Association of variation in the sugarcane transcriptome with sugar content

Abstract

Background: Sugarcane is a major crop of the tropics cultivated mainly for its high sucrose content. The crop is genetically less explored due to its complex polyploid genome. Sucrose synthesis and accumulation are complex processes influenced by physiological, biochemical and genetic factors, and the growth environment. The recent focus on the crop for fibre and biofuel has led to a renewed interest on understanding the molecular basis of sucrose and biomass traits. This transcriptome study aimed to identify genes that are associated with and differentially regulated during sucrose synthesis and accumulation in the mature stage of sugarcane. Patterns of gene expression in high and low sugar genotypes as well as mature and immature culm tissues were studied using RNA-Seq of culm transcriptomes.

Results: In this study, 28 RNA-Seq libraries from 14 genotypes of sugarcane differing in their sucrose content were used for studying the transcriptional basis of sucrose accumulation. Differential gene expression studies were performed using SoGI (Saccharum officinarum Gene Index, 3.0), SAS (sugarcane assembled sequences) of sugarcane EST database (SUCEST) and SUGIT, a sugarcane Iso-Seq transcriptome database. In total, about 34,476 genes were found to be differentially expressed between high and low sugar genotypes with the SoGI database, 20,487 genes with the SAS database and 18,543 genes with the SUGIT database at FDR < 0.01, using the Baggerley's test. Further, differential gene expression analyses were conducted between immature (top) and mature (bottom) tissues of the culm. The DEGs were functionally annotated using GO classification and the genes consistently associated with sucrose accumulation were identified.

Conclusions: The large number of DEGs may be due to the large number of genes that influence sucrose content or are regulated by sucrose content. These results indicate that apart from being a primary metabolite and storage and transport sugar, sucrose may serve as a signalling molecule that regulates many aspects of growth and development in sugarcane. Further studies are needed to confirm if sucrose regulates the expression of the identified DEGs or vice versa. The DEGs identified in this study may lead to identification of genes/pathways regulating sucrose accumulation and/or regulated by sucrose levels in sugarcane. We propose identifying the master regulators of sucrose if any in the future.

Keywords: High and low sugar genotypes; Sucrose; Sucrose genes; Sugarcane transcriptome; Transcriptome.

Fig. 1

Workflow for transcriptome sequencing, RNA-Seq experiments and the identification of differentially expressed transcripts. High and low sugar genotypes as well as mature and immature culm samples of sugarcane genotypes were compared in the study

Fig. 2

Volcano plot depiction of the DGEs in different groups using a, b, c) Saccharum officinarum gene indices, SoGI; d, e, f) Sugarcane Iso-Seq transcriptome database, SUGIT; g, h, i) Sugarcane assembled sequences, SAS. LST, low sugar top internode; LSB, low sugar bottom internode; HST, high sugar top internode; HSB, high sugar bottom internode; In HSB vs LSB, HSB shows upregulation of transcripts, whereas in LST vs LSB, LST shows upregulation of transcripts; in HST vs HSB, HSB shows a clear upregulation of transcripts using SAS database, though very few transcripts showed differential expression in other two databases. Please note that there was no DGE detected in HST vs LST

Fig. 3

Graphical representation of DGE experiments with three different databases, Saccharum officinarum gene indices, SoGI; Sugarcane Iso-Seq transcriptome database, SUGIT; Sugarcane assembled sequences, SAS. LST, low sugar top internode; LSB, low sugar bottom internode; HST, high sugar top internode; HSB, high sugar bottom internode

Fig. 4

Venn diagrams depicting the common and unique DEGs obtained from three different comparisons between mature and immature culm tissues of high and low sugar genotypes. LST, low sugar top internode; LSB, low sugar bottom internode; HST, high sugar top internode; HSB, high sugar bottom internode; a, b and c), RNA-Seq using Saccharum officinarum gene indices, SoGI; Sugarcane Iso-Seq transcriptome database, SUGIT; Sugarcane assembled sequences, SAS, respectively

Fig. 5

A schematic representation of global and differential gene expression between and within the groups using three databases. a, b, c) Saccharum officinarum gene indices, SoGI (121,342 ESTs); d, e, f) SUGIT-Sugarcane Iso-Seq transcriptome database (107,598 contigs) and f, g, h) Sugarcane assembled sequences, SAS (43,141) contigs. LST, low sugar top internode; LSB, low sugar bottom internode; HST, high sugar top internode; HSB, high sugar bottom internode. The numbers within the intersection are the number of DEGs between the groups compared. The numbers in the circle gives the number of transcripts expressed against the reference database with a RPKM cut off of >0

Fig. 6

Graphical representation of the expression pattern of sucrose synthase transcripts in various comparisons. LST, low sugar top internode samples; LSB, low sugar bottom internode samples; HST, high sugar top internode samples; HSB, high sugar bottom internode samples; T-top tissue; B-bottom tissue. Shown here are the sucrose synthase (SuSy) transcripts from Saccharum officinarum gene indices, SoGI database, showing differential expression in top two comparisons. (a) HSB vs LSB; (b) LST vs LSB, while there is no differential expression in case of lower two comparisons (c) HST vs HSB; (d) HST vs LST at FDR<0.01. X-axis shows the genotypes while Y-axis represents RPKM values

Fig. 7

Graphical representation of the expression pattern of some of SuSy transcripts in various comparisons. LST, low sugar top internode sample; LSB, low sugar bottom internode sample; HST, high sugar top internode sample; HSB, high sugar bottom internode sample; T-top tissue; B-bottom tissue. Shown here are the sucrose phosphate synthase III, sucrose non-fermented related protein kinase, sucrose transport protein, impaired sucrose induction 1- like protein transcripts from Saccharum officinarum gene indices, SoGI database, showing differential expression in top two comparisons (a) HSB vs LSB; (b) LST vs LSB while there is no differential expression in lower two comparisons (c) HST vs HSB; (d) HST vs LST at FDR < 0.01. X-axis shows the genotypes while Y-axis represents RPKM values