Evolutionary mechanisms of long-term genome diversification associated with niche partitioning in marine picocyanobacteria

Marine picocyanobacteria of the genera Prochlorococcus and Synechococcus are the most abundant photosynthetic organisms on Earth, an ecological success thought to be linked to the differential partitioning of distinct ecotypes into specific ecological niches. However, the underlying processes that governed the diversification of these microorganisms and the appearance of niche-related phenotypic traits are just starting to be elucidated. Here, by comparing 81 genomes, including 34 new Synechococcus, we explored the evolutionary processes that shaped the genomic diversity of picocyanobacteria. Time-calibration of a core-protein tree showed that gene gain/loss occurred at an unexpectedly low rate between the different lineages, with for instance 5.6 genes gained per million years (My) for the major Synechococcus lineage (sub-cluster 5.1), among which only 0.71/My have been fixed in the long term. Gene content comparisons revealed a number of candidates involved in nutrient adaptation, a large proportion of which are located in genomic islands shared between either closely or more distantly related strains, as identified using an original network construction approach. Interestingly, strains representative of the different ecotypes co-occurring in phosphorus-depleted waters (Synechococcus clades III, WPC1, and sub-cluster 5.3) were shown to display different adaptation strategies to this limitation. In contrast, we found few genes potentially involved in adaptation to temperature when comparing cold and warm thermotypes. Indeed, comparison of core protein sequences highlighted variants specific to cold thermotypes, notably involved in carotenoid biosynthesis and the oxidative stress response, revealing that long-term adaptation to thermal niches relies on amino acid substitutions rather than on gene content variation. Altogether, this study not only deciphers the respective roles of gene gains/losses and sequence variation but also uncovers numerous gene candidates likely involved in niche partitioning of two key members of the marine phytoplankton

ÉTUDE GÉNOMIQUE, MÉTAGÉNOMIQUE ET PHYSIOLOGIQUE DE LA DIVERSITÉ PIGMENTAIRE CHEZ LES CYANOBACTÉRIES DU GENRE SYNECHOCOCCUS

Picocyanobacteria of the Synechococcus genus are present in all marine environments. This ubiquity is partly explained by the large pigment diversity found within this genus, allowing them to efficiently capture photons over a broad spectral range. Most strains have a fixed pigmentation but some of them are able to match their pigmentation with the ambient light quality by a physiological process called type IV chromatic acclimation (CA4). An original targeted metagenomics approach, combining sophisticated techniques, was developed to study the diversity and distribution of pigment types of Synechococcus in situ. The interest of our approach has been demonstrated after specific optimizations. Availability of 25 new genomes of Synechococcus has allowed us to make significant advances in the understanding of molecular mechanism of CA4. A cluster of 4 to 6 genes, encoding a phycobilin lyase and several transcriptional regulators, is consistently present in all strains capable of this phenotypic plasticity. Two distinct configurations of this cluster, named CA4-A and CA4-B, have been discovered and were found in different Synechococcus lineages. These two types of clusters have undergone distinct evolutionary processes. In addition, some phenotypic peculiarities between strains having these two types of genomic clusters have been demonstrated. This thesis raises new hypotheses about the regulation of this phenotypic plasticity as well as the biochemical mechanisms involved. Keywords : Synechococcus, pigment diversity, genetic diversity, targeted metagenomics, chromatic acclimationLes picocyanobactéries du genre Synechococcus sont présentes dans tous les types d'environnements marins. Cette ubiquité s'explique en partie par la grande diversité pigmentaire de ces cellules, leur permettant de capturer efficacement la lumière sur une large gamme spectrale. La plupart des souches ont une composition pigmentaire fixe mais certaines sont capables d'ajuster leur pigmentation en fonction de la lumière incidente par un processus physiologique appelé acclimatation chromatique de type IV (AC4). Une approche de métagénomique ciblée originale, combinant des techniques sophistiquées, a été développée afin d'étudier la diversité et la distribution des différents types pigmentaires de Synechococcus in situ. L'intérêt de cette approche a pu être démontré après des optimisations spécifiques. La disponibilité de 25 nouveaux génomes de Synechococcus a permis de faire d'importantes avancées sur la compréhension du mécanisme d'AC4. Un cluster de 4 à 6 gènes, codant pour une phycobiline-lyase et plusieurs régulateurs transcriptionnels, est systématiquement présent chez toutes les souches capables de cette plasticité phénotypique. Deux configurations bien distinctes de ce cluster, nommées AC4-A et AC4-B, ont été découvertes et se retrouvent dans des lignées différentes de Synechococcus. Ces deux types de clusters auraient été soumis à des processus évolutifs distincts. Par ailleurs, certaines singularités phénotypiques entre les souches possédant ces deux types de clusters génomiques ont pu être démontrées. Ce travail de thèse soulève de nouvelles hypothèses sur la régulation de cette plasticité phénotypique ainsi que sur les mécanismes biochimiques associés

Connecting thermal physiology and latitudinal niche partitioning in marine Synechococcus

International audienceMarine Synechococcus cyanobacteria constitute a monophyletic group that displays a wide latitudinal distribution, ranging from the equator to the polar fronts. Whether these organisms are all physiologically adapted to stand a large temperature gradient or stenotherms with narrow growth temperature ranges has so far remained unexplored. We submitted a panel of six strains, isolated along a gradient of latitude in the North Atlantic Ocean, to long- and short-term variations of temperature. Upon a downward shift of temperature, the strains showed strikingly distinct resistance, seemingly related to their latitude of isolation, with tropical strains collapsing while northern strains were capable of growing. This behaviour was associated to differential photosynthetic performances. In the tropical strains, the rapid photosystem II inactivation and the decrease of the antioxydant β-carotene relative to chl a suggested a strong induction of oxidative stress. These different responses were related to the thermal preferenda of the strains. The northern strains could grow at 10 °C while the other strains preferred higher temperatures. In addition, we pointed out a correspondence between strain isolation temperature and phylogeny. In particular, clades I and IV laboratory strains were all collected in the coldest waters of the distribution area of marine Synechococus. We, however, show that clade I Synechococcus exhibit different levels of adaptation, which apparently reflect their location on the latitudinal temperature gradient. This study reveals the existence of lineages of marine Synechococcus physiologically specialised in different thermal niches, therefore suggesting the existence of temperature ecotypes within the marine Synechococcus radiation

A gene island with two possible configurations is involved in chromatic acclimation in marine Synechococcus.

Synechococcus, the second most abundant oxygenic phototroph in the marine environment, harbors the largest pigment diversity known within a single genus of cyanobacteria, allowing it to exploit a wide range of light niches. Some strains are capable of Type IV chromatic acclimation (CA4), a process by which cells can match the phycobilin content of their phycobilisomes to the ambient light quality. Here, we performed extensive genomic comparisons to explore the diversity of this process within the marine Synechococcus radiation. A specific gene island was identified in all CA4-performing strains, containing two genes (fciA/b) coding for possible transcriptional regulators and one gene coding for a phycobilin lyase. However, two distinct configurations of this cluster were observed, depending on the lineage. CA4-A islands contain the mpeZ gene, encoding a recently characterized phycoerythrobilin lyase-isomerase, and a third, small, possible regulator called fciC. In CA4-B islands, the lyase gene encodes an uncharacterized relative of MpeZ, called MpeW. While mpeZ is expressed more in blue light than green light, this is the reverse for mpeW, although only small phenotypic differences were found among chromatic acclimaters possessing either CA4 island type. This study provides novel insights into understanding both diversity and evolution of the CA4 process

Differential expression of <i>mpeW</i> or <i>mpeZ</i> genes in <i>Synechococcus</i> cultures acclimated to BL <i>vs</i>.

<div><p><b>GL as measured by real time PCR (20 µmol photons m^-2 s^-1 in both conditions)</b>. </p> <p>Mean and standard deviation were calculated from 3 biological replicates. Only differential transcript levels above or below the dotted lines (log₂(FC) > 1 or <-1) were considered as significant.</p></div

Maximum likelihood analysis of AraC-like proteins (289 aa positions) from marine <i>Synechococcus</i>.

<p>For each strain, the phylogenetic affiliation is mentioned between brackets and the pigment type is indicated by colored circles (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0084459#pone-0084459-t001" target="_blank">Table 1</a> for details). Series of four numbers shown at nodes correspond to bootstrap values for ML analyses, Bayesian posterior probabilities (PP, ranging between 0 and 1), and bootstrap values for Neighbor-Joining and Parsimony methods, respectively. Only values higher than 0.60 for PP and 60% for bootstrap values are shown on the phylogenetic tree. The scale bar represents 0.4 substitutions per amino acid.</p

Correlations between growth rate (µ, days) and half maximal acclimation time (T₅₀, days) for twelve <i>Synechococcus</i> strains.

<p>Shifts to another light quality (BL to GL and vice versa) were performed under two light intensities. LL and HL correspond respectively to 20 and 75 µmol photons m^-2 s^-1 of BL or GL. Data correspond to two biological replicates for each strain and each light condition. </p

CyanoLyase: a database of phycobilin lyase sequences, motifs and functions

International audienceCyanoLyase (http://cyanolyase.genouest.org/) is a manually curated sequence and motif database of phycobilin lyases and related proteins. These enzymes catalyze the covalent ligation of chromo-phores (phycobilins) to specific binding sites of phycobiliproteins (PBPs). The latter constitute the building bricks of phycobilisomes, the major light-harvesting systems of cyanobacteria and red algae. Phycobilin lyases sequences are poorly anno-tated in public databases. Sequences included in CyanoLyase were retrieved from all available genomes of these organisms and a few others by similarity searches using biochemically characteri-zed enzyme sequences and then classified into 3 clans and 32 families. Amino acid motifs were computed for each family using Protomata learner. CyanoLyase also includes BLAST and a novel pattern matching tool (Protomatch) that allow users to rapidly retrieve and annotate lyases from any new genome. In addition, it provides phylogen-etic analyses of all phycobilin lyases families, de-scribes their function, their presence/absence in all genomes of the database (phyletic profiles) and predicts the chromophorylation of PBPs in each strain. The site also includes a thorough bibliog-raphy about phycobilin lyases and genomes included in the database. This resource should be useful to scientists and companies interested in natural or artificial PBPs, which have a number of biotechnological applications, notably as fluores-cent markers

Phenotypic variability of chromatic acclimation in marine <i>Synechococcus</i>.

<p>Temporal changes of the Exc_495:545 after shifts from BL to GL (and vice versa) at two irradiances: LL (circles; 20 µmol photons m^-2 s^-1) and HL (triangles; 75 µmol photons m^-2 s^-1). The color (blue or green) of lines and symbols matches the ambient light color under which cultures were shifted at time zero. Error bars indicate standard deviation for two biological replicates. Four distinct CA4 phenotypic groups were observed. (<b>A</b>,<b>B</b>) Group 1 strains: MINOS11, A15-62, PROS-U-1, M11.1 and RS9915; (<b>C</b>,<b>D</b>) group 2: BIOS-U3-1, CC9311, WH8020, CC9902 and RS9916; (<b>E</b>,<b>F</b>) group 3: BL107 and CC9902; (<b>G</b>,<b>H</b>): group 4 : RCC307. Note the different x-axis scale for BL107. </p

Maximum likelihood analysis of four phycobilin lyase sequences of the E/F clan (380 aa positions) from marine <i>Synechococcus</i>.

<p>For each strain, the phylogenetic affiliation is mentioned between brackets and the pigment type is indicated by colored circles (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0084459#pone-0084459-t001" target="_blank">Table 1</a> for details). Series of four numbers shown at nodes correspond to bootstrap values for ML analyses, Bayesian posterior probabilities (PP, ranging between 0 and 1), and bootstrap values for Neighbor-Joining and Parsimony methods, respectively. Only values higher than 0.60 for PP and 60% for bootstrap values are shown on the phylogenetic tree. The scale bar represents 0.6 substitutions per amino acid. The branch bearing the CpeY cluster was shortened for readability (its full length initially was 3.85 substitutions per amino acid).</p