Repeatome evolution across space and time: Unravelling repeats dynamics in the plant genus Erythrostemon Klotzsch (Leguminosae Juss).

Fluctuations in genomic repetitive fractions (repeatome) are known to impact several facets of evolution, such as ecological adaptation and speciation processes. Therefore, investigating the divergence of repetitive elements can provide insights into an important evolutionary force. However, it is not clear how the different repetitive element clades are impacted by the different factors such as ecological changes and/or phylogeny. To discuss this, we used the Neotropical legume genus Erythrostemon (Caesalpinioideae) as a model, given its ancient origin (~33 Mya), lineage-specific niche conservatism, macroecological heterogeneity, and disjunct distribution in Meso- and South American (MA and SA respectively) lineages. We performed a comparative repeatomic analysis of 18 Erythrostemon species to test the impact of environmental variables over repeats diversification. Overall, repeatome composition was diverse, with high abundances of satDNAs and Ty3/gypsy-Tekay transposable elements, predominantly in the MA and SA lineages respectively. However, unexpected repeatome profiles unrelated to the phylogeny/biogeography were found in a few MA (E. coccineus, E. pannosus and E. placidus) and SA (E. calycinus) species, related to reticulate evolution and incongruence between nuclear and plastid topology, suggesting ancient hybridizations. The plesiomorphic Tekay and satDNA pattern was altered in the MA-sensu stricto subclade with a striking genomic differentiation (expansion of satDNA and retraction of Tekay) associated with the colonization of a new environment in Central America around 20 Mya. Our data reveal that the current species-specific Tekay pool was the result of two bursts of amplification probably in the Miocene, with distinct patterns for the MA and SA repeatomes. This suggests a strong role of the Tekay elements as modulators of the genome-environment interaction in Erythrostemon, providing macroevolutionary insights about mechanisms of repeatome differentiation and plant diversification across space and time.

How diverse a monocentric chromosome can be? Repeatome and centromeric organization of Juncus effusus (Juncaceae).

Juncus is the largest genus of Juncaceae and was considered holocentric for a long time. Recent findings, however, indicated that 11 species from different clades of the genus have monocentric chromosomes. Thus, the Juncus centromere organization and evolution need to be reassessed. We aimed to investigate the major repetitive DNA sequences of two accessions of Juncus effusus and its centromeric structure by employing whole-genome analyses, fluorescent in situ hybridization, CENH3 immunodetection, and chromatin immunoprecipitation sequencing. We showed that the repetitive fraction of the small J. effusus genome (~270 Mbp/1C) is mainly composed of Class I and Class II transposable elements (TEs) and satellite DNAs. Three identified satellite DNA families were mainly (peri)centromeric, with two being associated with the centromeric protein CENH3, but not strictly centromeric. Two types of centromere organization were discerned in J. effusus: type 1 was characterized by a single CENH3 domain enriched with JefSAT1-155 or JefSAT2-180, whereas type 2 showed multiple CENH3 domains interrupted by other satellites, TEs or genes. Furthermore, while type 1 centromeres showed a higher degree of satellite identity along the array, type 2 centromeres had less homogenized arrays along the multiple CENH3 domains per chromosome. Although the analyses confirmed the monocentric organization of J. effusus chromosomes, our data indicate a more dynamic arrangement of J. effusus centromeres than observed for other plant species, suggesting it may constitute a transient state between mono- and holocentricity.

Repeat-based phylogenomics shed light on unclear relationships in the monocentric genus Juncus L. (Juncaceae).

The repetitive fraction (repeatome) of eukaryotic genomes is diverse and usually fast evolving, being an important tool for clarify plant systematics. The genus Juncus L. comprises 332 species, karyotypically recognized by having holocentric chromosomes. However, four species were recently described as monocentric, yet our understanding of their genome evolution is largely masked by unclear phylogenetic relationships. Here, we reassess the current Juncus systematics using low-coverage genome skimming data of 33 taxa to construct repeats, nuclear rDNA and plastome-based phylogenetic hypothesis. Furthermore, we characterize the repeatome and chromosomal distribution of Juncus-specific centromeric repeats/CENH3 protein to test the monocentricity reach in the genus. Repeat-base phylogenies revealed topologies congruent with the rDNA tree, but not with the plastome tree. The incongruence between nuclear and plastome chloroplast dataset suggest an ancient hybridization in the divergence of Juncotypus and Tenageia sections 40 Myr ago. The phylogenetic resolution at section level was better fitted with the rDNA/repeat-based approaches, with the recognition of two monophyletic sections (Stygiopsis and Tenageia). We found specific repeatome trends for the main lineages, such as the higher abundances of TEs in the Caespitosi and Iridifolii + Ozophyllum clades. CENH3 immunostaining confirmed the monocentricity of Juncus, which can be a generic synapomorphy for the genus. The heterogeneity of the repeatomes, with high phylogenetic informativeness, identified here may be correlated with their ancient origin (56 Mya) and reveals the potential of comparative genomic analyses for understanding plant systematics and evolution.

How diverse is heterochromatin in the Caesalpinia group? Cytogenomic characterization of Erythrostemon hughesii Gagnon & G.P. Lewis (Leguminosae: Caesalpinioideae).

MAIN CONCLUSION: Cytogenomic characterization of Erythrostemon hughesii reveals a heterogeneity of repeats in its subtelomeric heterochromatin. Comparative analyses with other Caesalpinia group species reveal a significant reduction in the abundance of Ty3-gypsy/Chromovirus Tekay retrotransposons during its evolution. In numerically stable karyotypes, repetitive DNA variability is one of the main causes of genome and chromosome variation and evolution. Species from the Caesalpinia group (Leguminosae) are karyotypically characterized by 2n = 24, with small chromosomes and highly variable CMA+ heterochromatin banding patterns that correlate with environmental variables. Erythrostemon hughesii differs from other species of the group examined to date for having subtelomeric CMA+ bands; this contrasts with most species in the group which have proximal bands. Here we analyse the repeatome of E. hughesii using genome skimming and chromosomal mapping approaches to characterize the identity of the most abundant repetitive elements and their physical location. The repetitive fraction of E. hughesii comprises 28.73% of the genome. The most abundant elements were retrotransposons (RT) with long terminal repeats (LTR-RT; 9.76%) and satellite DNAs (7.83%). Within the LTR-RTs, the most abundant lineages were: Ty1/copia-Ale (1%), Ty3/gypsy CRM (0.88%) and Ty3/gypsy Athila (0.75%). Using fluorescent in situ hybridization four satellite DNAs and several LTR-RT elements were shown to be present in most subtelomeric CMA+ bands. These results highlight how the repeatome in E. hughesii, a species from Oaxaca state in Mexico, is clearly distinct from Northeast Brazilian species of the Caesalpinia group, mainly due to its high diversity of repeats in its subtelomeric heterochromatic bands and low amount of LTR-RT Ty3/gypsy-Tekay elements. Comparative sequence analysis of Tekay elements from different species is congruent with a clade-specific origin of this LTR-RT after the divergence of the Caesalpinia group. We hypothesize that repeat-rich heterochromatin may play a role in leading to faster genomic divergence between individuals, increasing speciation and diversification.

Evolutionary convergence or homology? Comparative cytogenomics of Caesalpinia group species (Leguminosae) reveals diversification in the pericentromeric heterochromatic composition.

MAIN CONCLUSION: We demonstrated by cytogenomic analysis that the proximal heterochromatin of the Northeast Brazilian species of Caesalpinia group is enriched with phylogenetically conserved Ty3/Gypsy-Tekay RT, but diverge in the presence of Ty3/Gypsy-Athila RT and satDNA. The Caesalpinia Group includes 225 species and 27 monophyletic genera of which four occur in Northeastern Brazil: Erythrostemon (1 sp.), Cenostigma (7 spp.), Libidibia (1 sp.), and Paubrasilia (1 sp.). The last three genera are placed in different clades in the Caesalpinia Group phylogeny, and yet they are characterized by having a numerically stable karyotype 2n = 24 (16 M+8A) and GC-rich heterochromatic bands (chromomycin A3 positive/CMA+ bands) in the proximal chromosome regions. To characterize the composition of their heterochromatin and test for the homology of these chromosomal regions, genomic DNA was extracted from Cenostigma microphyllum, Libidibia ferrea, and Paubrasilia echinata, and sequenced at low coverage using the Illumina platform. The genomic repetitive fractions were characterized using a Galaxy/RepeatExplorer-Elixir platform. The most abundant elements of each genome were chromosomally located by fluorescent in situ hybridization (FISH) and compared to the CMA+ heterochromatin distribution. The repetitive fraction of the genomes of C. microphyllum, L. ferrea, and P. echinata were estimated to be 41.70%, 38.44%, and 72.51%, respectively. Ty3/Gypsy retrotransposons (RT), specifically the Tekay lineage, were the most abundant repeats in each of the three genomes. FISH mapping revealed species-specific patterns for the Tekay elements in the proximal regions of the chromosomes, co-localized with CMA+ bands. Other species-specific patterns were observed, e.g., for the Ty3/Gypsy RT Athila elements which were found in all the proximal heterochromatin of L. ferrea or restricted to the acrocentric chromosomes of C. microphyllum. This Athila labeling co-localized with satellite DNAs (satDNAs). Although the Caesalpinia Group diverged around 55 Mya, our results suggest an ancestral colonization of Tekay RT in the proximal heterochromatin. Thus, the present-day composition of the pericentromeric heterochromatin in these Northeast Brazilian species is a combination of the maintenance of an ancestral Tekay distribution with a species-specific accumulation of other repeats.