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A Trypanosoma brucei kinesin heavy chain promotes parasite growth by triggering host arginase activity.

De Muylder G, Daulouède S, Lecordier L, Uzureau P, Morias Y, Van Den Abbeele J, Caljon G, Hérin M, Holzmuller P, Semballa S, Courtois P, Vanhamme L, Stijlemans B, De Baetselier P, Barrett MP, Barlow JL, McKenzie AN, Barron L, Wynn TA, Beschin A, Vincendeau P, Pays E - PLoS Pathog. (2013)

Bottom Line: By screening a cDNA library of T. brucei with an antibody neutralizing the arginase-inducing activity of parasite released factors, we identified a Kinesin Heavy Chain isoform, termed TbKHC1, as responsible for this effect.This TbKHC1 activity was IL-4Rα-independent and did not mirror M2 activation of myeloid cells.By treating infected mice with ornithine or with NO synthase inhibitor, we observed that during the first wave of parasitaemia the parasite growth-promoting effect of TbKHC1-mediated arginase activation resulted more from increased polyamine production than from reduction of NO synthesis.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Molecular Parasitology, IBMM, Université Libre de Bruxelles (ULB), Gosselies, Belgium.

ABSTRACT

Background: In order to promote infection, the blood-borne parasite Trypanosoma brucei releases factors that upregulate arginase expression and activity in myeloid cells.

Methodology/principal findings: By screening a cDNA library of T. brucei with an antibody neutralizing the arginase-inducing activity of parasite released factors, we identified a Kinesin Heavy Chain isoform, termed TbKHC1, as responsible for this effect. Following interaction with mouse myeloid cells, natural or recombinant TbKHC1 triggered SIGN-R1 receptor-dependent induction of IL-10 production, resulting in arginase-1 activation concomitant with reduction of nitric oxide (NO) synthase activity. This TbKHC1 activity was IL-4Rα-independent and did not mirror M2 activation of myeloid cells. As compared to wild-type T. brucei, infection by TbKHC1 KO parasites was characterized by strongly reduced parasitaemia and prolonged host survival time. By treating infected mice with ornithine or with NO synthase inhibitor, we observed that during the first wave of parasitaemia the parasite growth-promoting effect of TbKHC1-mediated arginase activation resulted more from increased polyamine production than from reduction of NO synthesis. In late stage infection, TbKHC1-mediated reduction of NO synthesis appeared to contribute to liver damage linked to shortening of host survival time.

Conclusion: A kinesin heavy chain released by T. brucei induces IL-10 and arginase-1 through SIGN-R1 signaling in myeloid cells, which promotes early trypanosome growth and favors parasite settlement in the host. Moreover, in the late stage of infection, the inhibition of NO synthesis by TbKHC1 contributes to liver pathogenicity.

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Expression and localization of T. brucei TbKHC1.(A) Detection of TbKHC1 transcripts by Northern blotting of RNA from procyclic forms (PF), bloodstream (BF) slender (SL) and stumpy (ST) forms, BF TbKHC1 knock-out (KO), overexpressor (OV) and TbKHC1 knock-down (RNAi) trypanosomes (d: days after doxycyclin induction; ds RNA: double-stranded RNA; histone H2B mRNA: loading control). (B) Detection of TbKHC1 with anti-rTbKHC1 polyclonal antibody in whole extracts from the indicated trypanosome forms (Tub: Tubulin loading control). (C) Parasite staining with anti-PRF monoclonal 2C12 antibody. Nucleus (N) and kinetoplast (K) are stained in blue with 4′,6-Diamidino-2-phenylindole (DAPI). (D) Localization of TbKHC1 on WT, KO and OV trypanosomes with anti-rTbKHC1 polyclonal antibody. (E) Immunoprecipitation of TbKHC1, actin, enolase and PDI-2 from 35S-metabolically labelled WT parasite total extracts (1) or supernatants (2).
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ppat-1003731-g002: Expression and localization of T. brucei TbKHC1.(A) Detection of TbKHC1 transcripts by Northern blotting of RNA from procyclic forms (PF), bloodstream (BF) slender (SL) and stumpy (ST) forms, BF TbKHC1 knock-out (KO), overexpressor (OV) and TbKHC1 knock-down (RNAi) trypanosomes (d: days after doxycyclin induction; ds RNA: double-stranded RNA; histone H2B mRNA: loading control). (B) Detection of TbKHC1 with anti-rTbKHC1 polyclonal antibody in whole extracts from the indicated trypanosome forms (Tub: Tubulin loading control). (C) Parasite staining with anti-PRF monoclonal 2C12 antibody. Nucleus (N) and kinetoplast (K) are stained in blue with 4′,6-Diamidino-2-phenylindole (DAPI). (D) Localization of TbKHC1 on WT, KO and OV trypanosomes with anti-rTbKHC1 polyclonal antibody. (E) Immunoprecipitation of TbKHC1, actin, enolase and PDI-2 from 35S-metabolically labelled WT parasite total extracts (1) or supernatants (2).

Mentions: Pleomorphic trypanosomes, differentiating in the bloodstream from proliferative slender forms into quiescent stumpy forms, can induce long-standing infection in mammals and perform cyclical transmission in tsetse flies while differentiating into procyclic forms. In contrast, monomorphic trypanosomes, resulting from prolonged cultivation in vitro, are unable to develop long-lasting infection and cyclical transmission. The TbKHC1 gene was found to be expressed preferentially in slender bloodstream forms (Fig. 2A,B). Parasite immunolabeling with both monoclonal 2C12 and polyclonal antibodies generated against recombinant TbKHC1 (rTbKHC1) vanished in TbKHC1 KO parasites or RNAi-mediated TbKHC1 knock-down (KD) parasites, while it increased in trypanosomes over-expressing TbKHC1 (Fig. 2A–D), confirming that TbKHC1 is the genuine target of the anti-PRF 2C12 antibody.


A Trypanosoma brucei kinesin heavy chain promotes parasite growth by triggering host arginase activity.

De Muylder G, Daulouède S, Lecordier L, Uzureau P, Morias Y, Van Den Abbeele J, Caljon G, Hérin M, Holzmuller P, Semballa S, Courtois P, Vanhamme L, Stijlemans B, De Baetselier P, Barrett MP, Barlow JL, McKenzie AN, Barron L, Wynn TA, Beschin A, Vincendeau P, Pays E - PLoS Pathog. (2013)

Expression and localization of T. brucei TbKHC1.(A) Detection of TbKHC1 transcripts by Northern blotting of RNA from procyclic forms (PF), bloodstream (BF) slender (SL) and stumpy (ST) forms, BF TbKHC1 knock-out (KO), overexpressor (OV) and TbKHC1 knock-down (RNAi) trypanosomes (d: days after doxycyclin induction; ds RNA: double-stranded RNA; histone H2B mRNA: loading control). (B) Detection of TbKHC1 with anti-rTbKHC1 polyclonal antibody in whole extracts from the indicated trypanosome forms (Tub: Tubulin loading control). (C) Parasite staining with anti-PRF monoclonal 2C12 antibody. Nucleus (N) and kinetoplast (K) are stained in blue with 4′,6-Diamidino-2-phenylindole (DAPI). (D) Localization of TbKHC1 on WT, KO and OV trypanosomes with anti-rTbKHC1 polyclonal antibody. (E) Immunoprecipitation of TbKHC1, actin, enolase and PDI-2 from 35S-metabolically labelled WT parasite total extracts (1) or supernatants (2).
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Related In: Results  -  Collection

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ppat-1003731-g002: Expression and localization of T. brucei TbKHC1.(A) Detection of TbKHC1 transcripts by Northern blotting of RNA from procyclic forms (PF), bloodstream (BF) slender (SL) and stumpy (ST) forms, BF TbKHC1 knock-out (KO), overexpressor (OV) and TbKHC1 knock-down (RNAi) trypanosomes (d: days after doxycyclin induction; ds RNA: double-stranded RNA; histone H2B mRNA: loading control). (B) Detection of TbKHC1 with anti-rTbKHC1 polyclonal antibody in whole extracts from the indicated trypanosome forms (Tub: Tubulin loading control). (C) Parasite staining with anti-PRF monoclonal 2C12 antibody. Nucleus (N) and kinetoplast (K) are stained in blue with 4′,6-Diamidino-2-phenylindole (DAPI). (D) Localization of TbKHC1 on WT, KO and OV trypanosomes with anti-rTbKHC1 polyclonal antibody. (E) Immunoprecipitation of TbKHC1, actin, enolase and PDI-2 from 35S-metabolically labelled WT parasite total extracts (1) or supernatants (2).
Mentions: Pleomorphic trypanosomes, differentiating in the bloodstream from proliferative slender forms into quiescent stumpy forms, can induce long-standing infection in mammals and perform cyclical transmission in tsetse flies while differentiating into procyclic forms. In contrast, monomorphic trypanosomes, resulting from prolonged cultivation in vitro, are unable to develop long-lasting infection and cyclical transmission. The TbKHC1 gene was found to be expressed preferentially in slender bloodstream forms (Fig. 2A,B). Parasite immunolabeling with both monoclonal 2C12 and polyclonal antibodies generated against recombinant TbKHC1 (rTbKHC1) vanished in TbKHC1 KO parasites or RNAi-mediated TbKHC1 knock-down (KD) parasites, while it increased in trypanosomes over-expressing TbKHC1 (Fig. 2A–D), confirming that TbKHC1 is the genuine target of the anti-PRF 2C12 antibody.

Bottom Line: By screening a cDNA library of T. brucei with an antibody neutralizing the arginase-inducing activity of parasite released factors, we identified a Kinesin Heavy Chain isoform, termed TbKHC1, as responsible for this effect.This TbKHC1 activity was IL-4Rα-independent and did not mirror M2 activation of myeloid cells.By treating infected mice with ornithine or with NO synthase inhibitor, we observed that during the first wave of parasitaemia the parasite growth-promoting effect of TbKHC1-mediated arginase activation resulted more from increased polyamine production than from reduction of NO synthesis.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Molecular Parasitology, IBMM, Université Libre de Bruxelles (ULB), Gosselies, Belgium.

ABSTRACT

Background: In order to promote infection, the blood-borne parasite Trypanosoma brucei releases factors that upregulate arginase expression and activity in myeloid cells.

Methodology/principal findings: By screening a cDNA library of T. brucei with an antibody neutralizing the arginase-inducing activity of parasite released factors, we identified a Kinesin Heavy Chain isoform, termed TbKHC1, as responsible for this effect. Following interaction with mouse myeloid cells, natural or recombinant TbKHC1 triggered SIGN-R1 receptor-dependent induction of IL-10 production, resulting in arginase-1 activation concomitant with reduction of nitric oxide (NO) synthase activity. This TbKHC1 activity was IL-4Rα-independent and did not mirror M2 activation of myeloid cells. As compared to wild-type T. brucei, infection by TbKHC1 KO parasites was characterized by strongly reduced parasitaemia and prolonged host survival time. By treating infected mice with ornithine or with NO synthase inhibitor, we observed that during the first wave of parasitaemia the parasite growth-promoting effect of TbKHC1-mediated arginase activation resulted more from increased polyamine production than from reduction of NO synthesis. In late stage infection, TbKHC1-mediated reduction of NO synthesis appeared to contribute to liver damage linked to shortening of host survival time.

Conclusion: A kinesin heavy chain released by T. brucei induces IL-10 and arginase-1 through SIGN-R1 signaling in myeloid cells, which promotes early trypanosome growth and favors parasite settlement in the host. Moreover, in the late stage of infection, the inhibition of NO synthesis by TbKHC1 contributes to liver pathogenicity.

Show MeSH
Related in: MedlinePlus