Structural basis for the recruitment and activation of the
            <i>Legionella</i>
            phospholipase VipD by the host GTPase Rab5

Lucas, María; Gaspar, Andrew H.; Pallara, Chiara; Rojas, Adriana L.; Fernández‐Recio, Juan; Machner, Matthias P.; Hierro, Aitor

doi:10.1073/pnas.1405391111

Cited by 47 publications

(55 citation statements)

References 54 publications

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“…Several L. pneumophila effectors have been found to have enzymatic activities that are regulated through autoinhibition. The GEF activity of RalF is autoinhibited by a C-terminal domain that functions as a cap that senses host membranes (48)(49)(50), and VipD is a phospholipase that is autoinhibited by an amino-terminal domain that binds to the host protein Rab5 (51)(52)(53). Thus, future studies on the mechanism by which SidJ regulates SidE function could reveal additional strategies by which the activities of L. pneumophila effectors are controlled spatially and temporally during infection.…”

Section: Discussionmentioning

confidence: 99%

Toxicity and SidJ-Mediated Suppression of Toxicity Require Distinct Regions in the SidE Family of Legionella pneumophila Effectors

Havey

Roy

2015

Infect Immun

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Intracellular bacteria use a variety of strategies to evade degradation and create a replicative niche. Legionella pneumophila is an intravacuolar pathogen that establishes a replicative niche through the secretion of more than 300 effector proteins. The function of most effectors remains to be determined. Toxicity in yeast has been used to identify functional domains and elucidate the biochemical function of effectors. A library of L. pneumophila effectors was screened using an expression plasmid that produces low levels of each protein. This screen identified the effector SdeA as a protein that confers a strong toxic phenotype that inhibits yeast replication. The toxicity of SdeA was suppressed in cells producing the effector SidJ. The effector SdeA is a member of the SidE family of L. pneumophila effector proteins. All SidE orthologs encoded by the Philadelphia isolate of Legionella pneumophila were toxic to yeast, and SidJ suppressed the toxicity of each. We identified a conserved central region in the SidE proteins that was sufficient to mediate yeast toxicity. Surprisingly, SidJ did not suppress toxicity when this central region was produced in yeast. We determined that the amino-terminal region of SidE was essential for SidJ-mediated suppression of toxicity. Thus, there is a genetic interaction that links the activity of SidJ and the amino-terminal region of SidE, which is required to modulate the toxic activity displayed by the central region of the SidE protein. This suggests a complex mechanism by which the L. pneumophila effector SidJ modulates the function of the SidE proteins after translocation into host cells.L egionella pneumophila is a Gram-negative, facultative intravacuolar pathogen that is the causative agent of Legionnaires' disease in humans (1-3). L. pneumophila is phagocytosed by host cells and creates a vacuole that supports replication by inhibiting the fusion of host endocytic vesicles and subverting the transport of secretory vesicles (4-7). The bacterially encoded Dot/Icm type IV secretion system (T4SS) is essential for the creation of this Legionella-containing vacuole (LCV) (8, 9). Proteins secreted by the T4SS, known as effectors, have been shown to both decorate the LCV and modify the location and function of host proteins, especially Rab GTPases (8,10,11). Determining the function of these effector proteins has been complicated by the observation that deletion of a single effector or groups of effectors does not typically result in a strong replication defect within host cells (12)(13)(14).The use of the yeast Saccharomyces cerevisiae as a system to study L. pneumophila effector proteins has been successful in previous studies (15)(16)(17)(18)(19). Examples include studies on the effector protein RalF, which was shown to inhibit yeast replication when overproduced, and the effector proteins YlfA and YlfB were both identified in an early yeast toxicity screen (16). Effector toxicity in yeast has also been used to study effector activities. The L. pneumophila effector AnkX is capa...

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Section: Discussionmentioning

confidence: 99%

Toxicity and SidJ-Mediated Suppression of Toxicity Require Distinct Regions in the SidE Family of Legionella pneumophila Effectors

Havey

Roy

2015

Infect Immun

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“…LpnE binds phosphatidylinositol 3-phosphate, is required for invasion and the establishment of an infection in macrophages, amoebae as well as A/J mice (117, 118) (supplemental Table S11). PE-on Dot/Icm effectors included RalF, a guanine nucleotide exchange factor activating ARF on LCVs (119); VipD, a phospholipase A which blocks endosome fusion with LCVs (120, 121); AnkX interfering with fusion of the LCVs with late endosomes (122,123); RavZ inhibiting autophagy during infection (124); and SidH which is negatively regulated by another effector protein, the E3 ubiquitin ligase LubX (112, 125) (supplemental Table S11). …”

Section: Overview Of L Pneumophila E and Pe Soluble Whole Cellmentioning

confidence: 99%

Life Stage-specific Proteomes of Legionella pneumophila Reveal a Highly Differential Abundance of Virulence-associated Dot/Icm effectors

Auraß

Gerlach

Becher

et al. 2016

Molecular & Cellular Proteomics

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Major differences in the transcriptional program underlying the phenotypic switch between exponential and post-exponential growth of Legionella pneumophila were formerly described characterizing important alterations in infection capacity. Additionally, a third state is known where the bacteria transform in a viable but nonculturable state under stress, such as starvation. We here describe phase-related proteomic changes in exponential phase (E), postexponential phase (PE) bacteria, and unculturable microcosms (UNC) containing viable but nonculturable state cells, and identify phasespecific proteins. We present data on different bacterial subproteomes of E and PE, such as soluble whole cell proteins, outer membrane-associated proteins, and extracellular proteins. In total, 1368 different proteins were identified, 922 were quantified and 397 showed differential abundance in E/PE. The quantified subproteomes of soluble whole cell proteins, outer membrane-associated proteins, and extracellular proteins; 841, 55, and 77 proteins, respectively, were visualized in Voronoi treemaps. 95 proteins were quantified exclusively in E, such as cell division proteins MreC, FtsN, FtsA, and ZipA; 33 exclusively in PE, such as motility-related proteins of flagellum biogenesis FlgE, FlgK, and FliA; and 9 exclusively in unculturable microcosms soluble whole cell proteins, such as hypothetical, as well as transport/binding-, and metabolism-related proteins. A high frequency of differentially abundant or phase-exclusive proteins was observed among the 91 quantified effectors of the major virulence-associated protein secretion system Dot/Icm (> 60%). 24 were E-exclusive, such as LepA/B, YlfA, MavG, Lpg2271, and 13 were PE-exclusive, such as RalF, VipD, Lem10. The growth phase-related specific abundance of a subset of Dot/Icm virulence effectors was confirmed by means of Western blotting. We therefore conclude that many effectors are predominantly abundant at either E or PE which suggests their phase specific function. The distinct temporal or spatial presence of such proteins might have important implications for functional assignments in the future or for use as lifestage specific markers for pathogen analysis. Molecular

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“…VipD, VpdA, VdpB, and VpdC are translocated by the Dot/Icm type IVB secretion system into the host cell (164)(165)(166). Once in the cytosol, VipD binds Rab5 and Rab22 (167,168), two key regulators of endosomal trafficking from early endosomes, which induces a structural rearrangement that triggers VipD's PLA 1 activity, resulting in inhibition of phagosomal maturation by catalyzing PIP 3 depletion and altering the protein composition of endosomes (169). Furthermore, VipD hydrolyzes PE and PC on the mitochondrial membrane, contributing to cytochrome c release, caspase 3 activation, and apoptosis induction (166).…”

Section: And Pi 45-biphosphate [Pi(45)p 2 ] (141)mentioning

confidence: 99%

Bacterial Sphingomyelinases and Phospholipases as Virulence Factors

Flores-Dı́az

Monturiol-Gross

Naylor

et al. 2016

Microbiol Mol Biol Rev

165

143

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SUMMARYBacterial sphingomyelinases and phospholipases are a heterogeneous group of esterases which are usually surface associated or secreted by a wide variety of Gram-positive and Gram-negative bacteria. These enzymes hydrolyze sphingomyelin and glycerophospholipids, respectively, generating products identical to the ones produced by eukaryotic enzymes which play crucial roles in distinct physiological processes, including membrane dynamics, cellular signaling, migration, growth, and death. Several bacterial sphingomyelinases and phospholipases are essential for virulence of extracellular, facultative, or obligate intracellular pathogens, as these enzymes contribute to phagosomal escape or phagosomal maturation avoidance, favoring tissue colonization, infection establishment and progression, or immune response evasion. This work presents a classification proposal for bacterial sphingomyelinases and phospholipases that considers not only their enzymatic activities but also their structural aspects. An overview of the main physiopathological activities is provided for each enzyme type, as are examples in which inactivation of a sphingomyelinase- or a phospholipase-encoding gene impairs the virulence of a pathogen. The identification of sphingomyelinases and phospholipases important for bacterial pathogenesis and the development of inhibitors for these enzymes could generate candidate vaccines and therapeutic agents, which will diminish the impacts of the associated human and animal diseases.

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Structural basis for the recruitment and activation of the Legionella phospholipase VipD by the host GTPase Rab5

Cited by 47 publications

References 54 publications

Toxicity and SidJ-Mediated Suppression of Toxicity Require Distinct Regions in the SidE Family of Legionella pneumophila Effectors

Toxicity and SidJ-Mediated Suppression of Toxicity Require Distinct Regions in the SidE Family of Legionella pneumophila Effectors

Life Stage-specific Proteomes of Legionella pneumophila Reveal a Highly Differential Abundance of Virulence-associated Dot/Icm effectors

Bacterial Sphingomyelinases and Phospholipases as Virulence Factors

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