Periodic Reporting for period 4 - SYMBIOSES (Novel Symbioses)
Berichtszeitraum: 2025-07-01 bis 2025-12-31
Cells can acquire fundamentally new properties by incorporating other cells as endosymbionts. This process has shaped the evolution of life in profound ways, the origins of mitochondria and chloroplasts being the most prominent examples. Yet the early events that enabled such extant associations to form and stabilise cannot be observed in retrospect. SYMBIOSES addressed this by developing nanotechnology to physically introduce bacteria into eukaryotic host cells and observe the emergence of endosymbiotic relationships under controlled conditions. Over the course of the project, novel endosymbioses were studied experimentally, vertical transmission and endosymbiont-mediated biosynthesis were demonstrated, host adaptation to the fitness costs of endosymbiosis was characterised, and FluidFM was established as a broadly applicable platform spanning organelle biology and synthetic endosymbiosis.
Over the course of the project, advances were made on two interconnected fronts: nanotechnology development and experimental endosymbiosis biology. On the technology side, FluidFM, which combines atomic force microscopy, optical microscopy, and nanofluidics, was developed into a versatile platform for single-cell manipulation. Mitochondria were transplanted between living mammalian cells: the aspirated organelles fused with the host mitochondrial network and maintained functionality across generations, opening new directions in organelle physiology and laying groundwork for therapeutic applications. A further technological advance was Live-seq, a method for extracting cytoplasmic RNA from a living cell without destroying it, enabling temporal transcriptomic recording at single-cell resolution. FluidFM was also specifically adapted for fungal cells, which possess a rigid cell wall, establishing a method for injection into and extraction from individual cells across yeasts and filamentous fungi. Building on these foundations, bacteria were implanted directly into the cytoplasm of an early-diverging filamentous fungus. Bacterial strains differed markedly in behaviour: some proliferated rapidly and became entrapped, while others achieved vertical transmission and enabled endosymbiont-mediated biosynthesis of a natural product. Endosymbiosis imposed fitness costs on the host that were offset through positive selection and host adaptation, allowing the stabilising dynamics of a nascent endosymbiosis to be characterised experimentally. In addition, a free-living soil bacterium was used to probe the earliest phase of endosymbiosis formation. Despite an initial antagonistic phase marked by compromised host fitness and immune activation, vertical transmission and repeated selective passaging led to transcriptional relaxation of host defence responses, documenting a shift from antagonism to commensalism. Host responses to bacterial implantation were also studied in enteroids, providing insight into how mammalian host cells respond to intruding bacteria.
SYMBIOSES established an experimental platform for studying endosymbiosis in a prospective manner. Live-seq makes longitudinal transcriptomic tracking of individual living cells possible, changing what can be asked about how cells respond to stimuli over time. Organelle transplantation between living cells opens a new approach to studying organelle physiology and has potential therapeutic relevance. The experimental endosymbiosis system makes it possible to study the conditions under which symbioses stabilise or collapse, with direct implications for understanding intracellular pathogens and for the rational design of beneficial symbioses for biotechnological applications.