Study on 14-3-3 related proteins and signal transduction for control of African tyrpanosomiasis.

14-3-3蛋白富含中枢神经系统，普遍存在于动植物真核生物中。目前已有不少蛋白与14-3-3结合，推测该蛋白在细胞周期、细胞凋亡等信号转导途径中具有重要作用。我们对14-3例布氏锥虫（Tb）进行了检查，并注意到以下几点：1) 14-3-3 2个同型（I、II）基因均可克隆。结果产生了重组蛋白和针对它们的敏感单特异性大鼠抗体。2)虽然14-3-3特有的基序是保守的，但锥虫的14-3-3特征与哺乳动物有很大的不同。3)我们开发了一种名为pQuadra （pQ）的有效载体构建，在诱导前无泄漏地进行dsRNAi。4)在14-3-3应用pQ对原环型（pr）进行了dsRNAi检测。在形态学上，pr损伤为奇异的多鞭毛状，动体定向或细胞核传递障碍。pr的生长受到抑制。5) pQ对dsRNAi诱导后的血流形态（bs）的影响比pr更早出现，其生长完全受到抑制。6)bs的形态变化与添加磷酸蛋白（pp）抑制剂冈田酸对bs培养的影响非常相似。根据14-3-3与pp的关系分析，我们非常接近非洲锥虫病的治疗靶点。

Protein 14-3-3 was described to be rich in CNS and ubiquitously found in eukaryotic organisms of plants and animals. Quite a many proteins have been reported to bind to 14-3-3 then it is estimated the protein has important roles in the cell cycle, apoptosis and other signal transduction paths.We examined 14-3-3 of Trypanosoma b.brucei (Tb) and remark the followings.1) The genes of 2 isotypes (I, II) of 14-3-3 could be cloned. The result produced the recombinant proteins and sensitive mono-specific rat antibodies against them.2) Though 14-3-3 distinctive motifs are conserved, the characteristics of trypanosome 14-3-3 are quite different form those of mammals.3) We developed solely effective vector construct named pQuadra (pQ) to perform dsRNAi without leakage before induction.4) Procyclic forms (pr) were examined by dsRNAi on 14-3-3 applying pQ. Morphologically pr was damaged to bizarre multiflagellated form and disoriented kinetoplsts or nuclei delivery. The growth of pr was suppressed.5)On blood stream forms (bs) after induction of dsRNAi with pQ more drastic effect appeared earlier than pr and the growth was entirely suppressed6) The morphological change of bs was very similar to the effect of okadaic acid addition to culture of bs, which is protein phosphates (pp) inhibitor. Analysis based on relation between 14-3-3 and pp we came very close to the therapeutic target of African trypanosomiasis.