scholarly journals Structural basis for arginine glycosylation of host substrates by bacterial effector proteins

2018 ◽  
Vol 9 (1) ◽  
Author(s):  
Jun Bae Park ◽  
Young Hun Kim ◽  
Youngki Yoo ◽  
Juyeon Kim ◽  
Sung-Hoon Jun ◽  
...  
Keyword(s):  
Effector Proteins ◽  
Structural Basis ◽  
Bacterial Effector Proteins

scholarly journals Host Pathways Important for Coxiella burnetii Infection Revealed by Genome-Wide RNA Interference Screening

mBio ◽  
2013 ◽  
Vol 4 (1) ◽  
Author(s):  
Justin A. McDonough ◽  
Hayley J. Newton ◽  
Scott Klum ◽  
Rachel Swiss ◽  
Hervé Agaisse ◽  
...  
Keyword(s):  
Coxiella Burnetii ◽  
Secretion System ◽  
Effector Proteins ◽  
Host Factors ◽  
Content Type ◽  
Host Determinants ◽  
Genome Wide ◽  
Type Ivb Secretion ◽  
Type Ivb Secretion System ◽  
Bacterial Effector Proteins

ABSTRACTCoxiella burnetiiis an intracellular pathogen that replicates within a lysosome-like vacuole. A Dot/Icm type IVB secretion system is used byC. burnetiito translocate effector proteins into the host cytosol that likely modulate host factor function. To identify host determinants required forC. burnetiiintracellular growth, a genome-wide screen was performed using gene silencing by small interfering RNA (siRNA). Replication ofC. burnetiiwas measured by immunofluorescence microscopy in siRNA-transfected HeLa cells. Newly identified host factors included components of the retromer complex, which mediates cargo cycling between the endocytic pathway and the Golgi apparatus. Reducing the levels of the retromer cargo-adapter VPS26-VPS29-VPS35 complex or retromer-associated sorting nexins abrogatedC. burnetiireplication. Several genes, when silenced, resulted in enlarged vacuoles or an increased number of vacuoles withinC. burnetii-infected cells. Silencing of theSTX17gene encoding syntaxin-17 resulted in a striking defect in homotypic fusion of vacuoles containingC. burnetii, suggesting a role for syntaxin-17 in regulating this process. Lastly, silencing host genes needed forC. burnetiireplication correlated with defects in the translocation of Dot/Icm effectors, whereas, silencing of genes that affected vacuole morphology, but did not impact replication, did not affect Dot/Icm translocation. These data demonstrate thatC. burnetiivacuole maturation is important for creating a niche that permits Dot/Icm function. Thus, genome-wide screening has revealed host determinants involved in sequential events that occur duringC. burnetiiinfection as defined by bacterial uptake, vacuole transport and acidification, activation of the Dot/Icm system, homotypic fusion of vacuoles, and intracellular replication.IMPORTANCEQ fever in humans is caused by the bacteriumCoxiella burnetii. Infection withC. burnetiiis marked by its unique ability to replicate within a large vacuolar compartment inside cells that resembles the harsh, acidic environment of a lysosome. Central to its pathogenesis is the delivery of bacterial effector proteins into the host cell cytosol by a Dot/Icm type IVB secretion system. These proteins can interact with and manipulate host factors, thereby leading to creation and maintenance of the vacuole that the bacteria grow within. Using high-throughput genome-wide screening in human cells, we identified host factors important for several facets ofC. burnetiiinfection, including vacuole transport and membrane fusion events that promote vacuole expansion. In addition, we show that maturation of theC. burnetiivacuole is necessary for creating an environment permissive for the Dot/Icm delivery of bacterial effector proteins into the host cytosol.

scholarly journals Structural basis for mRNA recognition by human RBM38

2020 ◽  
Vol 477 (1) ◽  
pp. 161-172 ◽  
Author(s):  
Kaiyue Qian ◽  
Mengyu Li ◽  
Junchao Wang ◽  
Min Zhang ◽  
Mingzhu Wang
Keyword(s):  
Rna Binding ◽  
Structural Information ◽  
Protein Translation ◽  
Effector Proteins ◽  
Structural Basis ◽  
Binding Property ◽  
Rna Recognition ◽  
Recognition Mechanism ◽  
Rrm Domain ◽  
Regulate Protein

RNA-binding protein RBM38 was reported to bind the mRNA of several p53-related genes through its RRM domain and to up-regulate or down-regulate protein translation by increasing mRNA stability or recruitment of other effector proteins. The recognition mechanism, however, for RNA-binding of RBM38 remains unclear. Here, we report the crystal structure of the RRM domain of human RBM38 in complex with a single-stranded RNA. Our structural and biological results revealed that RBM38 recognizes G(U/C/A)GUG sequence single-stranded RNA in a sequence-specific and structure-specific manner. Two phenylalanine stacked with bases of RNA were crucial for RNA binding, and a series of hydrogen bonds between the base atoms of RNA and main-chain or side-chain atoms of RBM38 determine the sequence-specific recognition. Our results revealed the RNA-recognition mechanism of human RBM38 and provided structural information for understanding the RNA-binding property of RBM38.

Structural basis of ubiquitin recognition by a bacterial OTU deubiquitinase LotA

2021 ◽  
Author(s):  
Norihiro Takekawa ◽  
Tomoko Kubori ◽  
Tomoya Iwai ◽  
Hiroki Nagai ◽  
Katsumi Imada
Keyword(s):  
Enzymatic Activity ◽  
Legionella Pneumophila ◽  
Pathogenic Bacteria ◽  
Catalytic Domain ◽  
Docking Simulation ◽  
Effector Proteins ◽  
Host Cells ◽  
Structural Basis ◽  
Ubiquitin Chain ◽  
Lota Lota

Pathogenic bacteria have acquired a vast array of eukaryotic-like proteins via intimate interaction with host cells. Bacterial effector proteins that function as ubiquitin ligases and deubiquitinases (DUBs) are remarkable examples of such molecular mimicry. LotA, a Legionella pneumophila effector, belongs to the ovarian tumor (OTU) superfamily, which regulates diverse ubiquitin signals by their DUB activities. LotA harbors two OTU domains that have distinct reactivities; the first one is responsible for the cleavage of the K6-linked ubiquitin chain, and the second one shows an uncommon preference for long chains of ubiquitin. Here, we report the crystal structure of a middle domain of LotA (LotA M ), which contains the second OTU domain. LotA M consists of two distinct subdomains, a catalytic domain having high structural similarity with human OTU DUBs and an extended helical lobe (EHL) domain, which is characteristically conserved only in Legionella OTU DUBs. The docking simulation of LotA M with ubiquitin suggested that hydrophobic and electrostatic interactions between the EHL of LotA M and the C-terminal region of ubiquitin are crucial for the binding of ubiquitin to LotA M . The structure-based mutagenesis demonstrated that the acidic residue in the characteristic short helical segment termed the ‘helical arm’ is essential for the enzymatic activity of LotA M . The EHL domain of the three Legionella OTU DUBs, LotA, LotB, and LotC, share the ‘helical arm’ structure, suggesting that the EHL domain defines the Lot-OTUs as a unique class of DUBs. Importance To successfully colonize, some pathogenic bacteria hijack the host ubiquitin system. Legionella OTU-like-DUBs (Lot-DUBs) are novel bacterial deubiquitinases found in effector proteins of L. pneumophila . LotA is a member of Lot-DUBs and has two OTU domains (OTU1 and OTU2). We determined the structure of a middle fragment of LotA (LotA M ), which includes OTU2. LotA M consists of the conserved catalytic domain and the Legionella OTUs-specific EHL domain. The docking simulation with ubiquitin and the mutational analysis suggested that the acidic surface in the EHL is essential for enzymatic activity. The structure of the EHL differs from those of other Lot-DUBs, suggesting that the variation of the EHL is related to the variable cleaving specificity of each DUB.

scholarly journals Effector Protein Cig2 Decreases Host Tolerance of Infection by Directing Constitutive Fusion of Autophagosomes with the Coxiella -Containing Vacuole

mBio ◽  
2016 ◽  
Vol 7 (4) ◽  
Author(s):  
Lara J. Kohler ◽  
Shawna R. Reed ◽  
Shireen A. Sarraf ◽  
David D. Arteaga ◽  
Hayley J. Newton ◽  
...  
Keyword(s):  
Coxiella Burnetii ◽  
Host Response ◽  
Galleria Mellonella ◽  
Secretion System ◽  
Effector Proteins ◽  
Host Cells ◽  
Effector Protein ◽  
Autophagy Pathway ◽  
Host Tolerance ◽  
Bacterial Effector Proteins

ABSTRACT Coxiella burnetii replicates in an acidified lysosome-derived vacuole. Biogenesis of the Coxiella -containing vacuole (CCV) requires bacterial effector proteins delivered into host cells by the Dot/Icm secretion system. Genetic and cell biological analysis revealed that an effector protein called Cig2 promotes constitutive fusion of autophagosomes with the CCV to maintain this compartment in an autolysosomal stage of maturation. This distinguishes the CCV from other pathogen-containing vacuoles that are targeted by the host autophagy pathway, which typically confers host resistance to infection by delivering the pathogen to a toxic lysosomal environment. By maintaining the CCV in an autolysosomal stage of maturation, Cig2 enabled CCV homotypic fusion and enhanced bacterial virulence in the Galleria mellonella (wax moth) model of infection by a mechanism that decreases host tolerance. Thus, C. burnetii residence in an autolysosomal organelle alters host tolerance of infection, which indicates that Cig2-dependent manipulation of a lysosome-derived vacuole influences the host response to infection. IMPORTANCE Coxiella burnetii is an obligate, intracellular bacterial pathogen that replicates inside a unique, lysosome-like compartment called the Coxiella -containing vacuole (CCV). Over 130 bacterial effector proteins are delivered into the host cell cytosol by the C. burnetii Dot/Icm type IV secretion system. Although the Dot/Icm system is essential for pathogenesis, the functions of most effectors remain unknown. Here we show that the effector protein Cig2 is essential for converting the CCV to an organelle that is similar to the autolysosome. Cig2 function promotes constitutive fusion between the CCV and autophagosomes generated by selective autophagy. Cig2-directed biogenesis of an autolysosomal vacuole is essential for the unique fusogenic properties of the CCV and for virulence in an animal model of disease. This work highlights how bacterial subversion of the host autophagy pathway can influence the cell biological properties of the CCV and influence the host response to infection.

scholarly journals Antibodies to synthetic peptide from the residue 33 to 42 domain of c-Ha-ras p21 block reconstitution of the protein with different effectors.

1989 ◽  
Vol 9 (9) ◽  
pp. 3904-3910 ◽  
Author(s):  
I Rey ◽  
P Soubigou ◽  
L Debussche ◽  
C David ◽  
A Morgat ◽  
...  
Keyword(s):  
Synthetic Peptide ◽  
Binding Pocket ◽  
Inhibitory Effect ◽  
Effector Proteins ◽  
Structural Basis ◽  
Ras Proteins ◽  
Ras P21 ◽  
Reduced Rate ◽  
Additional Support

Residues 32 to 40, which are conserved among ras proteins from different species, are likely to participate in interactions with the p21 effector system. With the goal of understanding the structural basis of the regulatory functions of c-Ha-ras p21, we produced rabbit antisera against a synthetic peptide corresponding to amino acids 33 to 42 of the protein. The affinity-purified antibodies interacted specifically with p21 and with the antigenic peptide. The epitope recognized by the antibodies appeared to be centered on threonine 35. The antibodies inhibited both in vitro p21-induced production of cyclic AMP in detergent extracts of RAS-defective yeast membranes and GAP-stimulated GTPase activity. However, monoclonal anti-ras antibodies Y13-259 and Y13-238 were not capable of specifically inhibiting interactions of p21 with these two putative effector proteins. The apparent inhibitory effect of Y13-259 on stimulation of p21 by GAP was due to a greatly reduced rate of exchange of nucleotides in the binding pocket of the protein. These findings provide additional support for the essential role of the residue 32 to 40 domain as the true effector site and further evidence of the involvement of GAP as a cellular effector of ras proteins.

scholarly journals Structural Profiling of Bacterial Effectors Reveals Enrichment of Host-Interacting Domains and Motifs

2021 ◽  
Vol 8 ◽  
Author(s):  
Yangchun Frank Chen ◽  
Yu Xia
Keyword(s):  
Protein Interactions ◽  
Effector Proteins ◽  
Host Cells ◽  
Structural Basis ◽  
Protein Protein Interactions ◽  
Specific Domain ◽  
Bacterial Proteins ◽  
Domain Interactions ◽  
Host Signaling ◽  
Bacterial Effectors

Effector proteins are bacterial virulence factors secreted directly into host cells and, through extensive interactions with host proteins, rewire host signaling pathways to the advantage of the pathogen. Despite the crucial role of globular domains as mediators of protein-protein interactions (PPIs), previous structural studies of bacterial effectors are primarily focused on individual domains, rather than domain-mediated PPIs, which limits their ability to uncover systems-level molecular recognition principles governing host-bacteria interactions. Here, we took an interaction-centric approach and systematically examined the potential of structural components within bacterial proteins to engage in or target eukaryote-specific domain-domain interactions (DDIs). Our results indicate that: 1) effectors are about six times as likely as non-effectors to contain host-like domains that mediate DDIs exclusively in eukaryotes; 2) the average domain in effectors is about seven times as likely as that in non-effectors to co-occur with DDI partners in eukaryotes rather than in bacteria; and 3) effectors are about nine times as likely as non-effectors to contain bacteria-exclusive domains that target host domains mediating DDIs exclusively in eukaryotes. Moreover, in the absence of host-like domains or among pathogen proteins without domain assignment, effectors harbor a higher variety and density of short linear motifs targeting host domains that mediate DDIs exclusively in eukaryotes. Our study lends novel quantitative insight into the structural basis of effector-induced perturbation of host-endogenous PPIs and may aid in the design of selective inhibitors of host-pathogen interactions.

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