Limits...
LRRFIP2 negatively regulates NLRP3 inflammasome activation in macrophages by promoting Flightless-I-mediated caspase-1 inhibition.

Jin J, Yu Q, Han C, Hu X, Xu S, Wang Q, Wang J, Li N, Cao X - Nat Commun (2013)

Bottom Line: Knockdown of Flightless-I significantly promotes NLRP3 inflammasome activation.LRRFIP2 enhances the interaction between Flightless-I and caspase-1, facilitating the inhibitory effect of Flightless-I on caspase-1 activation.Furthermore, silencing of Flightless-I abrogates the inhibitory effect of LRRFIP2 on NLRP3 inflammasome.

View Article: PubMed Central - PubMed

Affiliation: Institute of Immunology, Zhejiang University School of Medicine, 866 Yu-Hang-Tang Road, Hangzhou 310058, China.

ABSTRACT
The NLRP3 inflammasome is the most characterized inflammasome activated by cellular infection or stress, which is responsible for the maturation of proinflammatory cytokines IL-1β and IL-18. The precise molecular mechanism for negative regulation of NLRP3 inflammasome activation needs to be further defined. Here we identify leucine-rich repeat Fli-I-interacting protein 2 (LRRFIP2) as an NLRP3-associated protein and an inhibitor for NLRP3 inflammasome activation. LRRFIP2 binds to NLRP3 via its N terminus upon NLRP3 inflammasome activation, and also interacts with Flightless-I, a pseudosubstrate of caspase-1, via its Coil motif. Knockdown of Flightless-I significantly promotes NLRP3 inflammasome activation. LRRFIP2 enhances the interaction between Flightless-I and caspase-1, facilitating the inhibitory effect of Flightless-I on caspase-1 activation. Furthermore, silencing of Flightless-I abrogates the inhibitory effect of LRRFIP2 on NLRP3 inflammasome. These data demonstrate that LRRFIP2 inhibits NLRP3 inflammasome activation by recruiting the caspase-1 inhibitor Flightless-I, thus outlining a new mechanism for negative regulation of NLRP3 inflammasome.

Show MeSH

Related in: MedlinePlus

LRRFIP2 suppresses Alum-induced peritonitis in vivo.(a) Immunoblot analysis with indicated antibodies of PECs recovered 12 h after alum injection from LRRFIP2-silenced mice and control (Ctrl) mice (n=5–7). (b) IL-1β content in the lavage fluid 8 h after alum injection from LRRFIP2-silenced mice and Ctrl mice (n=5–7). **P=0.0074 (two-tailed Student’s t-test). (c) Inflammatory cell subset analysis by flow cytometry of PECs recovered 12 h after alum injection (n=5). *P=0.0282; **P=0.0029; ***P=0.0038 (analysis of variance). Data shown are representative of three separate experiments (a,c) or are means±s.e.m. of three independent experiments (b).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC3753543&req=5

f6: LRRFIP2 suppresses Alum-induced peritonitis in vivo.(a) Immunoblot analysis with indicated antibodies of PECs recovered 12 h after alum injection from LRRFIP2-silenced mice and control (Ctrl) mice (n=5–7). (b) IL-1β content in the lavage fluid 8 h after alum injection from LRRFIP2-silenced mice and Ctrl mice (n=5–7). **P=0.0074 (two-tailed Student’s t-test). (c) Inflammatory cell subset analysis by flow cytometry of PECs recovered 12 h after alum injection (n=5). *P=0.0282; **P=0.0029; ***P=0.0038 (analysis of variance). Data shown are representative of three separate experiments (a,c) or are means±s.e.m. of three independent experiments (b).

Mentions: Given that LRRFIP2 inhibits NLRP3 inflammasome activation in vitro, we further demonstrated the biological effect of LRRFIP2 in vivo by knocking down LRRFIP2 in a mouse peritonitis model. The siRNA oligos complexed with in vivo-jetPEI, a linear polyethylenimine, significantly reduced the endogenous protein level of LRRFIP2 in the peritoneal cavity (Fig. 6a). Mice transfected with siRNA–jetPEI complex was stimulated by Alum to trigger peritonitis. Silencing of LRRFIP2 enhanced caspase-1 activation in vivo but didn’t affect the expression of NLRP3 and pro-caspase-1 in peritoneal exudates cells (PECs; Fig. 6a). IL-1β secretion in the lavage fluid was significantly increased by in vivo LRRFIP2 knockdown (Fig. 6a,b), which was in line with the observation in vitro. Alum-induced recruitment of inflammatory cells was then analysed by flow cytometry. The number of total PECs recruited upon Alum challenge was markedly increased in mice silencing of LRRFIP2 (Fig. 6c). Neutrophils and Ly6C+ monocytes recruitment were also significantly increased in the si-LRRFIP2 transfection group (Fig. 6c). These findings further demonstrated that LRRFIP2 can inhibit NLRP3 inflammasome activation and subsequent immune cell accumulation in mouse peritonitis in vivo.


LRRFIP2 negatively regulates NLRP3 inflammasome activation in macrophages by promoting Flightless-I-mediated caspase-1 inhibition.

Jin J, Yu Q, Han C, Hu X, Xu S, Wang Q, Wang J, Li N, Cao X - Nat Commun (2013)

LRRFIP2 suppresses Alum-induced peritonitis in vivo.(a) Immunoblot analysis with indicated antibodies of PECs recovered 12 h after alum injection from LRRFIP2-silenced mice and control (Ctrl) mice (n=5–7). (b) IL-1β content in the lavage fluid 8 h after alum injection from LRRFIP2-silenced mice and Ctrl mice (n=5–7). **P=0.0074 (two-tailed Student’s t-test). (c) Inflammatory cell subset analysis by flow cytometry of PECs recovered 12 h after alum injection (n=5). *P=0.0282; **P=0.0029; ***P=0.0038 (analysis of variance). Data shown are representative of three separate experiments (a,c) or are means±s.e.m. of three independent experiments (b).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3753543&req=5

f6: LRRFIP2 suppresses Alum-induced peritonitis in vivo.(a) Immunoblot analysis with indicated antibodies of PECs recovered 12 h after alum injection from LRRFIP2-silenced mice and control (Ctrl) mice (n=5–7). (b) IL-1β content in the lavage fluid 8 h after alum injection from LRRFIP2-silenced mice and Ctrl mice (n=5–7). **P=0.0074 (two-tailed Student’s t-test). (c) Inflammatory cell subset analysis by flow cytometry of PECs recovered 12 h after alum injection (n=5). *P=0.0282; **P=0.0029; ***P=0.0038 (analysis of variance). Data shown are representative of three separate experiments (a,c) or are means±s.e.m. of three independent experiments (b).
Mentions: Given that LRRFIP2 inhibits NLRP3 inflammasome activation in vitro, we further demonstrated the biological effect of LRRFIP2 in vivo by knocking down LRRFIP2 in a mouse peritonitis model. The siRNA oligos complexed with in vivo-jetPEI, a linear polyethylenimine, significantly reduced the endogenous protein level of LRRFIP2 in the peritoneal cavity (Fig. 6a). Mice transfected with siRNA–jetPEI complex was stimulated by Alum to trigger peritonitis. Silencing of LRRFIP2 enhanced caspase-1 activation in vivo but didn’t affect the expression of NLRP3 and pro-caspase-1 in peritoneal exudates cells (PECs; Fig. 6a). IL-1β secretion in the lavage fluid was significantly increased by in vivo LRRFIP2 knockdown (Fig. 6a,b), which was in line with the observation in vitro. Alum-induced recruitment of inflammatory cells was then analysed by flow cytometry. The number of total PECs recruited upon Alum challenge was markedly increased in mice silencing of LRRFIP2 (Fig. 6c). Neutrophils and Ly6C+ monocytes recruitment were also significantly increased in the si-LRRFIP2 transfection group (Fig. 6c). These findings further demonstrated that LRRFIP2 can inhibit NLRP3 inflammasome activation and subsequent immune cell accumulation in mouse peritonitis in vivo.

Bottom Line: Knockdown of Flightless-I significantly promotes NLRP3 inflammasome activation.LRRFIP2 enhances the interaction between Flightless-I and caspase-1, facilitating the inhibitory effect of Flightless-I on caspase-1 activation.Furthermore, silencing of Flightless-I abrogates the inhibitory effect of LRRFIP2 on NLRP3 inflammasome.

View Article: PubMed Central - PubMed

Affiliation: Institute of Immunology, Zhejiang University School of Medicine, 866 Yu-Hang-Tang Road, Hangzhou 310058, China.

ABSTRACT
The NLRP3 inflammasome is the most characterized inflammasome activated by cellular infection or stress, which is responsible for the maturation of proinflammatory cytokines IL-1β and IL-18. The precise molecular mechanism for negative regulation of NLRP3 inflammasome activation needs to be further defined. Here we identify leucine-rich repeat Fli-I-interacting protein 2 (LRRFIP2) as an NLRP3-associated protein and an inhibitor for NLRP3 inflammasome activation. LRRFIP2 binds to NLRP3 via its N terminus upon NLRP3 inflammasome activation, and also interacts with Flightless-I, a pseudosubstrate of caspase-1, via its Coil motif. Knockdown of Flightless-I significantly promotes NLRP3 inflammasome activation. LRRFIP2 enhances the interaction between Flightless-I and caspase-1, facilitating the inhibitory effect of Flightless-I on caspase-1 activation. Furthermore, silencing of Flightless-I abrogates the inhibitory effect of LRRFIP2 on NLRP3 inflammasome. These data demonstrate that LRRFIP2 inhibits NLRP3 inflammasome activation by recruiting the caspase-1 inhibitor Flightless-I, thus outlining a new mechanism for negative regulation of NLRP3 inflammasome.

Show MeSH
Related in: MedlinePlus