A Screen of Coxiella burnetii Mutants Reveals Important Roles for Dot/Icm Effectors and Host Autophagy in Vacuole Biogenesis

PLoS Pathogens

Morayma Temoche-Diaz¹  Elizabeth L. Hartland¹  Emerson Crabill¹  Hayley J. Newton¹  Justin A. McDonough¹  Lara J. Kohler²  Craig R. Roy² 

[1] Department of Microbial Pathogenesis, Yale University School of Medicine, New Haven, Connecticut, United States of America;Department of Microbiology and Immunology, University of Melbourne, Parkville, Victoria, Australia

关键词: Coxiella burnetii;    Vacuoles;    Transposable elements;    Autophagic cell death;    Phenotypes;    Intracellular pathogens;    Lysosomes;    HeLa cells;   

DOI  :  10.1371/journal.ppat.1004286

学科分类：生物科学（综合）

来源: Public Library of Science

【 摘 要 】

Coxiella burnetii is an intracellular pathogen that replicates in a lysosome-derived vacuole. The molecular mechanisms used by this bacterium to create a pathogen-occupied vacuole remain largely unknown. Here, we conducted a visual screen on an arrayed library of C. burnetii NMII transposon insertion mutants to identify genes required for biogenesis of a mature Coxiella-containing vacuole (CCV). Mutants defective in Dot/Icm secretion system function or the PmrAB regulatory system were incapable of intracellular replication. Several mutants with intracellular growth defects were found to have insertions in genes encoding effector proteins translocated into host cells by the Dot/Icm system. These included mutants deficient in the effector proteins Cig57, CoxCC8 and Cbu1754. Mutants that had transposon insertions in genes important in central metabolism or encoding tRNA modification enzymes were identified based on the appearance filamentous bacteria intracellularly. Lastly, mutants that displayed a multi-vacuolar phenotype were identified. All of these mutants had a transposon insertion in the gene encoding the effector protein Cig2. Whereas vacuoles containing wild type C. burnetii displayed robust accumulation of the autophagosome protein LC3, the vacuoles formed by the cig2 mutant did not contain detectible amounts of LC3. Furthermore, interfering with host autophagy during infection by wild type C. burnetii resulted in a multi-vacuolar phenotype similar to that displayed by the cig2 mutant. Thus, a functional Cig2 protein is important for interactions between the CCV and host autophagosomes and this drives a process that enhances the fusogenic properties of this pathogen-occupied organelle.

【 授权许可】

CC BY