Gas exchange measurements of carboxysome mutants reveal insights into cyanobacterial carbon-concentrating mechanisms.

Gas exchange measurements are widely used in plant research to quantitatively resolve photosynthetic performance and CO2 assimilation rates, yet gas exchange techniques are less commonly applied to algae and cyanobacteria. Cyanobacteria have a well-characterized carbon-concentrating mechanism (CCM) that includes a proteinaceous microcompartment called the carboxysome. Carboxysome structural components have been extensively studied and consist of a nucleated core of rubisco encapsulated by a diverse family of structurally related shell proteins. In addition to the dominant shell proteins that compose the majority of carboxysome facets, accessory shell components are widely distributed throughout cyanobacterial phyla. Herein, we utilize gas exchange techniques to elucidate the photosynthetic impacts of carboxysome mutants in Synechoccocus elongatus PCC 7942, emphasizing analysis of three accessory shell proteins: CcmK3, CcmK4, and CcmP. We detail technical considerations important for increasing the throughput and accuracy of gas exchange measurements using a recently developed commercial aquatic chamber. We find distinct photosynthetic phenotypes associated with the roles of different accessory shell proteins, with context-specific photosynthetic impairment in ΔccmP mutants that contrast with the broader pleiotropic defects of ΔccmK3 ΔccmK4 strains under multiple environments. Our results have implications for understanding the role of accessory carboxysome shell components in facilitating carbon fixation and other carboxysomal functions.