Transcriptomic and photosynthetic analyses of Synechocystis sp. PCC6803 and Chlorogloeopsis fritschii sp. PCC6912 exposed to an M-dwarf spectrum under an anoxic atmosphere

Abstract

Introduction: Cyanobacteria appeared in the anoxic Archean Earth, evolving for the first time oxygenic photosynthesis and deeply changing the atmosphere by introducing oxygen. Starting possibly from UV-protected environments, characterized by low visible and far-red enriched light spectra, cyanobacteria spread everywhere on Earth thanks to their adaptation capabilities in light harvesting. In the last decade, few cyanobacteria species which can acclimate to far-red light through Far-Red Light Photoacclimation (FaRLiP) have been isolated. FaRLiP cyanobacteria were thus proposed as model organisms to study the origin of oxygenic photosynthesis as well as its possible functionality around stars with high far-red emission, the M-dwarfs. These stars are astrobiological targets, as their longevity could sustain life evolution and they demonstrated to host rocky terrestrial-like exoplanets within their Habitable Zone.

Methods: We studied the acclimation responses of the FaRLiP strain Chlorogloeopsis fritschii sp. PCC6912 and the non-FaRLiP strain Synechocystis sp. PCC6803 to the combination of three simulated light spectra (M-dwarf, solar and far-red) and two atmospheric compositions (oxic, anoxic). We first checked their growth, O₂ production and pigment composition, then we studied their transcriptional responses by RNA sequencing under each combination of light spectrum and atmosphere conditions.

Results and discussion: PCC6803 did not show relevant differences in gene expression when comparing the responses to M-dwarf and solar-simulated lights, while far-red caused a variation in the transcriptional level of many genes. PCC6912 showed, on the contrary, different transcriptional responses to each light condition and activated the FaRLiP response under the M-dwarf simulated light. Surprisingly, the anoxic atmosphere did not impact the transcriptional profile of the 2 strains significantly. Results show that both cyanobacteria seem inherently prepared for anoxia and to harvest the photons emitted by a simulated M-dwarf star, whether they are only visible (PCC6803) or also far-red photons (PCC6912). They also show that visible photons in the simulated M-dwarf are sufficient to keep a similar metabolism with respect to solar-simulated light.

Conclusion: Results prove the adaptability of the cyanobacterial metabolism and enhance the plausibility of finding oxygenic biospheres on exoplanets orbiting M-dwarf stars.

Keywords: FaRLiP; M-dwarf spectrum; RNA-Seq; anoxia; cyanobacteria; laboratory simulation experiments; transcriptomic analysis.

Figure 1

O₂ evolution of PCC6912 (A) and PCC6803 (B) in ATM TER and ATM MOD over 48 h of experiment. T₀ was set at 3.600 s to exclude the initial gas equilibration period inside the chamber. Bold lines represent the average of different biological replicates, with standard deviations reported as transparent areas. SOL, Solar light; M7, M-dwarf light; FR, Far-red light; ATM TER, oxic atmosphere; ATM MOD, anoxic atmosphere.

Figure 2

(A) on the right y-axis are plotted the normalized emission spectra of the light simulators utilized in the study. A dashed line with yellow fill pattern, a dotted line with orange fill pattern, and a dash-dot-dot line with dark red fill pattern are used respectively for the solar, M-dwarf, and far-red light spectra; on the left y-axis is plotted an example of in vivo absorption spectra of PCC6912 grown in the three light conditions under ATM TER; in vivo absorption spectra are normalized at 680 nm, while emission spectra of the light simulators are normalized to their respective peak emission in the range 380-780 nm; (B) example HPLC chromatograms at 705 nm of PCC6912 after 48 h of exposure to the different light conditions under ATM TER. Retention times for chlorophylls a, d, and f are highlighted with a red band. In gray is reported a control sample acclimated to FR light for 4 days at the same light intensity tested in these experiments. SOL, Solar light; M7, M-dwarf light; FR, Far-red light; ATM TER, oxic atmosphere.

Figure 3

PCA plot generated from the expression profiles of PCC6912 (A) and PCC6803 (B). Orange, red and yellow dots represent respectively M7, FR, SOL samples. Full and empty dots distinguish respectively ATM TER and ATM MOD samples. SOL, Solar light; M7, M-dwarf light; FR, Far-red light; ATM TER, oxic atmosphere; ATM MOD, anoxic atmosphere.

Figure 4

VST-normalized expression level of most variable genes in PCC6912 (A) and PCC6803 (B). Genes are clustered by Pearson correlation. For each strain, the top panels show the 50 genes which showed the most variation when comparing light conditions, while the bottom panels show the 6 genes with the most variation across atmosphere conditions. Functional categories according to the KEGG database are reported for each gene. SOL, Solar light; M7, M-dwarf light; FR, Far-red light; ATM TER, oxic atmosphere; ATM MOD, anoxic atmosphere.

Figure 5

Changes in expression of genes related to photosynthesis. The LFC values of photosystem and phycobilisome genes in M7 and FR compared to SOL are shown. Genes belonging to the FaRLiP group are highlighted. The statistical significance of individual LFCs is highlighted as follows: *, q<0.05; **, q<10^-3; ***, q<10^-5. PSII, photosystem II; PSI, photosystem I; PBS, phycobilisome; PQ, plastoquinone; PLC, plastocyanin; Fd, ferredoxin. SOL, Solar light; M7, M-dwarf light; FR, Far-red light. Created with BioRender.com.