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Carboxysomes are the specific CO2-fixing microcompartments in all cyanobacteria. Although it is known that the organization and subcellular localization of carboxysomes are dependent on external light conditions and are highly relevant to their functions, how carboxysome organization and function are actively orchestrated in natural diurnal cycles has remained elusive. Here, we explore the dynamic regulation of carboxysome positioning and carbon fixation in the model cyanobacterium Synechococcus elongatus PCC 7942 in response to diurnal light-dark cycles, using live-cell confocal imaging and Rubisco assays. We found that carboxysomes are prone to locate close to the central line along the short axis of the cell and exhibit a greater preference of polar distribution in the dark phase, coupled with a reduction in carbon fixation. Moreover, we show that deleting the gene encoding the circadian clock protein KaiA could lead to an increase in carboxysome numbers per cell and reduced portions of pole-located carboxysomes. Our study provides insight into the diurnal regulation of carbon fixation in cyanobacteria and the general cellular strategies of cyanobacteria living in natural habitat for environmental acclimation. [ABSTRACT FROM AUTHOR]

Carboxysomes are proteinaceous bacterial microcompartments that sequester the key enzymes for carbon fixation in cyanobacteria and some proteobacteria. They consist of a virus-like icosahedral shell, encapsulating several enzymes, including ribulose 1,5-bisphosphate carboxylase/oxygenase (RuBisCO), responsible for the first step of the Calvin-Benson-Bassham cycle. Despite their significance in carbon fixation and great bioengineering potentials, the structural understanding of native carboxysomes is currently limited to low-resolution studies. Here, we report the characterization of a native α-carboxysome from a marine cyanobacterium by single-particle cryoelectron microscopy (cryo-EM). We have determined the structure of its RuBisCO enzyme, and obtained low-resolution maps of its icosahedral shell, and of its concentric interior organization. Using integrative modeling approaches, we have proposed a complete atomic model of an intact carboxysome, providing insight into its organization and assembly. This is critical for a better understanding of the carbon fixation mechanism and toward repurposing carboxysomes in synthetic biology for biotechnological applications. [Display omitted] • Carboxysomes are prokaryotic microcompartments that sequester photosynthetic enzymes • We used single-particle cryo-EM analysis to characterize intact carboxysome complexes • We have proposed a model of how enzymes are arranged within carboxysomes • We also have generated a structural model of its shell architecture Carboxysomes are proteinaceous microcompartments found in cyanobacteria, which facilitate carbon fixation, the first step of photosynthesis, by sequestering the corresponding enzymes. Evans et al. used cryoelectron microscopy to characterize intact carboxysomes, which allowed them to establish a structural model for the carboxysome shell and for its internal enzyme organization. [ABSTRACT FROM AUTHOR]