Abstract：
Thioredoxin (Trx) is a widespread protein regulator of redox reactions in all organisms. It operates together with NADPH and thioredoxin reductase as a general protein disulfide catalytic system. Recently, Trx has been found to be related to the process by which apicomplexan protozoa invade host cells. In this study, Eimeria tenella thioredoxin (EtTrx1) was identified and its gene structural features, expression levels at different developmental stages, localization in sporozoites, roles in adhesion and invasion, and immunogenicity were investigated. Sequence analysis indicated that EtTrx1 contains a Trx domain with a WCGPC motif in 29-33 aa and a typical Trx fold, and belongs to thioredoxin family. EtTrx1 was detected on the surface of sporozoites using anti-EtTrx1 polyclonal antibodies under non-permeabilized conditions by indirect immunofluorescence assay (IFA) and also in a secretion form. EtTrx1 protein was highly transcribed and expressed in merozoites and sporozoites by quantitative PCR and western blot. The attachment assay showed that the adherence rates of yeast cells expressing EtTrx1 on the surface to host cells were 3.1-fold higher than those of the blank control. Specific anti-EtTrx1 antibodies inhibited the invasion of sporozoites into DF-1 cells. The highest inhibition rate was up to 36.75% compared to the control group. Immunization with recombinant EtTrx1 peptides also showed significant protection against lethal infections in chickens. It could offer moderate protective efficacy (Anticoccidial Index [ACI]: 163.70), induce humoral responses, and be an effective candidate for the development of new vaccines.
摘要：
硫氧还蛋白（Trx）是所有生物体中氧化还原反应的广泛蛋白质调节剂。它与NADPH和硫氧还蛋白还原酶一起作为通用的蛋白质二硫化物催化系统。最近,已发现Trx与顶丛原生动物侵入宿主细胞的过程有关。在这项研究中,硫氧还蛋白(EtTrx1)的鉴定及其基因结构特征,不同发育阶段的表达水平，在子孢子中定位，在粘附和侵入中的作用，和免疫原性进行了研究。序列分析表明，EtTrx1包含一个Trx结构域，其WCGPC基序在29-33个氨基酸和一个典型的Trx折叠，属于硫氧还蛋白家族。在非透化条件下，通过间接免疫荧光测定（IFA）使用抗EtTrx1多克隆抗体在子孢子表面上检测到EtTrx1，也以分泌形式检测到。通过定量PCR和蛋白质印迹，EtTrx1蛋白在裂殖子和子孢子中高度转录和表达。附着分析表明，在表面表达EtTrx1的酵母细胞对宿主细胞的粘附率是空白对照的3.1倍。特异性抗EtTrx1抗体抑制子孢子侵入DF-1细胞。与对照组相比，最高抑制率高达36.75％。用重组EtTrx1肽的免疫也显示了对鸡的致死性感染的显著保护。它可以提供适度的保护功效（抗球虫指数[ACI]：163.70），诱导体液反应，并成为开发新疫苗的有效候选者。
