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Mol Cell Proteomics. 2017 Mar;16(3):469-484. doi: 10.1074/mcp.M116.063602. Epub 2017 Jan 13.

A Human Proteome Array Approach to Identifying Key Host Proteins Targeted by Toxoplasma Kinase ROP18.

Author information

1
From the ‡Department of Biochemistry and Molecular Biology, Sun Yat-Sen University Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou 510080, China.
2
the §Department of Pharmacology and Molecular Sciences, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205.
3
the ¶Department of Ophthalmology, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21287.
4
the ‖State Key Lab of Ophthalmology, Guangdong Provincial Key Lab of Ophthalmology and Visual Science, Zhongshan Ophthalmic Center, Sun Yat-Sen University, Guangzhou 510060, China.
5
the **School of Life Sciences, Sun Yat-Sen University, Guangzhou 510275, China.
6
‡‡The Third Affiliated Hospital, Sun Yat-Sen University, Guangzhou 510630, China; and.
7
§§The First Affiliated Hospital, Sun Yat-Sen University, Guangzhou 510080, China.
8
From the ‡Department of Biochemistry and Molecular Biology, Sun Yat-Sen University Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou 510080, China; zhouxw2@mail.sysu.edu.cn hzhu4@jhmi.edu liaowq5@mail.sysu.edu.cn.
9
the §Department of Pharmacology and Molecular Sciences, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205; zhouxw2@mail.sysu.edu.cn hzhu4@jhmi.edu liaowq5@mail.sysu.edu.cn.

Abstract

Toxoplasma kinase ROP18 is a key molecule responsible for the virulence of Toxoplasma gondii; however, the mechanisms by which ROP18 exerts parasite virulence via interaction with host proteins remain limited to a small number of identified substrates. To identify a broader array of ROP18 substrates, we successfully purified bioactive mature ROP18 and used it to probe a human proteome array. Sixty eight new putative host targets were identified. Functional annotation analysis suggested that these proteins have a variety of functions, including metabolic process, kinase activity and phosphorylation, cell growth, apoptosis and cell death, and immunity, indicating a pleiotropic role of ROP18 kinase. Among these proteins, four candidates, p53, p38, UBE2N, and Smad1, were further validated. We demonstrated that ROP18 targets p53, p38, UBE2N, and Smad1 for degradation. Importantly, we demonstrated that ROP18 phosphorylates Smad1 Ser-187 to trigger its proteasome-dependent degradation. Further functional characterization of the substrates of ROP18 may enhance understanding of the pathogenesis of Toxoplasma infection and provide new therapeutic targets. Similar strategies could be used to identify novel host targets for other microbial kinases functioning at the pathogen-host interface.

PMID:
28087594
PMCID:
PMC5341007
DOI:
10.1074/mcp.M116.063602
[Indexed for MEDLINE]
Free PMC Article

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