗体发生交叉反应，进一步强化了CBP是y基因依赖蛋白的假设。
Carotenoids play important and diverse roles in insects. Recently, we purified and partially characterized a carotenoid-binding protein (CBP) from the wild type of Bombyx mori. In this report, we utilized immunoblotting, ELISA and immunocytochemistry to further characterize and localize the expression of CBP in the larval midgut and silk gland obtained from the wild type and four naturally occurring mutants linked to carotenoids transport. CBP was expressed throughout the 5th stadium, with highest expressions on days 4–5 in the silk gland and days 3–5 in the midgut. Immunoblotting analyses demonstrated the presence of CBP along the middle part of the midgut. Microscopic immunocytochemistry demonstrated that the 33 kDa CBP was uniformly expressed along the brush border of columnar cells in the epithelium of the midgut typifying its function in aiding absorption of dietary carotenoids. Similarly, CBP was highly expressed along the distal membrane of the middle part of the silk gland demonstrating its function in uptake of carotenoids from lipophorin. When the middle silk glands and midguts of the four mutants were incubated with rabbit anti-CBP antibody, only proteins of the Y-gene dominant mutants cross reacted with the antibody further accentuating the hypothesis that the CBP is a Y-gene dependent protein.