Prochlorococcus, the most abundant photosynthetic organism on Earth, diversified into high-light and low-light ecotype clades while its genome was drastically reduced and restructured. Yan et al. (2018) showed from five cultivated clades that major genome rearrangement coincided with genome reduction before the streamlined clades diverged, but left open the full picture: the structural transition from the non-streamlined LLIV clade to the ancestor of streamlined clades, the genomes of uncultivated partially streamlined clades, the cause of the discordance between the rearrangement-based and core-gene phylogenies, and whether loss of DNA repair genes causally accelerates rearrangement. These components remain unresolved, now within a sharpened drift-versus-selection-versus-mutation-rate causal debate.

Background

Prochlorococcus is a marine picocyanobacterium (a photosynthetic bacterium with cells roughly 0.6 μm across and compact genomes of about 1.6-1.8 million base pairs) and the most abundant free-living photosynthetic organism known; it is divided into high-light-adapted (HL, comprising HLI and HLII) and low-light-adapted (LL, comprising LLI, LLII/III, LLIV and several uncultured lineages) clades that occupy depth-stratified ecological niches. Its genomes are 'streamlined': unusually small, poor in G+C base composition (GC-poor), and depleted of redundant genes, a state widely attributed to selection for metabolic efficiency in nutrient-poor open-ocean waters, although an alternative drift-based account has recently gained experimental support (Chen et al. 2022, Nat Ecol Evol 6:183-194; Zhang et al. 2024, ISME J 18:wrad035). Early comparative work established that gene gain and loss, concentrated in hypervariable genomic islands (locally variable gene clusters prone to horizontal acquisition and deletion), drove much of the ecotype differentiation (Kettler et al. 2007, PLoS Genet 3:e231). Beyond local sequence change, genomes also evolve by large-scale rearrangement (inversions, translocations, duplications, deletions), which changes gene order; 'synteny' refers to conservation of gene order across genomes, and tools such as Mauve align whole bacterial genomes into 'locally colinear blocks' (LCBs), conserved stretches of collinear homology whose permutations summarize structural change. Yan et al. (2018, Appl Environ Microbiol 84:e01178-18) isolated two new LLI strains and used LCB permutations across 14 complete genomes to reconstruct a rearrangement-based phylogeny. They found that the streamlined HL, LLI, and LLII/III clades retain high mutual synteny and differ structurally from the non-streamlined LLIV clade, implying that the major rearrangement events coincided with genome reduction and predated divergence of the streamlined clades; that clade-specific backbone regions are enriched in genes for light-shock protection, DNA repair, and nutrient transport; and that the rearrangement-based tree conflicts with the 31-core-gene tree by grouping HL with LLII/III, which they attributed to a higher rearrangement rate in the more repair-gene-depleted HL and LLI lineages. Their analysis was, however, limited to the five clades with cultured whole-genome representatives: uncultured LL clades, known from single-cell and metagenomic surveys to exist and to occupy deep-euphotic and oxygen-minimum niches (Malmstrom et al. 2013, ISME J 7:184-198; Fuchsman et al. 2023, PeerJ 11:e14924), had no genome-structure data, so the structural relationship between LLIV-like ancestors and the streamlined clades, and the order and causes of reduction versus rearrangement versus diversification, could not be resolved. Genome sampling of these lineages has since widened — single-amplified genomes of uncultivated lineages (Berube et al. 2018, Sci Data 5:180154), metagenome-assembled genomes (Ustick et al. 2023, ISME J 17:1671-1679), and the first cultivated representatives of the deeply branching LLVII and LLVIII grades (Becker et al. 2024, mBio 15:e03497-23) are now available — but single-cell assemblies are too fragmented for whole-genome rearrangement analysis, and no post-2018 study has redone the rearrangement/synteny analysis across the deep-LL diversity.

Problem Statement

Determine the order and the causal drivers of genome reduction, genome rearrangement, and clade diversification in Prochlorococcus evolution. Concretely, the unresolved question posed by Yan et al. (2018) comprises linked components: (i) the genome-structural transition from the non-streamlined LLIV lineage to the most recent common ancestor of the highly streamlined clades, including the rearrangement architectures of the uncultivated, presumably partially streamlined LL lineages (e.g., the LLV and related deep-branching clades); (ii) the sequence of events, i.e., whether genome reduction preceded, accompanied, or followed the major rearrangement events and the ecological niche shifts marking clade differentiation; and (iii) which causal forces produced the observed structural evolution, including resolution of why the rearrangement-based phylogeny groups HL with LLII/III while the core-gene phylogeny does not. For (iii), Yan et al. (2018) attributed the discordance to elevated rearrangement or mutation rates in the DNA-repair-gene-depleted HL and LLI lineages but did not causally test that explanation; subsequent literature has added further candidate forces beyond those discussed by the source — notably drift acting through small effective population sizes (Chen et al. 2022; Zhang et al. 2024) and mobile-element activity (e.g., tycheposons, integrative chromosomal elements carrying tyrosine-recombinase genes that excise and reinsert at tRNA sites; Hackl et al. 2023) — which, together with selection for streamlining and elevated rearrangement or mutation rates from loss of DNA repair genes, define the candidate causal scenarios the causal-drivers question must now discriminate among. Resolving the question therefore requires fixing the relative timing of genome reduction, rearrangement, and the ecological niche shifts marking clade differentiation, and establishing, for each structural or causal claim, which candidate scenario the available data support and which they exclude, or, where several act jointly, their relative contribution and the evidence distinguishing them. A further question raised by the source's background — whether the clade-distinguishing rearrangements have functional consequences, such as effects on gene expression — is adjacent context and does not form part of the unresolved question posed by Yan et al. (2018).

References

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