Identificaton of novel Cystoisospora suis enolase 2 interaction proteins in IPEC-J2 cells using Co-IP/LC-MS.

IF 2 4区医学 Q3 PARASITOLOGY

Folia Parasitologica Pub Date : 2026-08-31 DOI:10.14411/fp.2026.009

Hung-Chuan Chiu, Yuhao Ao, Yaoxue Tang

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引用次数: 0

Abstract

Enolase 2 (ENO2) is a crucial an important glycolytic enzyme involved in glycolysis and is also recognised as a multifunctional protein. However, its interacting proteins in intestinal porcine epithelial cells (IPEC-J2 cells) remain unclear. This study investigates the interacting proteins of Cystoisospora suis (Biester et Murray, 1934) ENO2 (CsENO2) in IPEC-J2 cells. According to our previous research, CsENO2 is highly expressed in sporulated oocysts. Co-immunoprecipitation/liquid chromatography-mass spectrometry (Co-IP/LC-MS) was conducted using the pcDNA3.1-Flag vector carrying the enolase 2 gene, which was transfected into IPEC-J2 cells to co-immunoprecipitate with cellular proteins. The Co-IP/LC-MS-based protein sequencing identified three cellular proteins: NME2, RPS3, and Vimentin. Subsequently, cDNAs encoding the complete coding sequences of these proteins were cloned into the pGEX-4T-1 expression vector. Immunofluorescence assays revealed the localisation of CsENO2 in sporulated oocysts and sporozoites. Most CsENO2 protein was found in the cytoplasm of sporulated oocysts. In sporozoites, CsENO2 protein was primarily distributed around and outside the nucleus. This study aims to identify CsENO2-interacting proteins in IPEC-J2 cells and determine the localisation of CsENO2 within sporulated oocysts and sporozoites.

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用Co-IP/LC-MS鉴定IPEC-J2细胞中新型猪囊异孢子虫烯醇化酶2相互作用蛋白。

烯醇化酶2 （ENO2）是参与糖酵解的一种至关重要的糖酵解酶，也是公认的多功能蛋白。然而，其在猪肠上皮细胞（IPEC-J2细胞）中的相互作用蛋白尚不清楚。本研究研究了猪囊异孢子虫（Cystoisospora suis, Biester et Murray, 1934） ENO2 （CsENO2）在IPEC-J2细胞中的相互作用蛋白。根据我们之前的研究，CsENO2在孢子卵囊中高度表达。采用携带烯醇化酶2基因的pcDNA3.1-Flag载体进行共免疫沉淀/液相色谱-质谱（Co-IP/LC-MS），将其转染IPEC-J2细胞，与细胞蛋白共免疫沉淀。基于Co-IP/ lc - ms的蛋白测序鉴定出三种细胞蛋白：NME2， RPS3和Vimentin。随后，将编码这些蛋白完整编码序列的cdna克隆到pGEX-4T-1表达载体中。免疫荧光分析显示CsENO2在孢子卵囊和孢子子中的定位。CsENO2蛋白主要存在于孢子卵囊细胞质中。在孢子体中，CsENO2蛋白主要分布在细胞核周围和细胞核外。本研究旨在鉴定IPEC-J2细胞中CsENO2相互作用蛋白，并确定CsENO2在孢子卵囊和孢子子内的定位。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

Folia Parasitologica 医学-寄生虫学

CiteScore

2.70

自引率

0.00%

发文量

审稿时长

>12 weeks

期刊介绍： FOLIA PARASITOLOGICA, issued in online versions, is an international journal that covers the whole field of general, systematic, ecological and experimental parasitology. It publishes original research papers, research notes and review articles. Contributions from all branches of animal parasitology, such as morphology, taxonomy, biology, biochemistry, physiology, immunology, molecular biology and evolution of parasites, and host-parasite relationships, are eligible. Novelty and importance in the international (not local or regional) context are required. New geographical records of parasites, records of new hosts, regional parasite and/or host surveys (if they constitute the principal substance of manuscript), local/regional prevalence surveys of diseases, local/regional studies on epidemiology of well known diseases and of parasite impact on human/animal health, case reports, routine clinical studies and testing of established diagnostic or treatment procedures, will not be considered. One species description will also not be considered unless they include more general information, such as new diagnostic characters, host-parasite associations, phylogenetic implications, etc. Manuscripts found suitable on submission will be reviewed by at least two reviewers.