Common architectures in cyanobacteria Prochlorococcus cells visualized by X-ray diffraction imaging using X-ray free electron laser

Amane Kobayashi,Sachihiro Matsunaga,Takeshi Hirakawa,Mao Oide,Tomotaka Oroguchi,Yayoi Inui,Masayoshi Nakasako,Yuki Takayama,Masaki Yamamoto,Koji Okajima

doi:10.1038/s41598-021-83401-y

Amane Kobayashi, Sachihiro Matsunaga + Show 8 more

Open Access

https://doi.org/10.1038/s41598-021-83401-y

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Abstract

Visualization of intracellular structures and their spatial organization inside cells without any modification is essential to understand the mechanisms underlying the biological functions of cells. Here, we investigated the intracellular structure of cyanobacteria Prochlorococcus in the interphase by X-ray diffraction imaging using X-ray free-electron laser. A number of diffraction patterns from single cells smaller than 1 µm in size were collected with high signal-to-noise ratio with a resolution of up to 30 nm. From diffraction patterns, a set of electron density maps projected along the direction of the incident X-ray were retrieved with high reliability. The most characteristic structure found to be common among the cells was a C-shaped arrangement of 100-nm sized high-density spots, which surrounded a low-density area of 100 nm. Furthermore, a three-dimensional map reconstructed from the projection maps of individual cells was non-uniform, indicating the presence of common structures among cyanobacteria cells in the interphase. By referring to the fluorescent images for distributions of thylakoid membranes, nucleoids, and carboxysomes, we inferred and represented their spatial arrangements in the three-dimensional map. The arrangement allowed us to discuss the relevance of the intracellular organization to the biological functions of cyanobacteria.

Highlights

Visualization of intracellular structures and their spatial organization inside cells without any modification is essential to understand the mechanisms underlying the biological functions of cells
We describe intracellular geometrical characteristics in projection electron density maps obtained by X-ray free-electron laser (XFEL)-X-ray diffraction imaging (XDI) experiments (Fig. 1)
Distribution of three components, chlorophylls, deoxyribonucleic acid (DNA), and ribulose-1,5-bisphosphate carboxylase/oxygenase (RubisCO) molecules, in Prochlorococcus cells in the interphase were visualized by fluorescence light microscopy (Fig. 2A)

Summary

Introduction

Visualization of intracellular structures and their spatial organization inside cells without any modification is essential to understand the mechanisms underlying the biological functions of cells. We investigated the intracellular structure of cyanobacteria Prochlorococcus in the interphase by X-ray diffraction imaging using X-ray free-electron laser. If cytoplasm of cyanobacteria cell is enclosed completely by several layers of thylakoid membranes, cargos of vesicles would be secreted to exterior of cells through complicated pathways including membrane fusion with thylakoid membranes. Investigation of such biological activities requires structural data on the distribution of intracellular components of cyanobacteria. Since biological cells are composed of well-organized and fragile cellular components, any other imaging technique complementary to TEM is necessary to visualize structures of whole cells without sectioning and chemical labeling of specimens. Since cell structures are somewhat different even in the same phase in the cell cycle, the technique is required to provide a large number of cell images enough to elucidate any common structures among cells in a specific cell-cycle phase and to discuss heterogeneity

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