Toxin secretion and trafficking by Mycobacterium tuberculosis

AbstractThe tuberculosis necrotizing toxin (TNT) is the major cytotoxicity factor of Mycobacterium tuberculosis (Mtb) in macrophages. TNT is the C-terminal domain of the outer membrane protein CpnT and gains access to the cytosol to kill macrophages infected with Mtb. However, molecular mechanisms of TNT secretion and trafficking are largely unknown. A comprehensive analysis of the five type VII secretion systems of Mtb revealed that the ESX-4 system is required for export of CpnT and surface accessibility of TNT. Furthermore, the ESX-2 and ESX-4 systems are required for permeabilization of the phagosomal membrane in addition to the ESX-1 system. Thus, these three ESX systems need to act in concert to enable trafficking of TNT into the cytosol of Mtb-infected macrophages. These discoveries establish new molecular roles for the two previously uncharacterized type VII secretion systems ESX-2 and ESX-4 and reveal an intricate link between toxin secretion and phagosomal permeabilization by Mtb.

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Deciphering the Function of the Outer Membrane Protein OprD Homologue of Acinetobacter baumannii

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.06022-11 ◽

2012 ◽

Vol 56 (7) ◽

pp. 3826-3832 ◽

Cited By ~ 28

Author(s):

Manuella Catel-Ferreira ◽

Rony Nehmé ◽

Virginie Molle ◽

Jesús Aranda ◽

Emeline Bouffartigues ◽

...

Keyword(s):

Membrane Protein ◽

Acinetobacter Baumannii ◽

Outer Membrane ◽

Outer Membrane Protein ◽

Molecular Mechanisms ◽

Single Channel ◽

Resistance Mechanism ◽

Carbapenem Resistance ◽

Bacterial Survival ◽

Content Type

ABSTRACTThe increasing number of carbapenem-resistantAcinetobacter baumanniiisolates is a major cause for concern which restricts therapeutic options to treat severe infections caused by this emerging pathogen. To identify the molecular mechanisms involved in carbapenem resistance, we studied the contribution of an outer membrane protein homologue of thePseudomonas aeruginosaOprD porin. Suspected to be the preferred pathway of carbapenems inA. baumannii, theoprDhomologue gene was inactivated in strain ATCC 17978. Comparison of wild-type and mutant strains did not confirm the expected increased resistance to any antibiotic tested. OprD homologue sequence analysis revealed that this protein actually belongs to an OprD subgroup but is closer to theP. aeruginosaOprQ protein, with which it could share some functions, e.g., allowing bacterial survival under low-iron or -magnesium growth conditions or under poor oxygenation. We thus overexpressed and purified a recombinant OprD homologue protein to further examine its functional properties. As a specific channel, this porin presented rather low single-channel conductance, i.e., 28 pS in 1 M KCl, and was partially closed by micro- and millimolar concentrations of Fe3+and Mg2+, respectively, but not by imipenem and meropenem or basic amino acids. TheA. baumanniiOprD homologue is likely not involved in the carbapenem resistance mechanism, but as an OprQ-like protein, it could contribute to the adaptation of this bacterium to magnesium- and/or iron-depleted environments.

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The mating pair stabilization protein, TraN, of the F plasmid is an outer-membrane protein with two regions that are important for its function in conjugation

Microbiology ◽

10.1099/mic.0.28025-0 ◽

2005 ◽

Vol 151 (11) ◽

pp. 3527-3540 ◽

Cited By ~ 31

Author(s):

William A. Klimke ◽

Candace D. Rypien ◽

Barbara Klinger ◽

R. Alexander Kennedy ◽

J. Manuel Rodriguez-Maillard ◽

...

Keyword(s):

Membrane Protein ◽

Outer Membrane ◽

Outer Membrane Protein ◽

Recipient Cell ◽

Terminal Region ◽

Mating Pair ◽

Secretion Systems ◽

F Plasmid ◽

Type Iv ◽

Reducing Conditions

F plasmid TraN (602 aa, processed to 584 aa with 22 conserved cysteines), which is essential for F plasmid conjugation, is an outer-membrane protein involved in mating pair stabilization (MPS). Unlike R100 TraN, F TraN requires OmpA in the recipient cell for efficient MPS. The authors have identified three external loops (aa 172–187, 212–220 and 281–284) in the highly divergent region from aa 164 to aa 333 as candidates for interaction with OmpA. These loops were identified using both site-directed and random TnphoA/in mutagenesis to insert epitopes (31-aa or c-myc) into TraN and monitor their effect on sensitivity to external proteases and on mating ability. TraN is a hallmark protein of F-type IV secretion systems as demonstrated by blast searches of the databases. The C-terminal region is highly conserved and contains five of the six completely conserved cysteines. Mutation of these residues to serine demonstrated their importance in TraN function. TraN appears to require both intra- and intermolecular disulfide bond formation for its stability and structure as demonstrated by its instability in a dsbA mutant and its aberrant migration on SDS-polyacrylamide gels under non-reducing conditions or by cross-linking with bis(sulfosuccinimidyl)suberate (BS3). Thus, F TraN appears to have two domains: the N-terminal region is involved in OmpA interaction with OmpA during MPS; and the C-terminal region, which is rich in conserved cysteine residues, is essential for conjugation.

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Quantification and surface localization of the hemolysin A type 1 secretion system at the endogenous level and under conditions of overexpression

Applied and Environmental Microbiology ◽

10.1128/aem.01896-21 ◽

2021 ◽

Author(s):

Tobias Beer ◽

Sebastian Hänsch ◽

Klaus Pfeffer ◽

Sander H.J. Smits ◽

Stefanie Weidtkamp-Peters ◽

...

Keyword(s):

Escherichia Coli ◽

Membrane Protein ◽

Outer Membrane ◽

Outer Membrane Protein ◽

Extracellular Space ◽

Super Resolution ◽

Secretion Systems ◽

Endogenous Level ◽

Surface Localization ◽

Bacterial Secretion

Secretion systems are essential for Gram-negative bacteria as these nanomachineries allow a communication with the outside world by exporting proteins into the extracellular space or directly into the cytosol of a host cell. For example, type one secretion systems (T1SS) secrete a broad range of substrates across both membranes into the extracellular space. One well-known example is the hemolysin A (HlyA) T1SS from Escherichia coli (E. coli) , which consists of an ABC transporter (HlyB), a membrane fusion protein (HlyD), the outer membrane protein TolC and the substrate HlyA, a member of the family of RTX (repeats in toxins) toxins. Here, we determined the amount of TolC at the endogenous level (parental strain, UTI89) and under conditions of overexpression (T7 expression system, BL21(DE3)-BD). The overall amount of TolC was not influenced by the overexpression of the HlyBD complex. Moving one step further, we determined the localization of the HlyA T1SS by super-resolution microscopy. In contrast to other bacterial secretion systems, no polarization was observed with respect to endogenous or overexpression levels. Additionally, the cell growth and division cycle did not influence the polarization. Most importantly, the size of the observed T1SS clusters did not correlate with the recently proposed outer membrane islands. These data indicate that T1SS cluster at the outer membrane generating domains of so far not described identity. Importance Uropathogenic Escherichia coli (UPEC) strains cause about 110 million urinary tract infections each year worldwide representing a global burden to the healthcare system. UPEC secrete many virulence factors among these the TX toxin hemolysin A via a cognate T1SS into the extracellular space. In this study, we determined the endogenous copy number of the HlyA T1SS in UTI89 and analyzed the surface localization in BL21(DE3)-BD and UTI89, respectively. With approximately 800 copies of the T1SS in UTI89, this is one of the highest expressed bacterial secretion systems. Furthermore and in clear contrast to other secretion systems, no polarized surface localization was detected. Finally, quantitative analysis of the super-resolution data revealed that clusters of the HlyA T1SS are not related to the recently identified outer membrane protein islands. These data provide insights into the quantitative molecular architecture of the HlyA T1SS.

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BNIP3L/NIX-mediated mitophagy: molecular mechanisms and implications for human disease

Cell Death and Disease ◽

10.1038/s41419-021-04469-y ◽

2021 ◽

Vol 13 (1) ◽

Author(s):

Yue Li ◽

Wanqing Zheng ◽

Yangyang Lu ◽

Yanrong Zheng ◽

Ling Pan ◽

...

Keyword(s):

Membrane Protein ◽

Outer Membrane ◽

Outer Membrane Protein ◽

Neurological Disorders ◽

Human Disease ◽

Molecular Mechanisms ◽

Molecular Level ◽

Current Evidence ◽

Mitochondrial Outer Membrane ◽

Biological Functions

AbstractMitophagy is a highly conserved cellular process that maintains the mitochondrial quantity by eliminating dysfunctional or superfluous mitochondria through autophagy machinery. The mitochondrial outer membrane protein BNIP3L/Nix serves as a mitophagy receptor by recognizing autophagosomes. BNIP3L is initially known to clear the mitochondria during the development of reticulocytes. Recent studies indicated it also engages in a variety of physiological and pathological processes. In this review, we provide an overview of how BNIP3L induces mitophagy and discuss the biological functions of BNIP3L and its regulation at the molecular level. We further discuss current evidence indicating the involvement of BNIP3L-mediated mitophagy in human disease, particularly in cancer and neurological disorders.

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Molecular insights into type I secretion systems

Biological Chemistry ◽

10.1515/hsz-2013-0171 ◽

2013 ◽

Vol 394 (11) ◽

pp. 1371-1384 ◽

Cited By ~ 14

Author(s):

Michael H.H. Lenders ◽

Sven Reimann ◽

Sander H. J. Smits ◽

Lutz Schmitt

Keyword(s):

Membrane Protein ◽

Outer Membrane ◽

Outer Membrane Protein ◽

Atp Binding ◽

Type I ◽

Secretion Systems ◽

Inner Membrane ◽

Small Proteins ◽

Atp Binding Cassette Transporter ◽

Atp Binding Cassette

Abstract Type 1 secretion systems are one of the main machineries in Gram-negative bacteria involved in the secretion of a wide range of substrates from the cytoplasm across the inner and outer membrane in one step to the extracellular space. The range of substrates varies from small proteins up to large surface layer proteins of about 900 kDa. Most of the substrates have a non-cleavable C-terminal secretion signal and so-called GG repeats that are able to bind calcium ions. The translocator complex is composed of a trimeric outer membrane protein that provides a pore in the outer membrane. A multimeric membrane fusion protein spans the periplasm and forms a continuous channel connecting the outer membrane protein with a dimeric ATP-binding cassette transporter in the inner membrane. The ATP-binding cassette-transporter is thought to form a channel through the inner membrane and energizes the transport process. This review will provide a detailed view of the components of the translocator and will summarize structural as well as functional data.

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