OPEN Origin of marine planktonic cyanobacteria

OPEN Origin of marine planktonic cyanobacteria

Marine planktonic cyanobacteria contributed to the widespread oxygenation of the oceans towards the end of the Pre-Cambrian and their evolutionary origin represents a key transition in the geochemical evolution of the Earth surface. Little is known, however, about the evolutionary events that led to the appearance of marine planktonic cyanobacteria. I present here phylogenomic data from one hundred thirty-five proteins and two ribosomal RNAs, Bayesian relaxed molecular clock data from eighteen proteins, SSU and LSU, and Bayesian stochastic character mapping analyses from one hundred thirty-one cyanobacteria genomes with the aim to unravel key evolutionary steps involved in the origin of marine planktonic cyanobacteria. While filamentous cell types evolved early on at around two thousand six hundred to two thousand three hundred million years ago and likely dominated microbial mats in benthic environments for most of the Proterozoic, two thousand five hundred to five hundred forty-two million years ago, marine planktonic cyanobacteria evolved towards the end of the Proterozoic and early Phanerozoic. Crown groups of modern terrestrial and/or benthic coastal cyanobacteria appeared during the late Paleoproterozoic to early Mesoproterozoic. Decrease in cell diameter and loss of filamentous forms contributed to the evolution of unicellular planktonic lineages during the middle of the Mesoproterozoic, one thousand six hundred to one thousand million years ago, in freshwater environments. This study shows that marine planktonic cyanobacteria evolved from benthic marine and some diverged from freshwater ancestors during the Neoproterozoic, one thousand to five hundred forty-two million years ago.

Cyanobacteria have fundamentally transformed the geochemistry of our planet. Multiple lines of geochemical evidence support the occurrence of intervals of profound global environmental change at the beginning and end of the Proterozoic, two thousand five hundred to five hundred forty-two million years ago. While it is widely accepted that the presence of molecular oxygen in the early fossil record was the result of cyanobacteria activity, little is known about how cyanobacteria evolution, for example, habitat preference, may have contributed to changes in biogeochemical cycles through Earth history. Geochemical evidence has indicated that there was a first step-increase in the oxygenation of the Earth's surface, which is known as the Great Oxidation Event, in the early Paleoproterozoic, two thousand five hundred to one thousand six hundred million years ago. A second but much steeper increase in oxygen levels, known as the Neoproterozoic Oxygenation Event, occurred at around eight hundred to five hundred million years ago. Recent chromium isotope data point to low levels of atmospheric oxygen in the Earth's surface during the mid-Proterozoic, which is consistent with the late evolution of marine planktonic cyanobacteria during the Cryogenian; both types of evidence help explain the late emergence and diversification of metazoans.

Understanding the evolution of planktonic cyanobacteria is an essential question because their origin fundamentally transformed the nitrogen and carbon cycles towards the end of the Pre-Cambrian. It remains unclear, however, what evolutionary events led to the emergence of open-ocean planktonic forms within cyanobacteria, and how these events relate to geochemical evidence during the Pre-Cambrian. So far, it seems that ocean geochemistry, for example, euxinic conditions during the early- to mid-Proterozoic, and nutrient availability likely contributed to the apparent delay in diversification and widespread colonization of open ocean environments by planktonic cyanobacteria during the Neoproterozoic.

Marine phytoplankton today contribute to almost half of the Earth's total primary production. Within the cyanobacteria, only a few lineages colonized the open-ocean, i.e., Crocosphaera and relatives, cyanobacterium UCYN-A, Trichodesmium, as well as Prochlorococcus and Synechococcus.

From these lineages, nitrogen-fixing cyanobacteria are particularly important because they exert a control on primary productivity and the export of organic carbon to the deep ocean, by converting nitrogen gas into ammonium, which is later used to make amino acids and proteins. Marine picocyanobacteria, i.e., Prochlorococcus and Synechococcus, numerically dominate most phytoplankton assemblages in modern oceans contributing importantly to primary productivity. While some planktonic cyanobacteria are unicellular and free living cells, for example, Crocosphaera, Prochlorococcus, Synechococcus, others have established symbiotic relationships with prymnesiophyte algae. Amongst the filamentous forms, Trichodesmium are free-living and form aggregates. However, filamentous heterocyst-forming cyanobacteria, for example, Richelia, Calothrix, are found in association with diatoms such as Hemiaulus, Rhizosolenia, and Chaetoceros.

While environmental conditions might have prevented the widespread diversification of planktonic forms during most of the Pre-Cambrian, the evolutionary history of marine planktonic cyanobacteria, for example, habitat preferences, morphology, likely played an important role in the events surrounding the emergence of complex life in the oceans. Data from one hundred thirty-one cyanobacterial genomes was used to carry out large-scale multi-gene analyses of cyanobacteria; these analyses provide robust evidence for the early evolution of filamentous forms and mat-forming/benthic cyanobacteria and a delay in the emergence of marine planktonic cyanobacteria towards the end of the Pre-Cambrian. Two separate data sets, protein and nucleotide sequence data, and five different types of substitution models, including the CAT-GTR model, were used to explore the timing of key evolutionary events that led to the late emergence of planktonic cyanobacteria. Bayesian stochastic character mapping analyses were performed to study the evolutionary traits involved in the emergence of marine planktonic cyanobacteria such as loss of filamentous forms, and presumably intracellular communication, decrease in cell diameter, and shifts in habitat preference within cyanobacteria. This study also shows that marine planktonic cyanobacteria evolved from benthic marine and freshwater ancestors.

Results

Phylogenetic relationships. An increase in genome sequencing and taxon-sampling have allowed for broad coverage of a range of morphologies, lifestyles, and metabolisms within cyanobacteria. The analyses performed here included a large phylogenetic data set consisting of one hundred thirty-one genome taxa with a total of fifty-six thousand two hundred fifty-one amino acids and four thousand five hundred fifty-five base pairs. Whilst analyses have recovered well-supported monophyletic groups previously reported, new genomic data have revealed novel deep-branching relationships of major cyanobacteria lineages. In this study Pseudanabaena appears as an early divergent lineage within cyanobacteria occurring in eighty-eight percent of the Maximum Likelihood trees generated for each gene alignment, one hundred thirty-seven genes, generated in SATé two point two. A basal position for Pseudanabaena is consistent with recent large-scale multi gene studies. Previous studies suggesting that Pseudanabaena is a derived lineage were based on SSU rRNA datasets.

Genomic data have also clarified problematic phylogenetic relationships such as the positioning of the filamentous LPP group. New data strongly support sister relationships between the LPP clade and Prochlorothrix, Synechoccocous elongatus and the SynPro clade. While the inclusion of recently sequenced genomes suggest a new placement for Trichodesmium, more Oscillatoria-like genomes are needed to fully understand the placement of this important lineage. Modern marine planktonic cyanobacteria evolved within two major groups of cyanobacteria, here referred to as the Microcyanobacteria and the Macrocyanobacteria since they are well-supported monophyletic clades. Whilst the Microcyanobacteria contain lineages with smaller cell diameters, the Macrocyanobacteria contain lineages with larger cell diameters. The Macrocyanobacteria are the most taxonomically and ecologically diverse clade including lineages such as Synechocystis, Pleurocapsa, Microcystis, Trichodesmium and the Nostocales, amongst others.

Relaxed molecular clock analyses. Age divergences were estimated using two independent data sets, RNA and proteins, and applying a Bayesian approach. Four calibration points were implemented, three of which have been previously used. Relaxed molecular clock analyses were performed under the independent-rates model, which has been shown to be the best fitting molecular clock model for cyanobacteria based on Bayes Factors. Four different models of molecular evolution were implemented for proteins and RNA in MCMCtree and the CAT-GTR model for proteins and RNA in Phylobayes. The implementation of two different maximum ages for the origin of oxygenic photosynthesis resulted in different age estimates for the origin of filamentous forms. While an older maximum age predicts the origin of filamentous forms before the GOE with estimates ranging between two thousand six hundred sixty-five and two thousand five hundred fifty-nine million years ago, a younger maximum age predicts filamentous forms appearing around the time of the GOE between two thousand four hundred sixty and two thousand three hundred fifty-one million years ago. Overall an older maximum age tends to make ages older across all analyses.

Results were consistent across models of molecular evolution within each data set. There is strong evidence for a Neoproterozoic or early Cambrian origin for marine unicellular N-fixers.

Crocosphaera clade and the filamentous Nodularia spumigena CCY9414. Age estimates appear to be younger for Prochlorococcus and Synechococcus based on the nucleotide data set, in contrast to the protein data set. All analyses however provide robust evidence for the relatively late evolution of marine planktonic cyanobacteria. Other marine N-fixers evolved during the Phanerozoic such as Richelia and the cyanobacterium UCYN-A clade. Age estimates across all analyses are summarized and are mostly in broad agreement.

Bayesian trait evolution analyses. The earliest cyanobacteria were likely unicellular and inhabited low salinity environments. Living relatives of these early divergent lineages have been isolated from terrestrial, freshwater environments, hot-springs, and coastal marine habitats. Bayesian stochastic character mapping analyses revealed that filamentous cyanobacteria evolved early on and different molecular clock analyses indicate filamentous forms evolved around two thousand six hundred sixty-five to two thousand three hundred fifty-one million years ago and the GOE. Ancestors of early filamentous forms likely resembled modern relatives of Pseudanabaena and the LPP clade. All Basal Lineages and the Microcyanobacteria have retained small cell diameters exhibiting cells that are less than three micrometers, with most lineages exhibiting diameters that are less than two micrometers. Interestingly, further decrease in cell diameter characterizes the evolution of the marine Prochlorococcus within the SynPro clade. Also a switch from filamentous to unicellular cell types occurred around one thousand nine hundred ninety-four to one thousand four hundred twenty-one million years ago.

All analyses suggest that the Macrocyanobacteria clade, exhibiting larger cell diameters, may have evolved just after the GOE with age estimates ranging between two thousand three hundred eighty-six and one thousand eight hundred ninety-four million years ago. Within this clade two opposite evolutionary trends were found: one, an increase in cell diameter within the Nostocales, and two, a decrease in cell diameter within the clade containing Microcystis and Crocosphaera relatives. A switch from filamentous to unicellular forms also occurred around one thousand four hundred thirty-seven to one thousand forty-seven million years ago in freshwater habitats. Whilst unicellular marine N-fixing cyanobacteria and Nodularia spumigena CCY9414 diverged from freshwater ancestors, Trichodesmium evolved from filamentous coastal marine lineages.