Molecular mechanisms of Streptococcus pneumoniae‐targeted autophagy via pneumolysin, Golgi‐resident Rab41, and Nedd4‐1‐mediated K63‐linked ubiquitination

Streptococcus pneumoniae is the most common causative agent of community‐acquired pneumonia and can penetrate epithelial barriers to enter the bloodstream and brain. We investigated intracellular fates of S. pneumoniae and found that the pathogen is entrapped by selective autophagy in pneumolysin‐ and ubiquitin‐p62‐LC3 cargo‐dependent manners. Importantly, following induction of autophagy, Rab41 was relocated from the Golgi apparatus to S. pneumoniae‐containing autophagic vesicles (PcAV), which were only formed in the presence of Rab41‐positive intact Golgi apparatuses. Moreover, subsequent localization and regulation of K48‐ and K63‐linked polyubiquitin chains in and on PcAV were clearly distinguishable from each other. Finally, we found that E3 ligase Nedd4‐1 was recruited to PcAV and played a pivotal role in K63‐linked polyubiquitin chain (K63Ub) generation on PcAV, promotion of PcAV formation, and elimination of intracellular S. pneumoniae. These findings suggest that Nedd4‐1‐mediated K63Ub deposition on PcAV acts as a scaffold for PcAV biogenesis and efficient elimination of host cell‐invaded pneumococci.

[1]  A. Blumenthal,et al.  Functions of the WNT Signaling Network in Shaping Host Responses to Infection , 2019, Front. Immunol..

[2]  T. Mak,et al.  Emerging roles of HECT‐type E3 ubiquitin ligases in autophagy regulation , 2019, Molecular oncology.

[3]  K. Kim,et al.  Heterogeneity in pneumolysin expression governs the fate of Streptococcus pneumoniae during blood-brain barrier trafficking , 2018, PLoS pathogens.

[4]  I. Dikic,et al.  Ubiquitin signaling and autophagy , 2017, The Journal of Biological Chemistry.

[5]  Wannian Yang,et al.  The HECT E3 ubiquitin ligase NEDD4 interacts with and ubiquitylates SQSTM1 for inclusion body autophagy , 2017, Journal of Cell Science.

[6]  A. Ballabio,et al.  Molecular definitions of autophagy and related processes , 2017, The EMBO journal.

[7]  Wannian Yang,et al.  The E3 ubiquitin ligase NEDD4 is an LC3-interactive protein and regulates autophagy , 2017, Autophagy.

[8]  C. Rock,et al.  Pyruvate Oxidase as a Critical Link between Metabolism and Capsule Biosynthesis in Streptococcus pneumoniae , 2016, PLoS pathogens.

[9]  H. Harada,et al.  Golgi-Resident GTPase Rab30 Promotes the Biogenesis of Pathogen-Containing Autophagosomes , 2016, PloS one.

[10]  M. Ohnishi,et al.  Serotypes, antimicrobial susceptibility, and molecular epidemiology of invasive and non-invasive Streptococcus pneumoniae isolates in paediatric patients after the introduction of 13-valent conjugate vaccine in a nationwide surveillance study conducted in Japan in 2012-2014. , 2016, Vaccine.

[11]  S. Pyo,et al.  Streptococcus pneumoniae induces pyroptosis through the regulation of autophagy in murine microglia , 2015, Oncotarget.

[12]  V. Nizet,et al.  Streptococcal toxins: role in pathogenesis and disease , 2015, Cellular microbiology.

[13]  Yee‐Shin Lin,et al.  Insufficient Acidification of Autophagosomes Facilitates Group A Streptococcus Survival and Growth in Endothelial Cells , 2015, mBio.

[14]  E. Feeley,et al.  Ubiquitin systems mark pathogen-containing vacuoles as targets for host defense by guanylate binding proteins , 2015, Proceedings of the National Academy of Sciences.

[15]  G. Tonon,et al.  Che‐1‐induced inhibition of mTOR pathway enables stress‐induced autophagy , 2015, The EMBO journal.

[16]  Nektarios Tavernarakis,et al.  Unsaturated fatty acids induce non‐canonical autophagy , 2015, The EMBO journal.

[17]  C. Viret,et al.  Autophagy receptor NDP52 regulates pathogen-containing autophagosome maturation. , 2015, Cell host & microbe.

[18]  Pu Li,et al.  Streptococcus pneumoniae Induces Autophagy through the Inhibition of the PI3K-I/Akt/mTOR Pathway and ROS Hypergeneration in A549 Cells , 2015, PloS one.

[19]  A. Hakansson,et al.  Streptococcus pneumoniae biofilm formation and dispersion during colonization and disease , 2015, Front. Cell. Infect. Microbiol..

[20]  T. Yamaji,et al.  Establishment of HeLa Cell Mutants Deficient in Sphingolipid-Related Genes Using TALENs , 2014, PloS one.

[21]  S. Akira,et al.  Recruitment of the autophagic machinery to endosomes during infection is mediated by ubiquitin , 2013, The Journal of cell biology.

[22]  T. Mizushima,et al.  Phosphorylation of p62 activates the Keap1-Nrf2 pathway during selective autophagy. , 2013, Molecular cell.

[23]  J. Bijlsma,et al.  Streptococcus pneumoniae Invades Endothelial Host Cells via Multiple Pathways and Is Killed in a Lysosome Dependent Manner , 2013, PloS one.

[24]  M. Fukuda,et al.  Functional involvement of Rab1A in microtubule-dependent anterograde melanosome transport in melanocytes , 2012, Journal of Cell Science.

[25]  S. Hamada,et al.  The small GTPases Rab9A and Rab23 function at distinct steps in autophagy during Group A Streptococcus infection , 2012, Cellular microbiology.

[26]  M. Fukuda,et al.  Melanoregulin regulates retrograde melanosome transport through interaction with the RILP–p150Glued complex in melanocytes , 2012, Journal of Cell Science.

[27]  H. Stenmark,et al.  Nedd4-dependent lysine-11-linked polyubiquitination of the tumour suppressor Beclin 1 , 2011, The Biochemical journal.

[28]  C. Thompson,et al.  Ammonia-induced autophagy is independent of ULK1/ULK2 kinases , 2011, Proceedings of the National Academy of Sciences.

[29]  C. Sasakawa,et al.  A Tecpr1-dependent selective autophagy pathway targets bacterial pathogens. , 2011, Cell host & microbe.

[30]  Y. Koide,et al.  Rab GTPases Regulating Phagosome Maturation Are Differentially Recruited to Mycobacterial Phagosomes , 2011, Traffic.

[31]  T. Noda,et al.  Combinational Soluble N-Ethylmaleimide-sensitive Factor Attachment Protein Receptor Proteins VAMP8 and Vti1b Mediate Fusion of Antimicrobial and Canonical Autophagosomes with Lysosomes , 2010, Molecular biology of the cell.

[32]  Christian Rosenmund,et al.  Regulation of Rap2A by the Ubiquitin Ligase Nedd4-1 Controls Neurite Development , 2010, Neuron.

[33]  Takeshi Noda,et al.  An Initial Step of GAS-Containing Autophagosome-Like Vacuoles Formation Requires Rab7 , 2009, PLoS pathogens.

[34]  C. Sasakawa,et al.  Listeria monocytogenes ActA-mediated escape from autophagic recognition , 2009, Nature Cell Biology.

[35]  H. Izumiya,et al.  Application of lambda Red recombination system to Vibrio cholerae genetics: simple methods for inactivation and modification of chromosomal genes. , 2009, Gene.

[36]  S. Akira,et al.  Loss of the autophagy protein Atg16L1 enhances endotoxin-induced IL-1β production , 2008, Nature.

[37]  J. Guan,et al.  FIP200, a ULK-interacting protein, is required for autophagosome formation in mammalian cells , 2008, The Journal of cell biology.

[38]  Jeffrey N. Weiser,et al.  The role of Streptococcus pneumoniae virulence factors in host respiratory colonization and disease , 2008, Nature Reviews Microbiology.

[39]  Masaaki Komatsu,et al.  Homeostatic Levels of p62 Control Cytoplasmic Inclusion Body Formation in Autophagy-Deficient Mice , 2007, Cell.

[40]  C. Preisinger,et al.  Analysis of GTPase-activating proteins: Rab1 and Rab43 are key Rabs required to maintain a functional Golgi complex in human cells , 2007, Journal of Cell Science.

[41]  M. Lipsitch,et al.  SpxB Is a Suicide Gene of Streptococcus pneumoniae and Confers a Selective Advantage in an In Vivo Competitive Colonization Model , 2007, Journal of bacteriology.

[42]  K. Ishii,et al.  Key function for the Ubc13 E2 ubiquitin-conjugating enzyme in immune receptor signaling , 2006, Nature Immunology.

[43]  Mitsunori Fukuda,et al.  Rab3A and Rab27A cooperatively regulate the docking step of dense-core vesicle exocytosis in PC12 cells , 2006, Journal of Cell Science.

[44]  J. Swanson,et al.  Cytolysin‐dependent delay of vacuole maturation in macrophages infected with Listeria monocytogenes , 2006, Cellular microbiology.

[45]  Masaaki Komatsu,et al.  Impairment of starvation-induced and constitutive autophagy in Atg7-deficient mice , 2005, The Journal of cell biology.

[46]  Hiroshi Sagara,et al.  Escape of Intracellular Shigella from Autophagy , 2005, Science.

[47]  Takeshi Tokuhisa,et al.  The role of autophagy during the early neonatal starvation period , 2004, Nature.

[48]  S. Hamada,et al.  Autophagy Defends Cells Against Invading Group A Streptococcus , 2004, Science.

[49]  Y. Kadono,et al.  Segregation of TRAF6‐mediated signaling pathways clarifies its role in osteoclastogenesis , 2001, The EMBO journal.

[50]  Takeshi Noda,et al.  LC3, a mammalian homologue of yeast Apg8p, is localized in autophagosome membranes after processing , 2000, The EMBO journal.

[51]  E. Tuomanen,et al.  The Polymeric Immunoglobulin Receptor Translocates Pneumococci across Human Nasopharyngeal Epithelial Cells , 2000, Cell.

[52]  H. Roche,et al.  Unsaturated fatty acids , 1999, Proceedings of the Nutrition Society.

[53]  Joel A. Swanson,et al.  pH-dependent Perforation of Macrophage Phagosomes by Listeriolysin O from Listeria monocytogenes , 1997, The Journal of experimental medicine.

[54]  G. Pozzi,et al.  Competence for genetic transformation in encapsulated strains of Streptococcus pneumoniae: two allelic variants of the peptide pheromone , 1996, Journal of bacteriology.

[55]  I. Nakagawa,et al.  Rab35 GTPase recruits NDP52 to autophagy targets , 2017, The EMBO journal.

[56]  M. Fukuda,et al.  Large-scale Screening for Novel Rab Effectors Reveals Unexpected Broad Rab-binding Specificity* , 2008 .