Cell identity and filament architecture shape intercellular communication in the terminal heterocystous cyanobacterium Richelia rhizosoleniae SC01

ABSTRACT Intercellular communication is essential for metabolite exchange and cellular coordination in filamentous cyanobacteria, yet how filament organization influences this process remains poorly understood, particularly in terminal heterocyst-forming strains. Here, we compared the facultative symbiont Richelia rhizosoleniae SC01 (hereafter Richelia ; terminal heterocysts) with the free-living Anabaena sp. PCC 7120 (hereafter Anabaena sp.; intercalary heterocysts) and a septal junction mutant of Anabaena with impaired intercellular communication (CSVT22, ∆ fraC -∆ fraD ), using fluorescence recovery after photobleaching (FRAP) and fluorescence loss in photobleaching (FLIP). Richelia SC01 and Anabaena showed similar recovery ( R ) rates, indicating comparable average molecular exchange through septal junctions, with no significant correlation between R and filament length. By contrast, CSVT22 showed reduced recovery and accelerated, irregular decay, indicative of both disrupted septal connectivity and altered regulation of septal junctions, resulting in heterogeneous and less efficient intercellular exchange. Differences in fluorescence decay speed across cell types and strains suggest that cell identity and filament organization, including filament taper, shape patterns of intercellular exchange. FLIP analyses showed that Anabaena consistently exhibited greater exchange asymmetry, whereas Richelia SC01 vegetative cells displayed comparatively more symmetric exchange, despite pronounced filament tapering and terminal heterocyst organization. Transmission electron microscopy further showed that septal nanopores in Richelia SC01 are similar in size to those of Anabaena but exhibited marked variation in number per septal disk, suggesting a potential structural basis for variation in intercellular exchange patterns. Collectively, our findings show that combining FRAP and FLIP provides a quantitative framework to dissect the kinetics and spatial organization of intercellular exchange in filamentous cyanobacteria. IMPORTANCE Filamentous cyanobacteria rely on intercellular exchange through septal junctions to coordinate metabolism and development, yet the principles governing multicellular communication in bacterial filaments remain poorly understood. Here, we show that intercellular exchange emerges from the interplay between septal junction properties, cell identity, and filament architecture, generating distinct communication patterns across cyanobacterial lifestyles. Notably, despite its pronounced filament tapering and terminal heterocyst organization, Richelia SC01 exhibited comparatively symmetric exchange, demonstrating that filament polarity alone does not determine communication patterns. Our findings reveal that cyanobacterial multicellularity arises from the integration of structural organization and cell-specific properties, providing a framework for understanding coordinated physiology in filamentous cyanobacteria, including symbiotic and nitrogen-fixing systems.