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Identification of two novel host proteins interacting with Toxoplasma gondii 14-3-3 protein by yeast two-hybrid system

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Abstract

Toxoplasma gondii deploys many effector proteins in order to hijack and manipulate host cell signaling pathways, allowing parasite colonization, subversion of immune responses, and disease progression. T. gondii effector protein 14-3-3 (Tg14-3-3) promotes parasite dissemination inside the body, by enhancing the migratory ability of infected microglia and dendritic cells. Understanding both the mechanism of action and the host targets of Tg14-3-3 effector is important because of their importance to the parasite’s virulence. The aim of the present study was to explore the function of Tg14-3-3 by utilizing the yeast two-hybrid system (Y2HS) to identify novel Tg14-3-3 interactors/substrates in host cells. A human cDNA library was screened using Tg14-3-3 as the bait. Tg14-3-3 (RH strain, Type I) was cloned into the pGBKT7 vector and expressed in the Y2HGold yeast strain. The bait protein expression was validated by Western blotting analysis, auto-activation, and toxicity investigation compared with control (Y2HGold yeast strain transformed with empty pGBKT7 vector). Two positive Tg14-3-3 interactors identified by this screening, hCG1821272 and eIF5B (eukaryotic translation initiation factor 5B), were isolated and characterized. This approach made it possible to gain a better understanding of the function of Tg14-3-3 in regulating host proteins involved in key cellular processes, such as translational initiation and cell migration.

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Funding

This study was supported by the National Natural Science Foundation of China (Grant No. 31230073), the Fundamental Research Funds of Chinese Academy of Agricultural Sciences (Grant No. Y2016JC05), and the Agricultural Science and Technology Innovation Program (ASTIP) (Grant No. CAAS-ASTIP-2016-LVRI-03).

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Authors and Affiliations

  1. State Key Laboratory of Veterinary Etiological Biology, Key Laboratory of Veterinary Parasitology of Gansu Province, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou, China

    Fa-Cai Li, Qing Liu, Wen-Bin Yang, Jun-Ling Hou & Xing-Quan Zhu

  2. College of Animal Science and Veterinary Medicine, Shanxi Agricultural University, Taigu, China

    Qing Liu

  3. Faculty of Medicine and Health Sciences, School of Veterinary Medicine and Science, University of Nottingham, Sutton Bonington Campus, Loughborough, LE12 5RD, UK

    Hany M. Elsheikha

  4. College of Veterinary Medicine, Northwest A&F University, Yangling, China

    Wen-Bin Yang

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  1. Fa-Cai Li
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  2. Qing Liu
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  3. Hany M. Elsheikha
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  4. Wen-Bin Yang
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  5. Jun-Ling Hou
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  6. Xing-Quan Zhu
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Correspondence to Fa-Cai Li.

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Li, FC., Liu, Q., Elsheikha, H.M. et al. Identification of two novel host proteins interacting with Toxoplasma gondii 14-3-3 protein by yeast two-hybrid system. Parasitol Res 117, 1291–1296 (2018). https://doi.org/10.1007/s00436-018-5812-z

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  • DOI: https://doi.org/10.1007/s00436-018-5812-z

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