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Interaction between parasite-encoded JAB1/CSN5 and macrophage migration inhibitory factor proteins attenuates its proinflammatory function.

ABSTRACT: Multiple protozoans produce homologs of the cytokine MIF which play a role in immune evasion, invasion and pathogenesis. However, how parasite-encoded MIF activity is controlled remains poorly understood. Cytokine activity can be inhibited by intracellular binding partners that are released in the extracellular space during cell death. We investigated the presence of an endogenous parasite protein that was capable of interacting and interfering with MIF activity. A screen for protein-protein interaction was performed using immunoaffinity purification of amebic cell lysate with specific anti-Entamoeba histolytica MIF (EhMIF) antibody followed by mass spectrometry analysis, which revealed an E. histolytica-produced JAB1 protein (EhJAB1) as a potential binding partner. JAB1 was found to be highly conserved in protozoans. Direct interaction between the EhMIF and EhJAB1 was confirmed by several independent approaches with GST pull-down, co-immunoprecipitation, and Biolayer interferometry (BLI) assays. Furthermore, the C-terminal region outside the functional JAMM deneddylase motif was required for EhMIF binding, which was consistent with the top in silico predictions. In addition, EhJAB1 binding blocked EhMIF-induced IL-8 production by human epithelial cells. We report the initial characterization of a parasite-encoded JAB1 and uncover a new binding partner for a protozoan-produced MIF protein, acting as a possible negative regulator of EhMIF.

SUBMITTER: Ghosh S

PROVIDER: S-EPMC6035221 | biostudies-literature | 2018 Jul

REPOSITORIES: biostudies-literature

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Interaction between parasite-encoded JAB1/CSN5 and macrophage migration inhibitory factor proteins attenuates its proinflammatory function.

Ghosh Swagata S Leaton Laura Ann LA Farr Laura L Barfield Alexis A Moonah Shannon S

Scientific reports 20180706 1

Multiple protozoans produce homologs of the cytokine MIF which play a role in immune evasion, invasion and pathogenesis. However, how parasite-encoded MIF activity is controlled remains poorly understood. Cytokine activity can be inhibited by intracellular binding partners that are released in the extracellular space during cell death. We investigated the presence of an endogenous parasite protein that was capable of interacting and interfering with MIF activity. A screen for protein-protein int ...[more]

PMID: 29980718

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Project description:BackgroundThe danger-associated molecular patterns (DAMPs) are critical contributors to the progressive neuropathology and thereafter affect the functional outcomes following spinal cord injury (SCI). Up to now, the regulatory mechanisms on their inducible production from the living cells remain elusive, aside from their passive release from the necrotic cells. Thrombin is immediately activated by the damaged or stressed central nervous system (CNS), which potently mediates inflammatory astrocytic responses through proteolytic cleavage of protease-activated receptors (PARs). Therefore, SCI-activated thrombin is conceived to induce the production of DAMPs from astrocytes at lesion site.MethodsRat SCI model was established by the cord contusion at T8-T10. The expression of thrombin and macrophage migration inhibitory factor (MIF) was determined by ELISA and Western blot. The PAR1, PAR3, and PAR4 receptors of thrombin were examined by PCR and immunohistochemistry. Primary astrocytes were isolated and purified from the spinal cord, followed by stimulation with different concentrations of thrombin either for transcriptome sequencing or for analysis of thrombin-mediated expression of MIF and related signal pathways in the presence or absence of various inhibitors. The post-injury locomotor functions were assessed using the Basso, Beattie, and Bresnahan (BBB) locomotor scale.ResultsMIF protein levels were significantly elevated in parallel with those of thrombin induced by SCI. Immunostaining demonstrated that PAR1 receptor, together with MIF, was abundantly expressed in astrocytes. By transcriptome sequencing and bioinformatical analysis of thrombin-stimulated primary astrocytes, MIF was identified to be dynamically regulated by the serine protease. Investigation of the underlying mechanism using various inhibitors revealed that thrombin-activated PAR1 was responsible for the MIF production of astrocytes through modulation of JNK/NFκB pathway. Administration of PAR1 inhibitor at lesion sites following SCI significantly reduced the protein levels of MIF and ameliorated functional deficits of rat locomotion.ConclusionSCI-activated thrombin is a robust inducer of MIF production from astrocytes. Exploring the roles of thrombin in promoting the production of DAMPs from astrocytes at lesion site will provide an alternative strategy for the clinical therapy of CNS inflammation.

Dataset Information

Interaction between parasite-encoded JAB1/CSN5 and macrophage migration inhibitory factor proteins attenuates its proinflammatory function.

Publications

Interaction between parasite-encoded JAB1/CSN5 and macrophage migration inhibitory factor proteins attenuates its proinflammatory function.

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