Characterization and Processing of Superoxide Dismutase-Fused Vitellogenin in the Diapause Embryo Formation: A Special Developmental Pathway in the Brine Shrimp, Artemia parthenogenetica
Characterization and Processing of Superoxide Dismutase-Fused Vitellogenin in the Diapause Embryo Formation: A Special Developmental Pathway in the Brine Shrimp, Artemia parthenogenetica
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滞育胚胎形成中超氧化物歧化酶融合卵黄蛋白原的表征和加工：丰年虾孤雌生殖丰年虫的特殊发育途径
DOI：
10.1095/biolreprod.110.090340
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发表时间：
2011-07-01
影响因子：
3.6
通讯作者：
Yang, Wei-Jun
中科院分区：
文献类型：
--
作者：
Chen, Su;Chen, Dian-Fu;Yang, Wei-Jun
为了承受环境压力，卤虫通过卵生途径释放滞育包囊，而不是通过卵胎生途径产生游泳的无节幼体。滞育的胚胎被包裹在这样的囊肿中，可以存活很多年。卵黄蛋白原 (Vtg) 是卵黄蛋白（主要卵黄蛋白）的前体，对于胚胎发育至关重要。本研究比较了卵生和卵胎生之间的卵黄发生，这两种生殖模式发生在孤雌生殖中。通过N端氨基酸序列分析、PCR扩增以及cDNA文库构建和筛选，克隆了Vtg基因，该基因全长6778bp，开放阅读框6657bp，编码2219个氨基酸。从推导的一级结构来看，卤虫卵黄蛋白原 (ArVtg) 具有 6 个共有切割位点 R-X-X-R 的拷贝，并且在 N 末端含有一个超氧化物歧化酶 (SOD) 样结构域。对于甲壳类动物 Vtg 蛋白来说，这是一个不寻常的发现，此前仅在一种甲壳类动物大型水蚤中报道过。使用Northern印迹分析和原位杂交，在位于头胸部的结缔组织中卵黄发生的早期阶段观察到ArVtg基因表达，并且在卵巢中存在微量表达。蛋白质印迹分析和几个 N 末端序列显示，ArVtg 在每个共有切割位点被切割，并且在卵巢成熟的翻译后加工过程中形成了 10 多个亚基。其中，只有含有 SOD 的亚基（类似于 90 和 60 kDa）在卵生和卵胎生途径之间显示出不同的特征。这表明这些高浓度成分对于包膜滞育胚胎在长期细胞周期停滞期间具有重要功能，但迄今为止仍不清楚。
To withstand environmental stress, Artemia release diapause cysts via an oviparous pathway instead of producing swimming nauplius larvae by the ovoviviparous pathway. Encased in such a cyst, the embryos at diapause can survive for many years. Vitellogenin (Vtg), the precursor of vitellins, the main yolk proteins, is crucial for embryonic development. This study compares vitellogenesis between oviparity and ovoviviparity, the two reproductive modes occurring in A. parthenogenetica. A Vtg gene was cloned, based on N-terminal amino acid sequence analysis, PCR amplification, and cDNA library construction and screening, and was found to consist of 6778 bp with a 6657 bp open reading frame encoding 2219 amino acids. From the deduced primary structure, Artemia vitellogenin (ArVtg) was found to possess six copies of the consensus cleavage site, R-X-X-R, and to contain a superoxide dismutase (SOD)-like domain at the N-terminus. This is an unusual finding for crustacean Vtg proteins, having been reported only in one previous crustacean, Daphnia magna. Using Northern blot analysis and in situ hybridization, ArVtg gene expression was observed at early stages of vitellogenesis in the connective tissue located in the cephalothorax, with trace expression in the ovary. Western blot analysis and several N-terminal sequences revealed that ArVtg was cleaved at each consensus cleavage site and that more than 10 subunits were formed during posttranslational processing in ovarian maturation. Of these, only the SOD-containing subunits (similar to 90 and 60 kDa) showed different profiles between the oviparous and ovoviviparous pathways. This suggests that these high concentration components have an important function for the encysted diapaused embryos during long-term cell-cycle arrest, which has remained unknown up until now.