Abstract
<title>Abstract</title> <p> Background Photosynthetic cyanobacteria entrapped in artificial solid matrices, such as alginate hydrogels, can sustain active photosynthesis and bio-production of chemicals for months, yet the cellular adaptations underlying this remarkable resilience remain poorly understood. Results Here, we compared the physiological and proteomic responses of the model cyanobacterium <italic>Synechocystis</italic> sp. PCC 6803 entrapped within thin calcium-alginate hydrogel films with those of suspension-grown cells. Immobilized cells maintained relatively stable photosystem II (PSII) photochemical efficiency over three weeks despite strongly restricted biomass accumulation. By contrast, suspension cultures exhibited a progressive decline in phycobilisome connectivity and PSII photochemical yield during prolonged cultivation. To elucidate the molecular adaptations associated with immobilization, we applied a comparative label-free proteomics, which revealed extensive and time-dependent proteome remodelling following immobilization. Proteins involved in photoprotection, alternative electron sinks and respiratory terminal oxidases progressively increased in abundance, indicating an enhanced capacity for excitation-energy dissipation and redox balancing. In parallel, ribosomal proteins, chaperones and Rubisco subunits broadly decreased in abundance, whereas the stringent-response regulator SpoT increased, suggesting a regulated downshift in growth-related metabolism and reallocation of cellular resources towards maintenance. Increased abundance of inorganic carbon uptake systems, cell-surface and pilus-associated proteins and toxin-antitoxin modules further indicated acclimation to spatial confinement, diffusion limitations, and high local cell density within the hydrogel matrix. Conclusion Collectively, these findings demonstrate that entrapment in thin-layer hydrogel induces a coordinated, maintenance-oriented physiological state that preserves photosynthetic activity under growth-limited conditions, consistent with a longevity phenotype. This state shares key features with natural cyanobacterial biofilms and enables photosynthetic living materials to function as robust biocatalysts for long-term bio-production. </p>