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The Coxiella burnetii Dot/Icm system delivers a unique repertoire of type IV effectors into host cells and is required for intracellular replication.

Carey KL, Newton HJ, Lührmann A, Roy CR - PLoS Pathog. (2011)

Bottom Line: The identification of a transposon insertion mutation that disrupts the dot/icm locus was used to validate that this apparatus was essential for translocation of effectors.Importantly, this C. burnetii Dot/Icm-deficient mutant was found to be defective for intracellular replication.Thus, these data indicate that C. burnetii encodes a unique subset of bacterial effector proteins translocated into host cells by the Dot/Icm apparatus, and that the cumulative activities exerted by these effectors enables C. burnetii to successfully establish a niche inside mammalian cells that supports intracellular replication.

View Article: PubMed Central - PubMed

Affiliation: Section of Microbial Pathogenesis, Yale University School of Medicine, New Haven, Connecticut, United States of America.

ABSTRACT
Coxiella burnetii, the causative agent of human Q fever, is an intracellular pathogen that replicates in an acidified vacuole derived from the host lysosomal network. This pathogen encodes a Dot/Icm type IV secretion system that delivers bacterial proteins called effectors to the host cytosol. To identify new effector proteins, the functionally analogous Legionella pneumophila Dot/Icm system was used in a genetic screen to identify fragments of C. burnetii genomic DNA that when fused to an adenylate cyclase reporter were capable of directing Dot/Icm-dependent translocation of the fusion protein into mammalian host cells. This screen identified Dot/Icm effectors that were proteins unique to C. burnetii, having no overall sequence homology with L. pneumophila Dot/Icm effectors. A comparison of C. burnetii genome sequences from different isolates revealed diversity in the size and distribution of the genes encoding many of these effectors. Studies examining the localization and function of effectors in eukaryotic cells provided evidence that several of these proteins have an affinity for specific host organelles and can disrupt cellular functions. The identification of a transposon insertion mutation that disrupts the dot/icm locus was used to validate that this apparatus was essential for translocation of effectors. Importantly, this C. burnetii Dot/Icm-deficient mutant was found to be defective for intracellular replication. Thus, these data indicate that C. burnetii encodes a unique subset of bacterial effector proteins translocated into host cells by the Dot/Icm apparatus, and that the cumulative activities exerted by these effectors enables C. burnetii to successfully establish a niche inside mammalian cells that supports intracellular replication.

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Domain analysis and genomic comparisons for C. burnetii effectors identified in this study.The schematic shows the eighteen NM effectors identified in this study represented in blue. Below each NM effector is a representation of the size of the homologous reading frame encoded by the genes in the other sequenced strains of C. burnetii. The size of the predicted proteins is represented in kDa in brackets following the gene designation. Black lines represent the presence of homologous DNA that does not encode an open reading frame due to small deletions (vertical black line) and stop codons (red cross). In cases where multiple deletions occur the first deletion, representing the site of the frameshift, is displayed. The locations of conserved domains in each protein identified in a SMART database search are indicated according to the key provided.
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ppat-1002056-g002: Domain analysis and genomic comparisons for C. burnetii effectors identified in this study.The schematic shows the eighteen NM effectors identified in this study represented in blue. Below each NM effector is a representation of the size of the homologous reading frame encoded by the genes in the other sequenced strains of C. burnetii. The size of the predicted proteins is represented in kDa in brackets following the gene designation. Black lines represent the presence of homologous DNA that does not encode an open reading frame due to small deletions (vertical black line) and stop codons (red cross). In cases where multiple deletions occur the first deletion, representing the site of the frameshift, is displayed. The locations of conserved domains in each protein identified in a SMART database search are indicated according to the key provided.

Mentions: Sequence analysis revealed that none of the 18 C. burnetii Dot/Icm effectors have significant homology to proteins found in other organisms, demonstrating the unique nature of these proteins. CBU0077 from the strain NM RSA493 is highly conserved in the two different sequenced C. burnetii strains isolated from chronic Q-fever patients presenting with endocarditis (G Q212 and K Q154) and in the Dugway strain isolated from rodents [30], which does not appear to cause clinical disease. For the remaining effectors, there were polymorphisms in the coding regions that would indicate that these effectors have either been mutated or have not been acquired by one or more of the strains of C. burnetii. An alignment of these effectors in all sequenced strains is presented in Figure 2. A comparison of the NM genome to the genomes of the G, K and Dugway strains revealed nucleotide deletions or nonsense mutations in the NM genes that would predict the production of a shorter version of several effector proteins compared to homologues found in some of these other sequenced strains [31]. Interestingly, three of the NM effectors identified here, CBU1108, CBU1107 and CBU1776, are not present in the G Q212 strain (Figure 2). Thus, similar to Dot/Icm effectors in L. pneumophila, there is significant plasticity observed in the repertoire of effectors of C. burnetii when genomes from different isolates are compared [32]. Despite these polymorphisms, all of the genes encoding the NM Dot/Icm effectors identified here were transcribed during infection (Figure S4). Thus, the genes encoding truncated NM effectors are transcribed and the predicted translated products should be delivered into host cells during infection by the functional Dot/Icm translocation signal identified in the protein.


The Coxiella burnetii Dot/Icm system delivers a unique repertoire of type IV effectors into host cells and is required for intracellular replication.

Carey KL, Newton HJ, Lührmann A, Roy CR - PLoS Pathog. (2011)

Domain analysis and genomic comparisons for C. burnetii effectors identified in this study.The schematic shows the eighteen NM effectors identified in this study represented in blue. Below each NM effector is a representation of the size of the homologous reading frame encoded by the genes in the other sequenced strains of C. burnetii. The size of the predicted proteins is represented in kDa in brackets following the gene designation. Black lines represent the presence of homologous DNA that does not encode an open reading frame due to small deletions (vertical black line) and stop codons (red cross). In cases where multiple deletions occur the first deletion, representing the site of the frameshift, is displayed. The locations of conserved domains in each protein identified in a SMART database search are indicated according to the key provided.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3102713&req=5

ppat-1002056-g002: Domain analysis and genomic comparisons for C. burnetii effectors identified in this study.The schematic shows the eighteen NM effectors identified in this study represented in blue. Below each NM effector is a representation of the size of the homologous reading frame encoded by the genes in the other sequenced strains of C. burnetii. The size of the predicted proteins is represented in kDa in brackets following the gene designation. Black lines represent the presence of homologous DNA that does not encode an open reading frame due to small deletions (vertical black line) and stop codons (red cross). In cases where multiple deletions occur the first deletion, representing the site of the frameshift, is displayed. The locations of conserved domains in each protein identified in a SMART database search are indicated according to the key provided.
Mentions: Sequence analysis revealed that none of the 18 C. burnetii Dot/Icm effectors have significant homology to proteins found in other organisms, demonstrating the unique nature of these proteins. CBU0077 from the strain NM RSA493 is highly conserved in the two different sequenced C. burnetii strains isolated from chronic Q-fever patients presenting with endocarditis (G Q212 and K Q154) and in the Dugway strain isolated from rodents [30], which does not appear to cause clinical disease. For the remaining effectors, there were polymorphisms in the coding regions that would indicate that these effectors have either been mutated or have not been acquired by one or more of the strains of C. burnetii. An alignment of these effectors in all sequenced strains is presented in Figure 2. A comparison of the NM genome to the genomes of the G, K and Dugway strains revealed nucleotide deletions or nonsense mutations in the NM genes that would predict the production of a shorter version of several effector proteins compared to homologues found in some of these other sequenced strains [31]. Interestingly, three of the NM effectors identified here, CBU1108, CBU1107 and CBU1776, are not present in the G Q212 strain (Figure 2). Thus, similar to Dot/Icm effectors in L. pneumophila, there is significant plasticity observed in the repertoire of effectors of C. burnetii when genomes from different isolates are compared [32]. Despite these polymorphisms, all of the genes encoding the NM Dot/Icm effectors identified here were transcribed during infection (Figure S4). Thus, the genes encoding truncated NM effectors are transcribed and the predicted translated products should be delivered into host cells during infection by the functional Dot/Icm translocation signal identified in the protein.

Bottom Line: The identification of a transposon insertion mutation that disrupts the dot/icm locus was used to validate that this apparatus was essential for translocation of effectors.Importantly, this C. burnetii Dot/Icm-deficient mutant was found to be defective for intracellular replication.Thus, these data indicate that C. burnetii encodes a unique subset of bacterial effector proteins translocated into host cells by the Dot/Icm apparatus, and that the cumulative activities exerted by these effectors enables C. burnetii to successfully establish a niche inside mammalian cells that supports intracellular replication.

View Article: PubMed Central - PubMed

Affiliation: Section of Microbial Pathogenesis, Yale University School of Medicine, New Haven, Connecticut, United States of America.

ABSTRACT
Coxiella burnetii, the causative agent of human Q fever, is an intracellular pathogen that replicates in an acidified vacuole derived from the host lysosomal network. This pathogen encodes a Dot/Icm type IV secretion system that delivers bacterial proteins called effectors to the host cytosol. To identify new effector proteins, the functionally analogous Legionella pneumophila Dot/Icm system was used in a genetic screen to identify fragments of C. burnetii genomic DNA that when fused to an adenylate cyclase reporter were capable of directing Dot/Icm-dependent translocation of the fusion protein into mammalian host cells. This screen identified Dot/Icm effectors that were proteins unique to C. burnetii, having no overall sequence homology with L. pneumophila Dot/Icm effectors. A comparison of C. burnetii genome sequences from different isolates revealed diversity in the size and distribution of the genes encoding many of these effectors. Studies examining the localization and function of effectors in eukaryotic cells provided evidence that several of these proteins have an affinity for specific host organelles and can disrupt cellular functions. The identification of a transposon insertion mutation that disrupts the dot/icm locus was used to validate that this apparatus was essential for translocation of effectors. Importantly, this C. burnetii Dot/Icm-deficient mutant was found to be defective for intracellular replication. Thus, these data indicate that C. burnetii encodes a unique subset of bacterial effector proteins translocated into host cells by the Dot/Icm apparatus, and that the cumulative activities exerted by these effectors enables C. burnetii to successfully establish a niche inside mammalian cells that supports intracellular replication.

Show MeSH
Related in: MedlinePlus