photobleaching-enables-super-resolution-imaging-ftsz-ring

Photobleaching Enables Super-resolution Imaging of the FtsZ Ring in the Cyanobacterium <em>Prochlorococcus</em>

Super-resolution microscopy has been widely used to study protein interactions and subcellular structures in many organisms. In photosynthetic organisms, ...

Hong Kong University of Science and Technology

HKUST Shenzhen Research Institute

There are six known families of homing endonucleases, LAGLIDADG, GIY-YIG, HNH, His-Cys box, PD-(D/E)-XK, and EDxHD, which are characterized by their conserved residues. Previously, we discovered a novel homing endonuclease F-CphI encoded by ORF177 of cyanophage S-PM2. F-CphI does not resemble any characterized homing endonucleases. Instead, the C-terminus of F-CphI aligns well with the N-terminal catalytic domain of a Holliday junction DNA resolvase, phage T4 endonuclease VII (Endo VII).

F-CphI represents a new homing endonuclease family using the Endo VII catalytic motif.

The transcriptomic responses of bacteria to environmental stresses have been studied extensively, yet we know little about how the stressed cells respond to bacteriophage infection. Here, we conducted the first whole transcriptome sequencing (RNA-seq) study of stressed bacteria to phage infection by infecting the marine picocyanobacterium Prochlorococcus NATL2A with cyanomyovirus P-SSM2 under P limitation, a strong selective force in the oceans. Transcripts of the P-acquisition genes in the uninfected cells were enriched after P limitation, including the high-affinity phosphate-binding protein gene pstS. They were still enriched in the infected cells under P-limited conditions. In contrast, transcripts of adenosine triphosphate (ATP) synthase and ribosomal protein genes were depleted in the uninfected cells after P limitation but were enriched during phage infection of P-starved cells. Cyanophage P-SSM2 contains pstS, and pstS and its adjacent gene g247 of unknown function were the only phage genes that were enriched under P-limited conditions. We further found that the host pstS transcript number per cell decreased after infection, however, it was still much higher in the P-limited infected cells than that in the nutrient-replete cells. Moreover, phage pstS transcript number per cell was ∼ 20 times higher than the host copy, which may help maintain the host phosphate uptake rate during infection.

Transcriptomic response during phage infection of a marine cyanobacterium under phosphorus-limited conditions.

Marine chromophoric dissolved organic matter (CDOM) and its related fluorescent components (FDOM), which are widely distributed but highly photobleached in the surface ocean, are critical in regulating light attenuation in the ocean. However, the origins of marine FDOM are still under investigation. Here we show that cultured picocyanobacteria, Synechococcus and Prochlorococcus, release FDOM that closely match the typical fluorescent signals found in oceanic environments. Picocyanobacterial FDOM also shows comparable apparent fluorescent quantum yields and undergoes similar photo-degradation behaviour when compared with deep-ocean FDOM, further strengthening the similarity between them. Ultrahigh-resolution mass spectrometry (MS) and nuclear magnetic resonance spectroscopy reveal abundant nitrogen-containing compounds in Synechococcus DOM, which may originate from degradation products of the fluorescent phycobilin pigments. Given the importance of picocyanobacteria in the global carbon cycle, our results indicate that picocyanobacteria are likely to be important sources of marine autochthonous FDOM, which may accumulate in the deep ocean.

Picocyanobacteria and deep-ocean fluorescent dissolved organic matter share similar optical properties.

Superresolution imaging has revealed subcellular structures and protein interactions in many organisms. However, superresolution microscopy with lateral resolution better than 100 nm has not been achieved in photosynthetic cells due to the interference of a high-autofluorescence background. Here, we developed a photobleaching method to effectively reduce the autofluorescence of cyanobacterial and plant cells. We achieved lateral resolution of ~10 nm with stochastic optical reconstruction microscopy (STORM) in the sphere-shaped cyanobacterium  and the flowering plant  During the cell cycle of , we characterized the three-dimensional (3D) organization of the cell division protein FtsZ, which forms a ring structure at the division site and is important for cytokinesis of bacteria and chloroplasts. Although the FtsZ ring assembly process in rod-shaped bacteria has been studied extensively, it has rarely been studied in sphere-shaped bacteria. Similarly to rod-shaped bacteria, our results with  also showed the assembly of FtsZ clusters into incomplete rings and then complete rings during cell division. Differently from rod-shaped bacteria, the FtsZ ring diameter was not found to decrease during  cell division. We also discovered a novel double-Z-ring structure, which may be the Z rings of two daughter cells in a predivisional mother cell. Our results showed a quantitative picture of the  Z ring organization of sphere-shaped bacteria. Superresolution microscopy has not been widely used to study photosynthetic cells due to their high-autofluorescence background. Here, we developed a photobleaching method to reduce the autofluorescence of cyanobacteria and plant cells. After photobleaching, we performed superresolution imaging in the cyanobacterium  and the flowering plant  with ~10-nm resolution, which is the highest resolution in a photosynthetic cell. With this method, we characterized the 3D organization of the cell division protein FtsZ in  We found that the morphological variation of the FtsZ ring during cell division of the sphere-shaped cyanobacterium  is similar but not identical to that of rod-shaped bacteria. Our method might also be applicable to other photosynthetic organisms.

Three-Dimensional Superresolution Imaging of the FtsZ Ring during Cell Division of the Cyanobacterium .

 is the most abundant photosynthetic organism in the tropical and subtropical oceans. Here, we report the draft genome sequence of  XMU1401, which was isolated from the Western Tropical North Pacific Ocean.

Draft Genome Sequence of  Strain XMU1401, Isolated from the Western Tropical North Pacific Ocean.

 is the most abundant and smallest known free-living photosynthetic microorganism and is a key player in marine ecosystems and biogeochemical cycles.  can be broadly divided into high-light-adapted (HL) and low-light-adapted (LL) clades. In this study, we isolated two low-light-adapted I (LLI) strains from the western Pacific Ocean and obtained their genomic data. We reconstructed  evolution based on genome rearrangement. Our results showed that genome rearrangement might have played an important role in  evolution. We also found that the  clades with streamlined genomes maintained relatively high synteny throughout most of their genomes, and several regions served as rearrangement hotspots. Backbone analysis showed that different clades shared a conserved backbone but also had clade-specific regions, and the genes in these regions were associated with ecological adaptations., the most abundant and smallest known free-living photosynthetic microorganism, play a key role in marine ecosystems and biogeochemical cycles. The  genome evolution is a fundamental question related to how  clades adapted to different ecological niches. Recent studies revealed that the gene gain and loss is crucial to the clade differentiation. The significance of our research is that we interpreted the  genome evolution from the perspective of genome structure, and associated the genome rearrangement with the  clade differentiation and subsequent ecological adaptation.

Genome rearrangement shapes  ecological adaptation.

In this study, the transcriptional response of grouper to Singapore grouper iridovirus (SGIV) stimulation was characterized using RNA sequencing. Transcriptome sequencing of three test groups in the grouper was performed using the Illumina MiSeq platform. The three test groups were a control group, which was injected with PBS buffer; a high-susceptible (HS) group, which died shortly after the SGIV injection; and a high-resistance (HR) group, which survived the SGIV injection. In total, 38,253 unigenes were generated. When the HS group was compared with the control group, 885 unigenes were upregulated and 487 unigenes were downregulated. When the HR and control groups were compared, 1114 unigenes were upregulated and 420 were downregulated, and when the HR and HS groups were compared, 1010 unigenes were upregulated and 375 were downregulated. In the KEGG analysis, two immune-related pathways, the p53 and peroxisome proliferator-activated receptor pathways, were detected with highly significant enrichment. In addition, 7465 microsatellites and 22,1569 candidate single nucleotide polymorphisms were identified from our transcriptome data. The results suggested several pathways that are associated with traits of disease susceptibility or disease resistance, and provided extensive information about novel gene sequences, gene expression profiles, and genetic markers. This may contribute to vaccine research and a breeding program against SGIV infection in grouper.

Transcriptomics analysis reveals candidate genes and pathways for susceptibility or resistance to Singapore grouper iridovirus in orange-spotted grouper (Epinephelus coioides).

Article	Year
F-CphI represents a new homing endonuclease family using the Endo VII catalytic motif. Mobile DNA\| PubMed ID: 30100927	2018
Transcriptomic response during phage infection of a marine cyanobacterium under phosphorus-limited conditions. Environmental microbiology\| PubMed ID: 26522011	2016
Picocyanobacteria and deep-ocean fluorescent dissolved organic matter share similar optical properties. Nature communications\| PubMed ID: 28513605	2017
Three-Dimensional Superresolution Imaging of the FtsZ Ring during Cell Division of the Cyanobacterium . mBio\| PubMed ID: 29162705	2017
Draft Genome Sequence of Strain XMU1401, Isolated from the Western Tropical North Pacific Ocean. Genome announcements\| PubMed ID: 29326216	2018
Genome rearrangement shapes ecological adaptation. Applied and environmental microbiology\| PubMed ID: 29915114	2018
Transcriptomics analysis reveals candidate genes and pathways for susceptibility or resistance to Singapore grouper iridovirus in orange-spotted grouper (Epinephelus coioides). Developmental and comparative immunology\| PubMed ID: 30195709	2019

Qinglu Zeng

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