MIIM30011 Lecture 4 Bacterial membrane vesicles 2024 PDF

Bacterial membrane vesicles Maria Liaskos [email protected]  Overview of this lecture To understand what bacterial membrane vesicles are, what they contain and how they are made To understand the many functions of bacterial membrane vesic...

Bacterial membrane vesicles Maria Liaskos [email protected]  Overview of this lecture To understand what bacterial membrane vesicles are, what they contain and how they are made To understand the many functions of bacterial membrane vesicles that includes their contribution to mediating bacterial survival and pathogenesis in the host.  The type 0 secretion system (T0SS): Outer membrane vesicles released by Gram-negative bacteria Guerrero-Mandujano et. al., Traffic, Volume: 18, Issue: 7, Pages: 425-432, April 2017, DOI: (10.1111/tra.12488)  Bacteria release extracellular membrane vesicles Bacteria release extracellular vesicles, called Bacterial membrane vesicles (BMVs) BMVs are produced by all bacteria as part of their normal growth BMVs range from 20-300 nm in size Outer membrane vesicles (OMVs): Gram-negative bacteria Membrane vesicles (MVs): Gram-positive bacteria OMVs were first identified approximately 50 years ago in vitro Whereas MVs were identified 40 years later Most of our knowledge about BMVs is of OMVs Chaterjee & Das, 1967, J Gen Micro Vibrio cholerae OMVs OMVs were suggested to have a role in mediating disease OMVs were detected in various patient samples and multiple sites in vivo: – eg. serum of patients suffering bacteraemia – cerebral spinal fluid of patients with meningitis – stomach of patients infected with Helicobacter pylori OMVs were therefore suggested to be associated with mediating disease in the host Research within the past decade has clearly established that OMVs produced by Gram negative bacterial pathogens contribute to: Mediating disease in the host Promoting bacterial survival in a range of environments  OMVs produced by Gram-negative bacteria contain a range of biological cargo from their parent organism OMVs contain many of contents found in the parent bacterium Membrane: Lipopolysaccharide, proteins, peptidoglycan Contents: toxins, DNA, RNA and periplasmic proteins Due to their cargo, OMVs are highly immunostimulatory Licensed for use in humans (Neisseria) This cargo also allows OMVs to have many functions: Horizontal gene transfer Modulate immunity Antimicrobial functions Kaparakis-Liaskos and Ferrero. Nat Rev Immunol 2015 Bacterial outer membrane vesicles: mechanisms of production.  Biogenesis of OMVs OMVs are produced by Gram negative bacteria as part of their normal growth In liquid culture, solid culture, biofilms and in vivo Mechanism of OMV production OMVs were initially identified to be produced by ”blebbing” from the bacterial cell surface We now know there are 3 main mechanisms of OMV production Mechanisms of OMV Biogenesis #1. Blebbing Roier et. al., Nature Comm 2016  Production of Gram negative bacterial outer membrane vesicles (OMVs) 1) Blebbing 2) Bacterial lysis Roier et. al., Nature Comm 2016 Turnbull L. et. al., Nature Comm 2016  Mechanisms of OMV Biogenesis #2. Explosive cell lysis Turnbull L. et. al., Nature Comm 2016  Production of Gram negative bacterial outer membrane vesicles (OMVs) 1) Blebbing 2) Bacterial lysis 3) Membrane remodeling and integrity Roier et. al., Nature comm 2016 Turnbull L. et. al., Nature Comm 2016 H. pylori DtolB Turner et al Helicobacter 2015  Mechanisms of OMV Biogenesis #3. Membrane remodelling (Tol-Pal system) Tol-Pal proteins maintain bacterial cell membrane integrity The gene cluster is conserved across Gram-negative organisms Tol-Pal cluster is comprised of inner membrane proteins (TolA, TolQ, TolR), periplasmic protein (TolB) and an outer membrane peptidoglycan-associated protein (Pal) E. coli TolQ TolR TolA TolB Pal H. pylori 26695 hp1130 hp 1129 hp 1128 hp1127 hp1127 X X hp1126 hp1125 Turner L et. al. Helicobacter 2015  Production of Gram negative bacterial outer membrane vesicles (OMVs) 1) Blebbing 2) Bacterial lysis 3) Membrane remodeling and integrity Roier et. al., Nature Comm 2016 Turnbull L. et. al., Nature Comm 2016 H. pylori DtolB Turner et al Helicobacter 2015 What about bacterial membrane vesicles produced by Gram positive bacteria?  Bacterial membrane vesicles are also produced by Gram positive bacteria MV production by Gram-positive bacteria was first observed in 1990s (Dorward and Garon 1990) Staphylococcus aureus MVs identified in 2009 (Lee et al. 2009) MVs are produced by many Gram-positive bacteria Just beginning to understand the composition and functions of MVs Listeria monocytogenes Mycobacterium bovis S. aureus (MVs) Lee et al., 2009, Proteomics. Brown L. et. al., Nature Rev Micro 2019  Composition of MVs produced by Gram positive bacteria Gram-positive membrane vesicle (MV) Lipoproteins Cytoplasmic proteins & DNA enzymes RNA Peptidoglycan Brown L. et. al., Nature Rev Micro 2019 Bitto & Kaparakis-Liaskos, 2017. Int. J. Mol. Sci.  Mechanism of MV production by Gram positive bacteria MVs production by Gram positive bacteria was first identified in Bacillus subtilis in 2017 Expression of an endolysin encoded by a defective prophage triggers formation of MVs Endolysin weaken the peptidoglycan layer B. subtilis cells protrude cytoplasmic membrane material through holes in the peptidoglycan, which are then released as explosive MVs This process of MV production also occurs in other Gram positive bacteria Toyofuku M. et. al., Nature Rev Micro 2019 Summary: Mechanisms of BMV production by Gram negative and Gram positive bacteria Toyofuku M. et. al., Nature Rev Micro 2019 Bacteria produce a wide range of BMVs with varied cargo Toyofuku M, Schild S. Kaparakis-Liaskos M, Eberl L., Nature Reviews Microbiology 2023 Bacteria upregulate BMV production in response to stress Toyofuku M. et. al., Nature Rev Micro 2019  Functions of OMVs Pathogenesis mediated by bacterial outer membrane vesicles Outer membrane vesicles: a novel mechanism used by Gram negative bacteria to mediate NOD1 signalling H. pylori Nucleotide oligomerisation domain 1 OMVs (NOD1) detects Gram negative Shigella peptidoglycan (PG) GM-TriDAP NOD1 RIP2 Gram negative bacteria deliver PG into host epithelial cells via NF-kB invasion (Philpott et. al. J Immunol 2000) IL-8 hbDs a type 4 secretion system (Viala et al. Nat. Immunol 2004), or Outer membrane vesicles (OMVs) (Kaparakis et al. Cell. Micro 2010)  OMVs can enter human epithelial cells via multiple mechanisms to interact with NOD1 and be cleared by the host Entry TLR detection Intracellular detection and degradation Kaparakis-Liaskos et. al., Nature Rev Immunol 2015, 15(6): 375-87. Can OMVs activate other host pathogen recognition receptors (PRRs) to mediate inflammation and pathogenesis in the host?  The TLR family of bacterial pathogen recognition molecules (PRMs) Wilson, et. al., Bacterial Pathogenesis, a molecular approach  The TLR family of bacterial pathogen recognition molecules (PRMs) Wilson, et. al., Bacterial Pathogenesis, a molecular approach  Bacterial products recognised by pathogen-recognition molecules (PRM) Nod1/2 TLR9 Modified from Aderem & Ulevitch Nature 406: 782 Outer membrane vesicles can activate PRRs and modulate epithelial cell responses at the host mucosal surface. Stimulation of NODs and TLRs Activation of inflammasomes Immune modulation via sRNA delivery OMVs OMVs MVs OMVs TLR4 TLR2 Lipid raft TLR NLRP3 Autophagy Caspase-11 sRNA sRNA NOD1 Epithelial cells Submucosa Nucleus Caspase-1 Endosome NF-ĸB Macrophage ↓ Cytokine production Epithelial cell ↓ Neutrophil infiltration Submucosa IL-8 IL-18 IL-1β Gilmore W et. al., Molecular Immunology 2021 Pathogenesis mediated by OMVs: OMVs interact with multiple host pathogen recognition receptors to drive inflammation NOD1 Kaparakis-Liaskos et. al., Nature Rev Immunol 2015, 15(6): 375-87. What about MVs produced by Gram-positive bacteria? Can they mediate an inflammatory response in host epithelial cells? MVs produced by Gram positive bacteria contain cargo that can mediate inflammation and pathogenesis in the host Bacillus anthracis MVs contain anthrax toxin Mycobacterium tuberculosis MVs contain triacylated lipoprotein Brown L. et. al., Nature Rev Micro 2015  Summary: BMVs produced by Gram negative and Gram positive bacteria mediate inflammation and pathogenesis Toyofuku M, Schild S. Kaparakis-Liaskos M, Eberl L., Nature Reviews Microbiology 2023 Non-pathogenic functions of bacterial membrane vesicles? How do BMVs help bacteria?  Bacterial advantages conferred by MVs: MVs function as decoys, an energy source and to promote bacterial survival Acquire nutrients Degrade antibiotics and transfer resistance Remove toxic or misfolded proteins Energy source Decoys to protect bacteria Adapted from Schwechheimer et. al., Nature Rev Micro 2015, 13(10): 605-19  Lecture Outcomes To know that bacterial MVs are considered a novel secretion system used by all bacteria To know what bacterial MVs are, their structure, what they contain and how they are generated To understand that there are different subtypes of MVs produced by Gram positive and Gram negative bacteria with different membrane structures and cargo To know that environmental factors can increase BMV production To know that BMVs produced by both Gram negative and Gram positive bacteria can mediate inflammation in the host by activating host innate immune receptors (PRRs). To know that BMVs can deliver virulence determinants to and into host cells to promote pathogenesis. To know that BMVs also have non-pathogenic roles and confer a selective advantage to bacteria eg. promote bacterial survival  Further reading Toyofuku, M., Schild, S., Kaparakis-Liaskos, M. et al. Composition and functions of bacterial membrane vesicles. Nat Rev Microbiol 21, 415–430 (2023). doi.org/10.1038/s41579-023-00875-5 Toyofuku, M., Nomura, N. & Eberl, L. Types and origins of bacterial membrane vesicles. Nat. Rev. Microbiol. 17, 13–24 (2019). doi.org/10.1038/ s41579-018-0112-2 Brown L, Wolf JM, Prados-Rosales R, Casadevall A. Through the wall: extracellular vesicles in Gram- positive bacteria, mycobacteria and fungi. Nat Rev Microbiol. 2015 Oct;13(10):620-30. doi: 10.1038/nrmicro3480. Kaparakis-Liaskos M, Ferrero RL. Immune modulation by bacterial outer membrane vesicles. Nat Rev Immunol. 2015 Jun;15(6):375-87. doi: 10.1038/nri3837.