Metabolic diversity in the chemoautotrophic Riftia pachyptila symbiont

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Along hydrothermal vent areas of the Eastern Pacific, the giant deep sea tube worm Riftia pachyptila lives in a highly specialized monospecific symbiosis with chemoautotrophic bacteria. While adult worms lack a digestive system, their sulfide-oxidizing endosymbionts fix CO2 from the hydrothermal environment and thus supply the host with organic carbon. In turn, the bacteria benefit from high and steady substrate concentrations inside the worm. The metagenome of the uncultured symbionts was recently sequenced and provides the basis for in-depth genomic and metabolic analyses. Previous global proteomic profiling revealed a variety of alternative metabolic pathways that were simultaneously expressed in the symbiont population of the same host, but which appear to fulfill redundant reactions, or which might even function in opposing directions. These results strongly suggest the existence of individual bacterial subpopulations inside the bacteria-containing host tissue, the trophosome. Within the trophosome lobules, the intracellular symbionts undergo a differentiation process: They develop from small, dividing rods to small cocci, and finally to large cocci. These individual morphotypes very likely coincide with different metabolic strategies and might explain the observed metabolic diversity. The proposed project will address this hypothesis in three subtasks: (I) Morphotype enrichment: To verify whether the bacterial cell cycle stages represent metabolically distinct symbiont subpopulations, the respective morphotypes will be enriched by gradient centrifugation and subjected to a comprehensive and comparative proteomic analysis. (II) Protein localization: Additionally, key enzymes involved in potentially redundant or antagonistic pathways will be localized in trophosome lobule sections by immunohistochemistry. (III) Energy limitation: To examine the symbionts response to energy limitation and to identify potential differences between limitation-specific metabolic patterns of the individual subpopulations, a comparative proteomic analysis will be performed. Enriched morphotype fractions from sulfur-rich (energy-rich) trophosome tissue and fractions from sulfur-depleted (energy-depleted) trophosomes will be compared. A whole-tissue quantitative proteomic analysis of sulfur-rich and of sulfur-depleted trophosome will complement the study.