A chromosome-level genome assembly for Erianthus fulvus provides insights into its biofuel potential and facilitates breeding for improvement of sugarcane

Abstract

Erianthus produces substantial biomass, exhibits a good Brix value, and shows wide environmental adaptability, making it a potential biofuel plant. In contrast to closely related sorghum and sugarcane, Erianthus can grow in degraded soils, thus releasing pressure on agricultural lands used for biofuel production. However, the lack of genomic resources for Erianthus hinders its genetic improvement, thus limiting its potential for biofuel production. In the present study, we generated a chromosome-scale reference genome for Erianthus fulvus Nees. The genome size estimated by flow cytometry was 937 Mb, and the assembled genome size was 902 Mb, covering 96.26% of the estimated genome size. A total of 35 065 protein-coding genes were predicted, and 67.89% of the genome was found to be repetitive. A recent whole-genome duplication occurred approximately 74.10 million years ago in the E. fulvus genome. Phylogenetic analysis showed that E. fulvus is evolutionarily closer to S. spontaneum and diverged after S. bicolor. Three of the 10 chromosomes of E. fulvus formed through rearrangements of ancestral chromosomes. Phylogenetic reconstruction of the Saccharum complex revealed a polyphyletic origin of the complex and a sister relationship of E. fulvus with Saccharum sp., excluding S. arundinaceum. On the basis of the four amino acid residues that provide substrate specificity, the E. fulvus SWEET proteins were classified as mono- and disaccharide sugar transporters. Ortho-QTL genes identified for 10 biofuel-related traits may aid in the rapid screening of E. fulvus populations to enhance breeding programs for improved biofuel production. The results of this study provide valuable insights for breeding programs aimed at improving biofuel production in E. fulvus and enhancing sugarcane introgression programs.

Keywords: Erianthus fulvus; QTLs; SWEET family; biofuel; cold stress; reference genome.

Figure 1

Circos plot showing the genome characteristics of E. fulvus. The nine rings from outside to inside are (A) chromosomes, (B)Gypsy elements, (C)copia elements, (D) FPKM values of genes in the stem, (E) FPKM values of genes in the leaf, (F) FPKM values of genes in the root, (G) GC content, (H) ortho-QTL genes, and (I) syntenic relationships of E. fulvus genes.

Figure 2

Evolutionary characteristics of the E. fulvus genome. (A) Hi-C interaction map. (B) Depiction of LTR evolution. (C) Representation of whole-genome duplication (WGD) events during E. fulvus evolution based on synonymous substitution rates (Ks). (D) Density distribution of Ks values for paired genes calculated within each syntenic block between E. fulvus and other species. (E) Genome evolution based on single-copy orthologs of 10 species from the PACMAD and BOP clades. A. thaliana was used as an outgroup. Numbers in green indicate expanded gene families, and numbers in red indicate contracted gene families.

Figure 3

Inter-genomic analyses. (A) Correspondence between chromosomes of E. fulvus and S. spontaneum. (B) Chromosome rearrangements based on AGK gene composition across E. fulvus and other species. (C) Collinearity and syntenic relationships among E. fulvus, O. sativa, and S. spontaneum.

Figure 4

Maximum likelihood (ML) phylogenetic tree based on genome-wide SNPs. The magenta clade contains E. fulvus (sky-blue star), and the blue clade includes accessions of S. officinarum (red star), S. robustum (green square), S. barberi (blue triangle), S. sinense (sky-blue triangle), and S. spontaneum (magenta circle). The red clade contains accessions of E. rockii (green star), the green clade contains N. porphyrocoma (red triangle), the sky-blue clade contains S. arundinaceum (green circle), and the yellow clade contains Miscanthus (red square and magenta star).

Figure 5

Representation of sucrose and starch metabolism. (A) Pathway depicting starch biosynthesis. (B) Pathway depicting starch degradation. (C) Depiction of sucrose biosynthesis during photosynthesis. (D) Depiction of sucrose biosynthesis from end products of starch degradation. (E) Depiction of sucrose biosynthesis from end products of gluconeogenesis I. (F) Heat map showing expression patterns of important genes involved in starch and sucrose metabolism. RC, LC, and SC = root, leaf, and stem control (under normal conditions of 28°C). R24, L24, and S24 = root, leaf, and stem after 24 h at 4°C. R72, L72, and S72 = root, leaf, and stem after 72 h at 4°C. (G) Diagram representing transcription, translation, and trafficking of EfSWEET proteins to their target destinations. Thirteen EfSWEET proteins were targeted to the plasma membrane and three to the vacuole. These EfSWEET proteins transport either disaccharides like sucrose (orange text, orange transporter in diagram) or monosaccharides like glucose and fructose (purple text and purple transporter in diagram).

Figure 6

Structural details of EfSWEET proteins. (A) Tyr61, Tyr183, Asp189, and V192 are vital for extracellular gating. (B) The proline tetrad (P27, P47, P149, and P166) is essential for the transport route. (C) Tyr48 and Leu169 are responsible for intracellular gating, and Asn76, Trp180, and Asn196 surround the binding pocket to facilitate transport. Residues are numbered according to 5XPD, and structure figures were created in PyMOL. (D) The mono- and disaccharide substrate-specific residues highlighted are shown in yellow (23, 54, 145, and 176) and mapped on the 5XPD structure indicated by the gray ribbon, with its substrate analog shown in a stick representation. (F) Conservation surface mapping of EfSWEET transmembrane domain, colored according to the conservation score derived from the MSA. (E and G) Structural superimposition of the transmembrane domain of the modeled 3D structure of EfSWEET proteins with AtSWEET13 (PDBID:5XPD). The 5XPD and its disaccharide substrate are shown in orange, whereas all sweet proteins are shown in gray cartoons. (E) Modeled with Alphafold2. (G) Modeled with Phyre2 homology models.

Figure 7

Adaptive response of E. fulvus under cold stress. (A) Putative cold tolerance model of E. fulvus. (B) Heatmap showing expression of important cold-regulated genes. (C) Neighbor-joining phylogenetic tree of the functional domains of E. fulvus DREB proteins. L24, S24, and R24 represent the leaf, stem, and root under cold stress at 24 h, and L72, S72, and R72 represent the leaf, stem, and root under cold stress at 72 h.