ABSTRACT
Three-dimensional (3D) structure information, now available at the proteome scale, may facilitate the detection of remote evolutionary relationships in protein superfamilies. Here, we illustrate this with the identification of a novel family of protein domains related to the ferredoxin-like superfold, by combining (i) transitive sequence similarity searches, (ii) clustering approaches, and (iii) the use of AlphaFold2 3D structure models. Domains of this family were initially identified in relation with the intracellular biomineralization of calcium carbonates by Cyanobacteria. They are part of the large heavy-metal-associated (HMA) superfamily, departing from the latter by specific sequence and structural features. In particular, most of them share conserved basic amino acids (hence their name CoBaHMA for Conserved Basic residues HMA), forming a positively charged surface, which is likely to interact with anionic partners. CoBaHMA domains are found in diverse modular organizations in bacteria, existing in the form of monodomain proteins or as part of larger proteins, some of which are membrane proteins involved in transport or lipid metabolism. This suggests that the CoBaHMA domains may exert a regulatory function, involving interactions with anionic lipids. This hypothesis might have a particular resonance in the context of the compartmentalization observed for cyanobacterial intracellular calcium carbonates.
Subject(s)
Amino Acid Sequence , Bacterial Proteins , Metals, Heavy , Models, Molecular , Bacterial Proteins/chemistry , Bacterial Proteins/metabolism , Bacterial Proteins/genetics , Metals, Heavy/chemistry , Metals, Heavy/metabolism , Protein Domains , Cyanobacteria/metabolism , Cyanobacteria/chemistry , Cyanobacteria/genetics , Ferredoxins/chemistry , Ferredoxins/metabolism , Protein FoldingABSTRACT
The formation of intracellular amorphous calcium carbonates (iACC) has been recently observed in a few cultured strains of Microcystis, a potentially toxic bloom-forming cyanobacterium found worldwide in freshwater ecosystems. If iACC-forming Microcystis are abundant within blooms, they may represent a significant amount of particulate Ca. Here, we investigate the significance of iACC biomineralization by Microcystis. First, the presence of iACC-forming Microcystis cells has been detected in several eutrophic lakes, indicating that this phenomenon occurs under environmental conditions. Second, some genotypic (presence/absence of ccyA, a marker gene of iACC biomineralization) and phenotypic (presence/absence of iACC) diversity have been detected within a collection of strains isolated from one single lake. This illustrates that this trait is frequent but also variable within Microcystis even at a single locality. Finally, one-third of publicly available genomes of Microcystis were shown to contain the ccyA gene, revealing a wide geographic and phylogenetic distribution within the genus. Overall, the present work shows that the formation of iACC by Microcystis is common under environmental conditions. While its biological function remains undetermined, this process should be further considered regarding the biology of Microcystis and implications on the Ca geochemical cycle in freshwater environments.
Subject(s)
Cyanobacteria , Microcystis , Microcystis/genetics , Phylogeny , Ecosystem , Lakes/microbiology , Calcium CarbonateABSTRACT
Metagenome-assembled genomes (MAGs) represent individual genomes recovered from metagenomic data. MAGs are extremely useful to analyze uncultured microbial genomic diversity, as well as to characterize associated functional and metabolic potential in natural environments. Recent computational developments have considerably improved MAG reconstruction but also emphasized several limitations, such as the nonbinning of sequence regions with repetitions or distinct nucleotidic composition. Different assembly and binning strategies are often used; however, it still remains unclear which assembly strategy, in combination with which binning approach, offers the best performance for MAG recovery. Several workflows have been proposed in order to reconstruct MAGs, but users are usually limited to single-metagenome assembly or need to manually define sets of metagenomes to coassemble prior to genome binning. Here, we present MAGNETO, an automated workflow dedicated to MAG reconstruction, which includes a fully-automated coassembly step informed by optimal clustering of metagenomic distances, and implements complementary genome binning strategies, for improving MAG recovery. MAGNETO is implemented as a Snakemake workflow and is available at: https://gitlab.univ-nantes.fr/bird_pipeline_registry/magneto. IMPORTANCE Genome-resolved metagenomics has led to the discovery of previously untapped biodiversity within the microbial world. As the development of computational methods for the recovery of genomes from metagenomes continues, existing strategies need to be evaluated and compared to eventually lead to standardized computational workflows. In this study, we compared commonly used assembly and binning strategies and assessed their performance using both simulated and real metagenomic data sets. We propose a novel approach to automate coassembly, avoiding the requirement for a priori knowledge to combine metagenomic information. The comparison against a previous coassembly approach demonstrates a strong impact of this step on genome binning results, but also the benefits of informing coassembly for improving the quality of recovered genomes. MAGNETO integrates complementary assembly-binning strategies to optimize genome reconstruction and provides a complete reads-to-genomes workflow for the growing microbiome research community.
Subject(s)
Metagenomics , Microbiota , Workflow , Metagenomics/methods , Metagenome/genetics , Genome, MicrobialABSTRACT
Cyanobacteria have massively contributed to carbonate deposition over the geological history. They are traditionally thought to biomineralize CaCO3 extracellularly as an indirect byproduct of photosynthesis. However, the recent discovery of freshwater cyanobacteria-forming intracellular amorphous calcium carbonates (iACC) challenges this view. Despite the geochemical interest of such a biomineralization process, its molecular mechanisms and evolutionary history remain elusive. Here, using comparative genomics, we identify a new gene (ccyA) and protein family (calcyanin) possibly associated with cyanobacterial iACC biomineralization. Proteins of the calcyanin family are composed of a conserved C-terminal domain, which likely adopts an original fold, and a variable N-terminal domain whose structure allows differentiating four major types among the 35 known calcyanin homologs. Calcyanin lacks detectable full-length homologs with known function. The overexpression of ccyA in iACC-lacking cyanobacteria resulted in an increased intracellular Ca content. Moreover, ccyA presence was correlated and/or colocalized with genes involved in Ca or HCO3- transport and homeostasis, supporting the hypothesis of a functional role of calcyanin in iACC biomineralization. Whatever its function, ccyA appears as diagnostic of intracellular calcification in cyanobacteria. By searching for ccyA in publicly available genomes, we identified 13 additional cyanobacterial strains forming iACC, as confirmed by microscopy. This extends our knowledge about the phylogenetic and environmental distribution of cyanobacterial iACC biomineralization, especially with the detection of multicellular genera as well as a marine species. Moreover, ccyA was probably present in ancient cyanobacteria, with independent losses in various lineages that resulted in a broad but patchy distribution across modern cyanobacteria.