Cantaloupe melon genome reveals 3D chromatin features and structural relationship with the ancestral cucurbitaceae karyotype

Abstract

Cucumis melo displays a large diversity of horticultural groups with cantaloupe melon the most cultivated type. Using a combination of single-molecule sequencing, 10X Genomics link-reads, high-density optical and genetic maps, and chromosome conformation capture (Hi-C), we assembled a chromosome scale C. melo var. cantalupensis Charentais mono genome. Integration of RNA-seq, MeDip-seq, ChIP-seq, and Hi-C data revealed a widespread compartmentalization of the melon genome, segregating constitutive heterochromatin and euchromatin. Genome-wide comparative and evolutionary analysis between melon botanical groups identified Charentais mono genome increasingly more divergent from Harukei-3 (reticulatus), Payzawat (inodorus), and HS (ssp. agrestis) genomes. To assess the paleohistory of the Cucurbitaceae, we reconstructed the ancestral Cucurbitaceae karyotype and compared it to sequenced cucurbit genomes. In contrast to other species that experienced massive chromosome shuffling, melon has retained the ancestral genome structure. We provide comprehensive genomic resources and new insights in the diversity of melon horticultural groups and evolution of cucurbits.

Keywords: genomics; omics; plant biology; plant evolution.

Graphical abstract

Figure 1

Charmono chromosome-level-assembly (A–D) Flowers and fruit of the monoecious C.melo var. cantalupensis Charmono cultivar. Female (A) and male flowers (B) and the whole (C) or cut fruit (D). Ov, ovary; sg, stigmate; st, stamen. Scale bars: (A), 5mm; (B), 2mm; (C and D), 5cm. (E) Neighbor-joining phylogenetics trees of 14 different C. melo accessions/varieties with 1,000 bootstrap replicates. The percentage of replicate trees in which the associated taxa clustered together in the bootstrap test are shown next to the branches. C. melo ssp. melo and C. melo ssp. agrestis are highlighted in green and orange, respectively. Cucumber, C. sativus, was used as outgroup. (F) Representations of chromosome connections between the physical positions on the reconstructed chromosome and genetic-map positions in Charmono genome. (G) Hi-C map of the 12 chromosome of Charmono genome at 50 Kb of resolution. Heatmap intensity scale is indicated by the interaction intensity color bar.

Figure 2

Charmono genome annotation and TE distribution (Aand B) Circos plot showing features of C. melo var. cantalupensisCharmono genome in 500 kb bins. Gene (A) and transposable element (B) density heatmaps. (C–F) Distribution of the epigenetic features. (C) H3K9ac, (D) Accessible chromatin, (E) H3K27me3 and (F) Global DNA methylation.(B) BUSCO completeness based on Viridiplantae dataset comparison between available melon annotations. Colors in the bar represent the different classes of the BUSCO assessment results. Red, yellow, purple, and blue indicate Missing, Fragmented, Duplicated, and single-copy category, respectively.(C) TE landscape surrounding Charmono genes. For all genes, 10 kb upstream the transcription start site (TSS) and 10 kb downstream transcription termination site (TTS) were analyzed.

Figure 3

Characterization of the leaf Charmonoepigenome (A) Immunofluorescence detection of H3K27me1 (green) and H3K27me3 (purple) and DAPI (white) in isolated nucleus. Scale bar represents 10 μm. (B) Integration of Hi-C, ChIP-seq, MeDIP-seq and RNA-seq data at the chromosome scale. The heatmap of intra-chromosomal interaction frequency of chromosome 2 is presented. The color bar shows the interaction frequency scale. (C) Integration of Hi-C, ChIP-seq, MeDIP-seq and RNA-seq data at the megabase scale. A 2D heatmap with the interaction frequency in region chr2-13000000-17,000,000 is presented. (D) Genomic and chromatin features of the melon A/B compartments. Boxplots showing comparisons of H3K9ac level, H3K27me3 level, gene density and TE density between A and B compartment. (E) Triangle heatmap of Hi-C interaction frequency. The H3K9ac, H3K27me3 and DNA methylation signals are presented in green, red and blue, respectively. The RNA-seq signal is represented in black. (F) Interacting domains are associated to specific chromatin marks. Unsupervised clustering of chromatin domains associated to DNA methylation, H3K27me3 and H3K9ac marks. The median read count plot of each classified domain related to the histone marks H3K9ac (green), H3K27me3 (orange), global DNA methylation (pink) and accessible chromatin regions signal (purple) is presented. (G) Repressive domains are larger than transcriptionally active domains. Boxplots representing the median size of the transcriptionally active domains (green), polycomb domains (purple) and repressive domains (pink). ∗∗∗ indicates pvalue =6.04 × 10⁻¹⁶, ∗∗ pvalue = 0.00164, ∗pvalue = 9.62 × 10⁻⁸ with Wilcoxon rank-sum test.

Figure 4

Genome-wide comparison of melon genomes (A) Dot plot alignments comparing the chromosome sequence identity between Charmono and melon sequenced genomes (DHL92, Payzawat, Harukei and HS). Percentage of identity filter >75%. (B and C) Number of substitutions (B) and InDels (C) per chromosome. Orange, green, purple and cyan colors indicate Charmonovs HS, CharmonovsPayzawat, Charmonovs DHL92 and CharmonovsHarukei-3 comparisons, respectively. (D) Percentage of substitutions affecting a coding sequence region (CDS) or a non-coding region for each genome comparison. (E) Percentage of non-triple or triple InDels for each genome comparison. (F) Examples of CharmonovsHarukei-3 InDels affecting genes. Red and blue triangles indicate the position of the insertion or deletion, respectively.

Figure 5

Cucurbitaceae paleoevolutionary history (A) TOP-Evolutionary scenario of the modern Cucurbitaceae (squash, watermelon, gourd, melon, pumpkin and cucumber) genomes from the ancestral Cucurbitaceae karyotype (ACK) and the ancestral eudicot karyotype (AEK). The modern genomes are illustrated at the bottom with different colors reflecting the origin from the 22 ancestral chromosomes from ACK. Duplication (WGD) and triplication (WGT) events are shown with red dots on the tree branches, along with the shuffling events (fusions and fissions). BOTTOM- Complete dot-plot based deconvolution of the observed synteny and paralogy (dot-plot diagonals) between ACK (yaxis) and the investigated species (dot-plot xaxis). The synteny (paralogous and orthologous genes) relationships delivered between the modern Cucurbitaceae genomes and ACK are illustrated in green circles, as case example for ACK protochromosome 22. (B) Gene conservation between Charmono(cantalupensis),Harukei-3 (reticulatus), Payzawat (inodorus) and HS (ssp. agrestis). (C) Dotplot-based deconvolution of the synteny between Charmono(cantalupensis), vsHarukei-3 (reticulatus), vs HS (ssp agrestis) and vsPayzawat (inodorus), illustrating shared inversions between CharmonovsPayzawat and vs HS (illuminated in brown) on chromosome 11 and shared translocation on chromosome 4 between Charmono versus Harukei-3 and versus HS (illuminated in green).