부산대학교 도서관

Autophagy is a key innate immune response to intracellular parasites that promotes their delivery to degradative lysosomes following detection in the cytosol or within damaged vacuoles. Like L isteria and S higella, which use specific mechanisms to avoid autophagic detection and capture, the bacterial pathogen F rancisella tularensis proliferates within the cytosol of macrophages without demonstrable control by autophagy. To examine how F rancisella evades autophagy, we screened a library of F . tularensis subsp. tularensis Schu S4 HimarFT transposon mutants in GFP- LC3-expressing murine macrophages by microscopy for clones localized within autophagic vacuoles after phagosomal escape. Eleven clones showed autophagic capture at 6 h post-infection, whose HimarFT insertions clustered to fourgenetic loci involved in lipopolysaccharidic and capsular O-antigen biosynthesis. Consistent with the HimarFT mutants, in-frame deletion mutants of two representative loci, FTT1236 and FTT1448c ( manC), lacking both LPS and capsular O-antigen, underwent phagosomal escape but were cleared from the host cytosol. Unlike wild-type F rancisella, the O-antigen deletion mutants were ubiquitinated, and recruited the autophagy adaptor p62/ SQSTM1 and LC3 prior to cytosolic clearance. Autophagy-deficient macrophages partially supported replication of both mutants, indicating that O-antigen-lacking F rancisella are controlled by autophagy. These data demonstrate the intracellular protective role of this bacterial surface polysaccharide against autophagy. [ABSTRACT FROM AUTHOR]