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Mincle-mediated translational regulation is required for strong nitric oxide production and inflammation resolution.

Lee WB, Kang JS, Choi WY, Zhang Q, Kim CH, Choi UY, Kim-Ha J, Kim YJ - Nat Commun (2016)

Bottom Line: Here we show that Mincle, the inducible receptor for mycobacterial cord factor, is the key switch for the transition of macrophages from cytokine expression to high nitric oxide production.In addition to its stimulatory role on TLR-mediated transcription, Mincle enhanced the translation of key genes required for nitric oxide synthesis through p38 and eIF5A hypusination, leading to granuloma resolution.Thus, Mincle has dual functions in the promotion and subsequent resolution of inflammation during anti-mycobacterial defence using both transcriptional and translational controls.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, College of Life Science and Biotechnology, Yonsei University, Seoul 03722, Republic of Korea.

ABSTRACT
In response to persistent mycobacteria infection, the host induces a granuloma, which often fails to eradicate bacteria and results in tissue damage. Diverse host receptors are required to control the formation and resolution of granuloma, but little is known concerning their regulatory interactions. Here we show that Mincle, the inducible receptor for mycobacterial cord factor, is the key switch for the transition of macrophages from cytokine expression to high nitric oxide production. In addition to its stimulatory role on TLR-mediated transcription, Mincle enhanced the translation of key genes required for nitric oxide synthesis through p38 and eIF5A hypusination, leading to granuloma resolution. Thus, Mincle has dual functions in the promotion and subsequent resolution of inflammation during anti-mycobacterial defence using both transcriptional and translational controls.

No MeSH data available.


Related in: MedlinePlus

TDM stimulation diminishes NLRP3-dependent inflammasome activation and IL-1β production.(a,b) WT and Mincle−/− BMDMs were stimulated with LPS or co-stimulated with LPS and TDM for 12 h or the indicated times and then treated with ATP for 1 h (L: LPS; T: TDM; A: ATP; UT: untreated). (a) Left: immunoblot analysis of proIL-1β, mature IL-1β (p17), procaspase-1 (proCasp-1) and cleaved caspase-1 (p10) in cell culture supernatants (SN) and whole-cell lysates (XT). Right: kinetic quantitative analysis of proIL-1β and mature IL-1β (p17) from the immunoblots. (b) Release of lactate dehydrogenase (LDH) (assessing cell death) from cells. (c) ELISA of released IL-1β from LPS-treated WT and Mincle−/− BMDMs, stimulated with TDM for 12 h and then treated with MSU, nigericin or poly(dA:dT) for 3 h. *P<0.05, **P<0.01, ***P<0.001 (two-tailed unpaired t-test). Data are representative of at least three independent experiments. (b,c: mean and s.d.).
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f1: TDM stimulation diminishes NLRP3-dependent inflammasome activation and IL-1β production.(a,b) WT and Mincle−/− BMDMs were stimulated with LPS or co-stimulated with LPS and TDM for 12 h or the indicated times and then treated with ATP for 1 h (L: LPS; T: TDM; A: ATP; UT: untreated). (a) Left: immunoblot analysis of proIL-1β, mature IL-1β (p17), procaspase-1 (proCasp-1) and cleaved caspase-1 (p10) in cell culture supernatants (SN) and whole-cell lysates (XT). Right: kinetic quantitative analysis of proIL-1β and mature IL-1β (p17) from the immunoblots. (b) Release of lactate dehydrogenase (LDH) (assessing cell death) from cells. (c) ELISA of released IL-1β from LPS-treated WT and Mincle−/− BMDMs, stimulated with TDM for 12 h and then treated with MSU, nigericin or poly(dA:dT) for 3 h. *P<0.05, **P<0.01, ***P<0.001 (two-tailed unpaired t-test). Data are representative of at least three independent experiments. (b,c: mean and s.d.).

Mentions: The stimulatory effect of Mincle on TLR signalling prompted us to examine its effect on inflammasome activation. To enhance inflammasome activation, exogenous ATP, a NLRP3 agonist, was added after varying durations of lipopolysaccharides (LPS) pretreatment15. LPS treatment enhanced proIL-1β messenger RNA (mRNA)/protein and enabled IL-1β maturation at 6 h after ATP addition (Fig. 1a and Supplementary Fig. 3). Although LPS treatment induced Mincle in WT macrophages, WT and Mincle−/− macrophages showed no difference in inflammatory responses in the absence of the Mincle ligand. Moreover, TDM alone was not able to induce proIL-1β mRNA, not to mention protein expression, before 48 h of stimulation (Supplementary Fig. 3). In contrast to sole treatment of TDM or LPS, co-treatment of TDM and LPS before ATP addition induced stronger proIL-1β mRNA and protein expression, which continued up to 48 h in WT macrophages (Supplementary Fig. 3). Furthermore, despite high-precursor protein levels, IL-1β secretion was abruptly reduced after 6 h (Fig. 1a and Supplementary Fig. 4a). In comparison to WT macrophages, Mincle−/− macrophages did not show extended proIL-1β mRNA and protein production, but produced much higher levels of mature IL-1β. Similar proIL-1β expression and IL-1β maturation were also observed with trehalose-6,6-dibehenate (TDB; a synthetic analogue of TDM) treatment (Supplementary Fig. 4b). However, the secretion levels of other cytokines, TNFα and IL-6, were not affected by TDM treatment or Mincle mutation (Supplementary Fig. 4c). In addition, caspase-1 maturation was similarly inhibited by TDM-mediated Mincle signalling (Fig. 1a). Therefore, TDM treatment is responsible for reduced pyroptotic cell death in WT macrophages (Fig. 1b). Taken together, these results demonstrate that TDM-mediated Mincle activation reduces NLRP3-inflammasome activation and cell death.


Mincle-mediated translational regulation is required for strong nitric oxide production and inflammation resolution.

Lee WB, Kang JS, Choi WY, Zhang Q, Kim CH, Choi UY, Kim-Ha J, Kim YJ - Nat Commun (2016)

TDM stimulation diminishes NLRP3-dependent inflammasome activation and IL-1β production.(a,b) WT and Mincle−/− BMDMs were stimulated with LPS or co-stimulated with LPS and TDM for 12 h or the indicated times and then treated with ATP for 1 h (L: LPS; T: TDM; A: ATP; UT: untreated). (a) Left: immunoblot analysis of proIL-1β, mature IL-1β (p17), procaspase-1 (proCasp-1) and cleaved caspase-1 (p10) in cell culture supernatants (SN) and whole-cell lysates (XT). Right: kinetic quantitative analysis of proIL-1β and mature IL-1β (p17) from the immunoblots. (b) Release of lactate dehydrogenase (LDH) (assessing cell death) from cells. (c) ELISA of released IL-1β from LPS-treated WT and Mincle−/− BMDMs, stimulated with TDM for 12 h and then treated with MSU, nigericin or poly(dA:dT) for 3 h. *P<0.05, **P<0.01, ***P<0.001 (two-tailed unpaired t-test). Data are representative of at least three independent experiments. (b,c: mean and s.d.).
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4837483&req=5

f1: TDM stimulation diminishes NLRP3-dependent inflammasome activation and IL-1β production.(a,b) WT and Mincle−/− BMDMs were stimulated with LPS or co-stimulated with LPS and TDM for 12 h or the indicated times and then treated with ATP for 1 h (L: LPS; T: TDM; A: ATP; UT: untreated). (a) Left: immunoblot analysis of proIL-1β, mature IL-1β (p17), procaspase-1 (proCasp-1) and cleaved caspase-1 (p10) in cell culture supernatants (SN) and whole-cell lysates (XT). Right: kinetic quantitative analysis of proIL-1β and mature IL-1β (p17) from the immunoblots. (b) Release of lactate dehydrogenase (LDH) (assessing cell death) from cells. (c) ELISA of released IL-1β from LPS-treated WT and Mincle−/− BMDMs, stimulated with TDM for 12 h and then treated with MSU, nigericin or poly(dA:dT) for 3 h. *P<0.05, **P<0.01, ***P<0.001 (two-tailed unpaired t-test). Data are representative of at least three independent experiments. (b,c: mean and s.d.).
Mentions: The stimulatory effect of Mincle on TLR signalling prompted us to examine its effect on inflammasome activation. To enhance inflammasome activation, exogenous ATP, a NLRP3 agonist, was added after varying durations of lipopolysaccharides (LPS) pretreatment15. LPS treatment enhanced proIL-1β messenger RNA (mRNA)/protein and enabled IL-1β maturation at 6 h after ATP addition (Fig. 1a and Supplementary Fig. 3). Although LPS treatment induced Mincle in WT macrophages, WT and Mincle−/− macrophages showed no difference in inflammatory responses in the absence of the Mincle ligand. Moreover, TDM alone was not able to induce proIL-1β mRNA, not to mention protein expression, before 48 h of stimulation (Supplementary Fig. 3). In contrast to sole treatment of TDM or LPS, co-treatment of TDM and LPS before ATP addition induced stronger proIL-1β mRNA and protein expression, which continued up to 48 h in WT macrophages (Supplementary Fig. 3). Furthermore, despite high-precursor protein levels, IL-1β secretion was abruptly reduced after 6 h (Fig. 1a and Supplementary Fig. 4a). In comparison to WT macrophages, Mincle−/− macrophages did not show extended proIL-1β mRNA and protein production, but produced much higher levels of mature IL-1β. Similar proIL-1β expression and IL-1β maturation were also observed with trehalose-6,6-dibehenate (TDB; a synthetic analogue of TDM) treatment (Supplementary Fig. 4b). However, the secretion levels of other cytokines, TNFα and IL-6, were not affected by TDM treatment or Mincle mutation (Supplementary Fig. 4c). In addition, caspase-1 maturation was similarly inhibited by TDM-mediated Mincle signalling (Fig. 1a). Therefore, TDM treatment is responsible for reduced pyroptotic cell death in WT macrophages (Fig. 1b). Taken together, these results demonstrate that TDM-mediated Mincle activation reduces NLRP3-inflammasome activation and cell death.

Bottom Line: Here we show that Mincle, the inducible receptor for mycobacterial cord factor, is the key switch for the transition of macrophages from cytokine expression to high nitric oxide production.In addition to its stimulatory role on TLR-mediated transcription, Mincle enhanced the translation of key genes required for nitric oxide synthesis through p38 and eIF5A hypusination, leading to granuloma resolution.Thus, Mincle has dual functions in the promotion and subsequent resolution of inflammation during anti-mycobacterial defence using both transcriptional and translational controls.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, College of Life Science and Biotechnology, Yonsei University, Seoul 03722, Republic of Korea.

ABSTRACT
In response to persistent mycobacteria infection, the host induces a granuloma, which often fails to eradicate bacteria and results in tissue damage. Diverse host receptors are required to control the formation and resolution of granuloma, but little is known concerning their regulatory interactions. Here we show that Mincle, the inducible receptor for mycobacterial cord factor, is the key switch for the transition of macrophages from cytokine expression to high nitric oxide production. In addition to its stimulatory role on TLR-mediated transcription, Mincle enhanced the translation of key genes required for nitric oxide synthesis through p38 and eIF5A hypusination, leading to granuloma resolution. Thus, Mincle has dual functions in the promotion and subsequent resolution of inflammation during anti-mycobacterial defence using both transcriptional and translational controls.

No MeSH data available.


Related in: MedlinePlus