Department of Medical Environmental Biology, Chung-Ang University College of Medicine, Dongjak-gu, Seoul, Republic of Korea.
Department of Pathology, Chung-Ang Univesity College of Medicine, Dongjak-gu, Seoul, Republic of Korea.
PLoS One. 2014 Jan 16;9(1):e85577. doi: 10.1371/journal.pone.0085577. eCollection 2014.
Clonorchis sinensis causes clonorchiasis, a potentially serious disease. Growth factor receptor-bound protein 2 (Grb2) is a cytosolic protein conserved among animals and plays roles in cellular functions such as meiosis, organogenesis and energy metabolism. In the present study, we report first molecular characters of growth factor receptor bound-protein (CsGrb2) from C. sinensis as counter part of Grb2 from animals and its possible functions in development and organogenesis of C. sinensis.
METHODOLOGY/PRINCIPAL FINDINGS: A CsGrb2 cDNA clone retrieved from the C. sinensis transcriptome encoded a polypeptide with a SH3-SH2-SH3 structure. Recombinant CsGrb2 was bacterially produced and purified to homogeneity. Native CsGrb2 with estimated molecular weight was identified from C. sinensis adult extract by western blotting using a mouse immune serum to recombinant CsGrb2. CsGrb2 transcripts was more abundant in the metacercariae than in the adults. Immunohistochemical staining showed that CsGrb2 was localized to the suckers, mesenchymal tissues, sperms in seminal receptacle and ovary in the adults, and abundantly expressed in most organs of the metacercariae. Recombinant CsGrb2 was evaluated to be little useful as a serodiagnostic reagent for C. sinesis human infections.
Grb2 protein found in C. sinensis was conserved among animals and suggested to play a role in the organogenesis, energy metabolism and mitotic spermatogenesis of C. sinensis. These findings from C. sinensis provide wider understanding on diverse function of Grb2 in lower animals such as platyhelminths.
华支睾吸虫可引起华支睾吸虫病，是一种潜在的严重疾病。生长因子受体结合蛋白 2（Grb2）是一种在动物中保守的细胞质蛋白，在细胞功能中发挥作用，如减数分裂、器官发生和能量代谢。本研究首次报道了华支睾吸虫的生长因子受体结合蛋白（CsGrb2）作为动物 Grb2 的对应物的分子特征，及其在华支睾吸虫发育和器官发生中的可能功能。
方法/主要发现：从华支睾吸虫转录组中获得的 CsGrb2 cDNA 克隆编码了一个具有 SH3-SH2-SH3 结构的多肽。重组 CsGrb2 经细菌表达和纯化，达到均一性。用抗重组 CsGrb2 的小鼠免疫血清进行 Western blot 分析，从华支睾吸虫成虫提取物中鉴定出天然 CsGrb2，其估计分子量为。CsGrb2 转录物在囊蚴中比成虫中更为丰富。免疫组织化学染色显示，CsGrb2 定位于成虫的吸盘、间质组织、精囊中的精子和卵巢中，在囊蚴的大多数器官中均大量表达。重组 CsGrb2 作为华支睾吸虫人体感染的血清学诊断试剂几乎没有用处。
在华支睾吸虫中发现的 Grb2 蛋白在动物中是保守的，提示其在华支睾吸虫的器官发生、能量代谢和有丝分裂精子发生中发挥作用。这些来自华支睾吸虫的发现为 Grb2 在扁形动物等低等动物中的多种功能提供了更广泛的理解。
