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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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SIGN-R1 receptor contribution to myeloid cell activation and liver injury.(A,B) Effects of rTbKHC1 on myeloid cells from non-infected control (Ctrl) or SIGN-R1 KO mice (A: relative expression of Arg1, Arg2, Il10 and iNOS genes; B: arginase activity in presence of indicated antibodies or sugars). Data are means ± SEM of 3 individual mice of one representative from 3 independent experiments. * p<0.05 compared to non-stimulated (-) cells. (C) Effects of rTbKHC1 on liver injury: microscopic analysis (hematoxylin-eosin staining, magnification 20×) of sections from WT- and TbKHC1 KO-infected mice at day 30 p.i. Anoxic infarcts (ai), periportal (>>) and lobular (>) mononuclear cell infiltrates were representative of 8 animals tested in 2 independent experiments. (D) Spontaneous NO and IL-10 secretions in spleen myeloid cell supernatants from WT- and TbKHC1 KO-infected mice treated with D-NAME or L-NAME (day 30 p.i.). (E) Idem as D in SIGN-R1 KO and control (Ctrl) mice. Data are means ± SEM of 3–4 individual mice of one representative from 3 independent experiments. ∇ p<0.05 comparing WT or TbKHC1 KO- infected mice to non-infected mice; £ p<0.05 comparing L-NAME and D-NAME treatment in WT- or TbKHC1 KO-infected mice; # p<0.05 comparing WT- and TbKHC1 KO-infected mice.
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ppat-1003731-g006: SIGN-R1 receptor contribution to myeloid cell activation and liver injury.(A,B) Effects of rTbKHC1 on myeloid cells from non-infected control (Ctrl) or SIGN-R1 KO mice (A: relative expression of Arg1, Arg2, Il10 and iNOS genes; B: arginase activity in presence of indicated antibodies or sugars). Data are means ± SEM of 3 individual mice of one representative from 3 independent experiments. * p<0.05 compared to non-stimulated (-) cells. (C) Effects of rTbKHC1 on liver injury: microscopic analysis (hematoxylin-eosin staining, magnification 20×) of sections from WT- and TbKHC1 KO-infected mice at day 30 p.i. Anoxic infarcts (ai), periportal (>>) and lobular (>) mononuclear cell infiltrates were representative of 8 animals tested in 2 independent experiments. (D) Spontaneous NO and IL-10 secretions in spleen myeloid cell supernatants from WT- and TbKHC1 KO-infected mice treated with D-NAME or L-NAME (day 30 p.i.). (E) Idem as D in SIGN-R1 KO and control (Ctrl) mice. Data are means ± SEM of 3–4 individual mice of one representative from 3 independent experiments. ∇ p<0.05 comparing WT or TbKHC1 KO- infected mice to non-infected mice; £ p<0.05 comparing L-NAME and D-NAME treatment in WT- or TbKHC1 KO-infected mice; # p<0.05 comparing WT- and TbKHC1 KO-infected mice.

Mentions: Since the in vitro induction of arginase activity by PRF and rTbKHC1 was inhibited by D-mannose (Fig. 3B,C), we treated infected mice with D-mannose. This treatment reduced WT parasitaemia to the level of TbKHC1 KO parasitaemia, but did not significantly affect TbKHC1 KO parasitaemia (Fig. 5F). To follow up on this in vivo finding and the in vitro observation that arginase activation by PRF and rTbKHC1 was inhibited by D-mannose as well as by an antibody recognizing coils of TbKHC1 (Fig. 1B, 3C), we monitored the course of WT- and TbKHC1 KO-infection in mice deficient for myeloid cell receptors able to bind both mannose and peptidic coils, namely MMR (CD206) and SIGN-R1 (CD209b), using appropriate congenic control mice [25]–[27]. In MMR KO mice, WT and TbKHC1 KO parasitaemias were not affected as compared to infection in WT mice (Fig. 5G). In SIGN-R1 KO animals the differential control of the first peak of parasitaemia between WT and TbKHC1 KO parasites vanished, due to the reduction of the WT parasite level to that of TbKHC1 KO parasites (Fig. 5H). Thus, either the absence of SIGN-R1 in infected mice or the absence of TbKHC1 in the parasite decreased T. brucei parasitaemia similarly. These data suggested that TbKHC1 interacts with the SIGN-R1 receptor. To evaluate this hypothesis, we tested the in vitro activity of rTbKHC1 on myeloid cells from SIGN-R1 KO mice. In contrast to results obtained with myeloid cells from control mice, in myeloid cells from SIGN-RI KO mice rTbKHC1 did not stimulate IL-10 and Arg1 gene expression (Fig. 6A) and did not increase arginase activity (Fig. 6B). Conversely, in myeloid cells from MMR KO mice rTbKHC1 induced expression of Il10 and Arg1 genes as in WT mouse myeloid cells (Fig. S5 in Text S1). Therefore, TbKHC1 appeared to trigger arginase activity through SIGN-R1-mediated signaling.


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)

SIGN-R1 receptor contribution to myeloid cell activation and liver injury.(A,B) Effects of rTbKHC1 on myeloid cells from non-infected control (Ctrl) or SIGN-R1 KO mice (A: relative expression of Arg1, Arg2, Il10 and iNOS genes; B: arginase activity in presence of indicated antibodies or sugars). Data are means ± SEM of 3 individual mice of one representative from 3 independent experiments. * p<0.05 compared to non-stimulated (-) cells. (C) Effects of rTbKHC1 on liver injury: microscopic analysis (hematoxylin-eosin staining, magnification 20×) of sections from WT- and TbKHC1 KO-infected mice at day 30 p.i. Anoxic infarcts (ai), periportal (>>) and lobular (>) mononuclear cell infiltrates were representative of 8 animals tested in 2 independent experiments. (D) Spontaneous NO and IL-10 secretions in spleen myeloid cell supernatants from WT- and TbKHC1 KO-infected mice treated with D-NAME or L-NAME (day 30 p.i.). (E) Idem as D in SIGN-R1 KO and control (Ctrl) mice. Data are means ± SEM of 3–4 individual mice of one representative from 3 independent experiments. ∇ p<0.05 comparing WT or TbKHC1 KO- infected mice to non-infected mice; £ p<0.05 comparing L-NAME and D-NAME treatment in WT- or TbKHC1 KO-infected mice; # p<0.05 comparing WT- and TbKHC1 KO-infected mice.
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Related In: Results  -  Collection

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ppat-1003731-g006: SIGN-R1 receptor contribution to myeloid cell activation and liver injury.(A,B) Effects of rTbKHC1 on myeloid cells from non-infected control (Ctrl) or SIGN-R1 KO mice (A: relative expression of Arg1, Arg2, Il10 and iNOS genes; B: arginase activity in presence of indicated antibodies or sugars). Data are means ± SEM of 3 individual mice of one representative from 3 independent experiments. * p<0.05 compared to non-stimulated (-) cells. (C) Effects of rTbKHC1 on liver injury: microscopic analysis (hematoxylin-eosin staining, magnification 20×) of sections from WT- and TbKHC1 KO-infected mice at day 30 p.i. Anoxic infarcts (ai), periportal (>>) and lobular (>) mononuclear cell infiltrates were representative of 8 animals tested in 2 independent experiments. (D) Spontaneous NO and IL-10 secretions in spleen myeloid cell supernatants from WT- and TbKHC1 KO-infected mice treated with D-NAME or L-NAME (day 30 p.i.). (E) Idem as D in SIGN-R1 KO and control (Ctrl) mice. Data are means ± SEM of 3–4 individual mice of one representative from 3 independent experiments. ∇ p<0.05 comparing WT or TbKHC1 KO- infected mice to non-infected mice; £ p<0.05 comparing L-NAME and D-NAME treatment in WT- or TbKHC1 KO-infected mice; # p<0.05 comparing WT- and TbKHC1 KO-infected mice.
Mentions: Since the in vitro induction of arginase activity by PRF and rTbKHC1 was inhibited by D-mannose (Fig. 3B,C), we treated infected mice with D-mannose. This treatment reduced WT parasitaemia to the level of TbKHC1 KO parasitaemia, but did not significantly affect TbKHC1 KO parasitaemia (Fig. 5F). To follow up on this in vivo finding and the in vitro observation that arginase activation by PRF and rTbKHC1 was inhibited by D-mannose as well as by an antibody recognizing coils of TbKHC1 (Fig. 1B, 3C), we monitored the course of WT- and TbKHC1 KO-infection in mice deficient for myeloid cell receptors able to bind both mannose and peptidic coils, namely MMR (CD206) and SIGN-R1 (CD209b), using appropriate congenic control mice [25]–[27]. In MMR KO mice, WT and TbKHC1 KO parasitaemias were not affected as compared to infection in WT mice (Fig. 5G). In SIGN-R1 KO animals the differential control of the first peak of parasitaemia between WT and TbKHC1 KO parasites vanished, due to the reduction of the WT parasite level to that of TbKHC1 KO parasites (Fig. 5H). Thus, either the absence of SIGN-R1 in infected mice or the absence of TbKHC1 in the parasite decreased T. brucei parasitaemia similarly. These data suggested that TbKHC1 interacts with the SIGN-R1 receptor. To evaluate this hypothesis, we tested the in vitro activity of rTbKHC1 on myeloid cells from SIGN-R1 KO mice. In contrast to results obtained with myeloid cells from control mice, in myeloid cells from SIGN-RI KO mice rTbKHC1 did not stimulate IL-10 and Arg1 gene expression (Fig. 6A) and did not increase arginase activity (Fig. 6B). Conversely, in myeloid cells from MMR KO mice rTbKHC1 induced expression of Il10 and Arg1 genes as in WT mouse myeloid cells (Fig. S5 in Text S1). Therefore, TbKHC1 appeared to trigger arginase activity through SIGN-R1-mediated signaling.

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