Anaerobic metabolism of Foraminifera thriving below the seafloor - PubMed (original) (raw)
Anaerobic metabolism of Foraminifera thriving below the seafloor
William D Orsi et al. ISME J. 2020 Oct.
Abstract
Foraminifera are single-celled eukaryotes (protists) of large ecological importance, as well as environmental and paleoenvironmental indicators and biostratigraphic tools. In addition, they are capable of surviving in anoxic marine environments where they represent a major component of the benthic community. However, the cellular adaptations of Foraminifera to the anoxic environment remain poorly constrained. We sampled an oxic-anoxic transition zone in marine sediments from the Namibian shelf, where the genera Bolivina and Stainforthia dominated the Foraminifera community, and use metatranscriptomics to characterize Foraminifera metabolism across the different geochemical conditions. Relative Foraminifera gene expression in anoxic sediment increased an order of magnitude, which was confirmed in a 10-day incubation experiment where the development of anoxia coincided with a 20-40-fold increase in the relative abundance of Foraminifera protein encoding transcripts, attributed primarily to those involved in protein synthesis, intracellular protein trafficking, and modification of the cytoskeleton. This indicated that many Foraminifera were not only surviving but thriving, under the anoxic conditions. The anaerobic energy metabolism of these active Foraminifera was characterized by fermentation of sugars and amino acids, fumarate reduction, and potentially dissimilatory nitrate reduction. Moreover, the gene expression data indicate that under anoxia Foraminifera use the phosphogen creatine phosphate as an ATP store, allowing reserves of high-energy phosphate pool to be maintained for sudden demands of increased energy during anaerobic metabolism. This was co-expressed alongside genes involved in phagocytosis and clathrin-mediated endocytosis (CME). Foraminifera may use CME to utilize dissolved organic matter as a carbon and energy source, in addition to ingestion of prey cells via phagocytosis. These anaerobic metabolic mechanisms help to explain the ecological success of Foraminifera documented in the fossil record since the Cambrian period more than 500 million years ago.
Conflict of interest statement
The authors declare that they have no conflict of interest.
Figures
Fig. 1. Census count of foraminifera tests and corresponding geochemical profiles in anoxic Namibian sediment.
a Density of the foraminifera species in the nine intervals processed. Green and brown colors inside the tests indicate the presence of cytoplasm. b The changing redox profile of in sediment pore water, note the accumulation of hydrogen sulfide with depth below 6 cm. All O2 was below detection immediately below the sediment surface. c Representative specimens of the species enumerated; brownish-green color indicates the presence of cytoplasm. Scale bar 100 µm.
Fig. 2. Foraminifera exhibit high levels of gene expression under anoxia.
a The relative abundance of total expressed ORFs per sample that were assigned to prokaryotes (Bacteria and Archaea) and eukaryotes (including Foraminifera). Multiple histograms per depth represent biological replicates. b The total number of ORFs that were assigned to eukaryotes per sample. Multiple histograms per depth represent biological replicates. c The relative abundance of expressed ORFs from different protist groups (from b), note the dominance of Foraminifera gene expression in the deepest, most anoxic sample at 28 cm. Fungal analogs grouping corresponds to the Labyrinthulomycetes. d The relative abundance of functional eukaryotic gene (KOG) families in the three sediment zones that were assigned to expressed Foraminifera ORFs. Pie charts represent average values from the biological replicates shown in a–c. CT core top sample.
Fig. 3. Phylogenetic analysis of Foraminifera affiliated 18S rRNA sequences recovered from the metatranscriptomes.
Two 18S rRNA sequences were detected in the metatranscriptomes that are affiliated with the (a) Stainforthiidae family and (b) Bolivina genus. The sequence affiliated to the Stainforthiidae family clearly cluster with the only two representative genera of the family, Stainforthia and Gallietellia but the position of the metatranscriptomic 18S rDNA sequence is not clearly resolved, but intact test of Stainforthia were observed in the sample (See Fig. 1). The metatranscriptomic 18S rDNA sequence related to Bolivina is nearly identical to reference sequences deposited on NCBI and that were generated from Bolivina specimens collected in Namibia in previous studies. Furthermore, Bolivina specimens dominated the morphological assemblages within the core (Fig. 1). The Bolivina and Stainforthia 18S rDNA contigs were generated by semiautomated greedy extension of 18S rDNA OTUs with trimmed metatranscriptomic paired-end reads (see “Methods”).
Fig. 4. Expression of Foraminifera ORFs involved in key anaerobic physiologies.
a Heatmap displaying the expression levels of Foraminifera ORFs involved in anaerobic energy production and physiology. Dendrogram shows hierarchical clustering (UPGMA) of the samples based on the RNA-seq data. One metatranscriptome from the core top and one from the 12 cm sample did not have any detectable expression of the ORFs of interest and are thus not shown. b Reconstruction of anaerobic cellular activities in Foraminifera including biomineralization, phagocytosis, CME, and transport of ingested cargo (Banning, Novel strains isolated from a coastal aquifer suggest a predatory role for flavobacteria) based on the gene expression data shown in a. c Reconstruction of potential anaerobic energy production pathways in Foraminifera based on the gene expression data shown in a. Red colors show genes that were expressed, red arrows show reactions that are predicted to occur based on the expression of the corresponding gene. Where expressed, gene abbreviations (e.g., Nrt) are shown in red boxes, that correspond to the same labels in a. Gene abbreviations displayed with white background are present in the genome of the benthic foraminifera species Globobulimina turgida and G. auriculata [10], but expression was not detected. These include FH fumarase, KGDH alpha-ketoglutarate dehydrogenase, PK pyruvate kinase, and ASCT acetate:succinate CoA-transferase. This updated representation of Foraminifera anaerobic energy production is modified from anaerobic energy metabolism pathways in eukaryotes that were previously reviewed [39, 40].
Fig. 5. Oxygen consumption and Foraminifera gene expression in a 10-day incubation.
a Oxygen consumption at the top (in seawater) and bottom (underneath the sediment) of the incubated sediments, the photo shows the experimental setup and the positioning of the two oxygen sensor spots where measurements were made. After the onset of anoxia after 20 h, the top and bottom of the flask remained anoxic for the duration of the incubation. The flask was incubated in the dark at 10 °C. The replicate measurements at each time point are those made on the four separate flasks incubated for the _t_1, _t_2, _t_3, _t_4 timepoints. b The relative abundance of bacterial and archaeal ORFs compared with total eukaryotic ORFs (top), and the relative abundance of ORFs from eukaryotic groups detected in the metatranscriptomes (bottom). c The number of ORFs assigned to Foraminifera (top) and the relative abundance of KOG categories within those foraminiferal ORFs (bottom).
Fig. 6. Foraminifera gene categories whose relative expression increased in the presence of anoxia.
The five gene categories are shown based on KOG annotations. The category “mitochondrial proteins” are those KOG annotations that have the word “mitochondria”, “mitochondrion”, or “mitochondrial”, in the KOG description. The replicates are shown for each time point are displayed as two individual points, and represent the fractional abundance of all reads mapping to ORFs with a given annotation.
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