Project description:The endosymbiont population of the hydrothermal vent tube worm Riftia pachyptila consists of a single 16S phylotype of sulfur-oxidizing gammaproteobacteria. The intracellular symbiont exhibits remarkable morphological heterogeneity, from small rod-shaped or coccoid cells to large cocci, which were suggested to be part of a common cell cycle. To assess whether these morphological differences are accompanied by distinct metabolic profiles, we physically enriched individual symbiont cells sizes by density gradient centrifugation and subjected these enrichments to metaproteomic analysis and statistical evaluation using clustering and random forests. Unlike previous molecular studies, which examined the metabolism of the symbiont population as whole, we were thus able to unravel comprehensive protein abundance patterns of individual symbiont subpopulations. Supported by microscopic analyses, our metaproteomic results show that Riftia symbiont cells of different sizes are stages of a physiological differentiation process: Small symbionts actively divide and may establish cellular symbiont-host interaction, as indicated by highest abundance of the cell division key protein FtsZ and highly abundant chaperones and porins in this initial phase. We furthermore present evidence that large symbionts, on the other hand, do not divide, but still replicate DNA, leading to DNA endoreduplication. Highest abundance of enzymes for CO2 fixation, carbon storage and biosynthesis indicates that in its late differentiation stage, the symbiont’s metabolism is efficiently geared on the production of organic material. We propose that this symbiont aging process enhances the productivity of the symbiosis as a whole.
Project description:The deep-sea tubeworm Riftia pachyptila is a model system for a mutualistic association: The adult worm has no digestive system, but completely relies on one phylotype of endosymbiotic chemosynthetic bacteria for nutrition. The bacteria, in turn, are provisioned by the host. Metabolism and physiology of this symbiosis, particularly of the uncultured symbiont, have been subject to various studies. Yet, how both partners interact on the molecular level remains largely unknown. To study these host-symbiont interactions in detail, we sequenced the R. pachyptila host transcriptome de novo, and conducted comprehensive metaproteomic comparisons of symbiont-containing and symbiont-free R. pachyptila tissues under energy-rich and energy-limiting conditions. Our results demonstrate that R. pachyptila invests a considerable part of its proteome to provision the symbionts with inorganic compounds. It acquires symbiont-derived biomass primarily by digesting parts of the symbiont population. The R. pachyptila immune system apparently not only protects the holobiont from pathogens, but is also involved in symbiont population control. The symbiont expresses a repertoire of proteins dedicated to communication with the host, including eukaryote-like proteins that may counteract phagocytosis. During energy limitation, i.e., when reduced sulfur compounds are lacking, the host apparently increases symbiont digestion. We show here an intricate network of interaction pathways that shapes the R. pachyptila holobiont. Together with the metabolic flexibility of the association under varying energy conditions, this probably forms the basis for the success of this tight association under the highly challenging deep-sea conditions.
Project description:For multidomain proteins, evolutionary changes may occur at the domain as well as the whole-protein level. An example is presented here, with suggestions for how such complicated relationships might be visualized. Earlier analysis of the Candidatus Maribeggiatoa str. Orange Guaymas (BOGUAY; Gammaproteobacteria) single-filament draft genome found evidence of gene exchange with the phylogenetically distant Cyanobacteria, particularly for sensory and signal transduction proteins. Because these are modular proteins, known to undergo frequent duplication, domain swapping, and horizontal gene transfer, a single domain was chosen for analysis. Recognition (REC) domains are short (~125 amino acids) and well conserved, simplifying sequence alignments and phylogenetic calculations. Over 100 of these were identified in the BOGUAY genome and found to have a wide range of inferred phylogenetic relationships. Two sets were chosen here for detailed study. One set of four BOGUAY ORFs has closest relatives among other Beggiatoaceae and Cyanobacteria. A second set of four has REC domains with more mixed affiliations, including other Beggiatoaceae, several sulfate-reducing Deltaproteobacteria and Firmicutes, magnetotactic Nitrospirae, one Shewanella and one Ferrimonas strain (both Gammaproteobacteria), and numerous Vibrio vulnificus and V. navarrensis strains (also Gammaproteobacteria). For an overview of the possible origins of the whole proteins and the surrounding genomic regions, color-coded BLASTP results were produced and displayed against cartoons showing protein domain structure of predicted genes. This is suggested as a visualization method for investigation of possible horizontally transferred regions, giving more detail than scans of DNA composition and codon usage but much faster than carrying out full phylogenetic analyses for multiple proteins. As expected, most of the predicted sensor histidine kinases investigated have two or more segments with distinct BLASTP affiliations. For the first set of BOGUAY ORFs, the flanking regions were also examined, and the results suggest they are embedded in genomic stretches with complex histories. An automated method of creating such visualizations could be generally useful; a wish list for its features is given.