title: Intracellular TLR4/MD-2 in macrophages senses Gram-negative bacteria and induces a unique set of LPS-dependent genes.
abstract: Toll-like receptor (TLR)4/MD-2, a sensor for LPS, delivers the MyD88-dependent signal from the cell surface, then traffics to endolysosomes and delivers the TRIF/TICAM-1-dependent signal. Both signals are thought to be dependent on cell surface TLR4/MD-2. Although TLR4/MD-2 is located also in recycling endosomes, the Golgi apparatus or the endoplasmic reticulum, little is known about a role for intracellular TLR4/MD-2 in LPS responses. We here studied intracellular LPS sensing in macrophages. PRAT4A (protein associated with TLR4 A) is a cochaperone for a general chaperone gp96 and required for cell surface expression of TLR4/MD-2. Cell surface TLR4/MD-2 was undetectable on PRAT4A(-/-) thioglycollate-elicited peritoneal macrophages (P-Macs) and bone marrow-derived macrophages (BM-Macs). LPS responses were all abolished in PRAT4A(-/-) P-Macs, whereas a part of LPS responses remained detectable in PRAT4A(-/-) BM-Macs. Of note, LPS responses in PRAT4A(-/-) BM-Macs were not necessarily dependent on TRIF/TICAM-1 signaling. PRAT4A(-/-) BM-Macs showed unimpaired production of both TRIF/TICAM-1-dependent chemokine RANTES (CCL5) and MyD88-dependent chemokine MCP-1 (CCL2). Moreover, up-regulation of co-stimulatory molecules, CD40 and CD86 was not altered. In contrast, TRIF/TICAM-1-dependent production of type I IFN was profoundly impaired. In response to heat-killed bacteria Escherichia coli, BM-Macs also required PRAT4A-independent TLR4/MD-2 for production of MCP-1 (CCL2) and RANTES (CCL5) and for up-regulation of CD40 and CD86, indicating that intracellular TLR4/MD-2 is able to sense phagocytosed bacteria and activate immune responses. These results demonstrate that intracellular TLR4/MD-2 is responsible for unique set of LPS responses.
authors.affiliation: Division of Infectious Genetics, Department of Microbiology and Immunology, Institute of Medical Science, University of Tokyo, 4-6-1 Shirokanedai, Minatoku, Tokyo 108-8639, Japan.
authors.forename: Takuma
authors.initials: T
authors.lastname: Shibata
authors.forename: Yuji
authors.initials: Y
authors.lastname: Motoi
authors.forename: Natsuko
authors.initials: N
authors.lastname: Tanimura
authors.forename: Natsuko
authors.initials: N
authors.lastname: Yamakawa
authors.forename: Sachiko
authors.initials: S
authors.lastname: Akashi-Takamura
authors.forename: Kensuke
authors.initials: K
authors.lastname: Miyake
mesh_terms: D000818:Animals; D001773:Blood Cells / Q000276:immunology / Q000378:metabolism; D001854:Bone Marrow Cells / Q000276:immunology / Q000378:metabolism; D016207:Cytokines / Q000096:biosynthesis; D005786:Gene Expression Regulation / Q000187:drug effects / Q000276:immunology; D006090:Gram-Negative Bacteria / Q000276:immunology*; D042541:Intracellular Space / Q000276:immunology / Q000378:metabolism; D008070:Lipopolysaccharides / Q000276:immunology* / Q000494:pharmacology; D008264:Macrophages / Q000276:immunology* / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D051197:Toll-Like Receptor 4 / Q000235:genetics / Q000378:metabolism*; D015854:Up-Regulation / Q000235:genetics / Q000276:immunology
chemical_list: D016207:Cytokines; D008070:Lipopolysaccharides; D051197:Toll-Like Receptor 4