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ATP/ADP translocases: a common feature of obligate intracellular amoebal symbionts related to Chlamydiae and Rickettsiae.

ABSTRACT: ATP/ADP translocases catalyze the highly specific transport of ATP across a membrane in an exchange mode with ADP. Such unique transport proteins are employed by plant plastids and have among the prokaryotes so far only been identified in few obligate intracellular bacteria belonging to the Chlamydiales and the Rickettsiales. In this study, 12 phylogenetically diverse bacterial endosymbionts of free-living amoebae and paramecia were screened for the presence of genes encoding ATP/ADP transport proteins. The occurrence of ATP/ADP translocase genes was found to be restricted to endosymbionts related to rickettsiae and chlamydiae. We showed that the ATP/ADP transport protein of the Parachlamydia-related endosymbiont of Acanthamoeba sp. strain UWE25, a recently identified relative of the important human pathogens Chlamydia trachomatis and Chlamydophila pneumoniae, is functional when expressed in the heterologous host Escherichia coli and demonstrated the presence of transcripts during the chlamydial developmental cycle. These findings indicate that the interaction between Parachlamydia-related endosymbionts and their amoeba hosts concerns energy parasitism similar to the interaction between pathogenic chlamydiae and their human host cells. Phylogenetic analysis of all known ATP/ADP translocases indicated that the genes encoding ATP/ADP translocases originated from a chlamydial ancestor and were, after an ancient gene duplication, transferred horizontally to rickettsiae and plants.

SUBMITTER: Schmitz-Esser S

PROVIDER: S-EPMC321502 | biostudies-literature | 2004 Feb

REPOSITORIES: biostudies-literature

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ATP/ADP translocases: a common feature of obligate intracellular amoebal symbionts related to Chlamydiae and Rickettsiae.

Schmitz-Esser Stephan S Linka Nicole N Collingro Astrid A Beier Cora L CL Neuhaus H Ekkehard HE Wagner Michael M Horn Matthias M

Journal of bacteriology 20040201 3

ATP/ADP translocases catalyze the highly specific transport of ATP across a membrane in an exchange mode with ADP. Such unique transport proteins are employed by plant plastids and have among the prokaryotes so far only been identified in few obligate intracellular bacteria belonging to the Chlamydiales and the Rickettsiales. In this study, 12 phylogenetically diverse bacterial endosymbionts of free-living amoebae and paramecia were screened for the presence of genes encoding ATP/ADP transport p ...[more]

PMID: 14729693

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Project description:Energy parasitism by ATP/ADP transport proteins is an essential, common feature of intracellular bacteria such as chlamydiae and rickettsiae, which are major pathogens of humans. Although several ATP/ADP transport proteins have so far been characterized, some fundamental questions regarding their function remained unaddressed. In this study, we focused on the detailed biochemical analysis of a representative ATP/ADP transporter (PamNTT1), from the amoeba symbiont Protochlamydia amoebophila (UWE25) to further clarify the principle of energy exploitation. We succeeded in the purification of the first bacterial nucleotide transporter (NTT) and its functional reconstitution into artificial lipid vesicles. Reconstituted PamNTT1 revealed high import velocities for ATP and an unexpected and previously unobserved stimulating effect of the luminal ADP on nucleotide import affinities. Latter preference of the nucleotide hetero-exchange is independent of the membrane potential, and therefore, PamNTT1 not only structurally but also functionally differs from the well-characterized mitochondrial ADP/ATP carriers. Reconstituted PamNTT1 exhibits a bidirectional orientation in lipid vesicles, but interestingly, only carriers inserted with the N-terminus directed to the proteoliposomal interior are functional. The data presented here comprehensively explain the functional basis of how the intracellular P. amoebophila manages to exploit the energy pool of its host cell effectively by using the nucleotide transporter PamNTT1. This membrane protein mediates a preferred import of ATP, which is additionally stimulated by a high internal (bacterial) ADP/ATP ratio, and the orientation-dependent functionality of the transporter ensures that it is not working in a mode that is detrimental to P. amoebophila. Heterologous expression and purification of high amounts of PamNTT1 provides the basis for its crystallization and detailed structure/function analyses. Furthermore, functional reconstitution of this essential chlamydial protein paves the way for high-throughput uptake studies in order to screen for specific inhibitors potentially suitable as anti-chlamydial drugs.

Project description:Gene transcription in bacteria is carried out by the multisubunit RNA polymerase (RNAP), which is composed of a catalytic core enzyme and a promoter-recognizing σ factor. The core enzyme comprises two α subunits, one β subunit, one β' subunit, and one ω subunit. The ω subunit plays critical roles in the assembly of the core enzyme and other cellular functions, including the regulation of bacterial growth, the stress response, and biofilm formation. However, the identity of an ω subunit for the obligate intracellular bacterium Chlamydia has not previously been determined. Here, we report the identification of the hypothetical protein CTL0286 as the probable chlamydial ω subunit based on sequence, synteny, and AlphaFold and AlphaFold-Multimer three-dimensional-structure predictions. Our findings indicate that CTL0286 functions as the missing ω subunit of chlamydial RNAP. Our extended analysis also indicates that all obligate intracellular bacteria have ω orthologs. IMPORTANCE Chlamydiae are obligate intracellular bacteria that replicate only inside eukaryotic cells. Previously, it has not been possible to identify a candidate gene encoding the chlamydial RNA polymerase ω subunit, and it has been hypothesized that the chlamydial RNA polymerase ω subunit was lost in the evolutionary process through which Chlamydiae reduced their genome size and proteome sizes to adapt to an obligate intracellular lifestyle. Here, we report the identification of the chlamydial RNA polymerase ω subunit, based on conserved sequence, conserved synteny, AlphaFold-predicted conserved three-dimensional structure, and AlfaFold-Multimer-predicted conserved interactions. Our identification of the previously elusive chlamydial RNA polymerase ω subunit sets the stage for investigation of its roles in regulation of gene expression during chlamydial growth, development, and stress responses, and sets the stage for preparation and study of the intact chlamydial RNA polymerase and its interactions with inhibitors.

Dataset Information

ATP/ADP translocases: a common feature of obligate intracellular amoebal symbionts related to Chlamydiae and Rickettsiae.

Publications

ATP/ADP translocases: a common feature of obligate intracellular amoebal symbionts related to Chlamydiae and Rickettsiae.

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