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A Dynamic, Ring-Forming Bactofilin Critical for Maintaining Cell Size in the Obligate Intracellular Bacterium Chlamydia trachomatis.

ABSTRACT: Bactofilins are polymer-forming cytoskeletal proteins that are widely conserved in bacteria. Members of this protein family have diverse functional roles such as orienting subcellular molecular processes, establishing cell polarity, and aiding in cell shape maintenance. Using sequence alignment to the conserved bactofilin domain, we identified a bactofilin ortholog, BacA_CT, in the obligate intracellular pathogen Chlamydia trachomatis. Chlamydia species are obligate intracellular bacteria that undergo a developmental cycle alternating between infectious nondividing elementary bodies (EBs) and noninfectious dividing reticulate bodies (RBs). As Chlamydia divides by a polarized division process, we hypothesized that BacA_CT may function to establish polarity in these unique bacteria. Utilizing a combination of fusion constructs and high-resolution fluorescence microscopy, we determined that BacA_CT forms dynamic, membrane-associated filament- and ring-like structures in Chlamydia's replicative RB form. Contrary to our hypothesis, these structures are distinct from the microbe's cell division machinery and do not colocalize with septal peptidoglycan or MreB, the major organizer of the bacterium's division complex. Bacterial two-hybrid assays demonstrated BacA_CT interacts homotypically but does not directly interact with proteins involved in cell division or peptidoglycan biosynthesis. To investigate the function of BacA_CT in chlamydial development, we constructed a conditional knockdown strain using a newly developed CRISPR interference system. We observed that reducing bacA_CT expression significantly increased chlamydial cell size. Normal RB morphology was restored when an additional copy of bacA_CT was expressed in trans during knockdown. These data reveal a novel function for chlamydial bactofilin in maintaining cell size in this obligate intracellular bacterium.

SUBMITTER: Brockett MR

PROVIDER: S-EPMC8281209 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BACKGROUND: The obligate intracellular growing bacterium Chlamydia trachomatis causes diseases like trachoma, urogenital infection and lymphogranuloma venereum with severe morbidity. Several serovars and genotypes have been identified, but these could not be linked to clinical disease or outcome. The related Chlamydophila pneumoniae, of which no subtypes are recognized, causes respiratory infections worldwide. We developed a multi locus sequence typing (MLST) scheme to understand the population genetic structure and diversity of these species and to evaluate the association between genotype and disease. RESULTS: A collection of 26 strains of C. trachomatis of different serovars and clinical presentation and 18 strains of C. pneumoniae were included in the study. For comparison, sequences of C. abortus, C. psittaci, C. caviae, C. felis, C. pecorum (Chlamydophila), C. muridarum (Chlamydia) and of Candidatus protochlamydia and Simkania negevensis were also included. Sequences of fragments (400 - 500 base pairs) from seven housekeeping genes (enoA, fumC, gatA, gidA, hemN, hlfX, oppA) were analysed. Analysis of allelic profiles by eBurst revealed three non-overlapping clonal complexes among the C. trachomatis strains, while the C. pneumoniae strains formed a single group. An UPGMA tree produced from the allelic profiles resulted in three groups of sequence types. The LGV strains grouped in a single cluster, while the urogenital strains were distributed over two separated groups, one consisted solely of strains with frequent occurring serovars (E, D and F). The distribution of the different serovars over the three groups was not consistent, suggesting exchange of serovar encoding ompA sequences. In one instance, exchange of fumC sequences between strains of different groups was observed. Cluster analyses of concatenated sequences of the Chlamydophila and Chlamydia species together with those of Candidatus Protochlamydia amoebophila and Simkania negevensis resulted in a tree identical to that obtained with 23S RNA gene sequences. CONCLUSION: These data show that C. trachomatis and C. pneumoniae are highly uniform. The difference in genetic diversity between C. trachomatis and C. pneumoniae is in concordance with a later assimilation to the human host of the latter. Our data supports the taxonomy of the order of Chlamydiales.

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A Dynamic, Ring-Forming Bactofilin Critical for Maintaining Cell Size in the Obligate Intracellular Bacterium Chlamydia trachomatis.

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