Dynamics of gene expression during development and expansion of vegetative stem internodes of bioenergy sorghum

Abstract

Background: Bioenergy sorghum accumulates 75% of shoot biomass in stem internodes. Grass stem internodes are formed during vegetative growth and elongate in response to developmental and environmental signals. To identify genes and molecular mechanisms that modulate the extent of internode growth, we conducted microscopic and transcriptomic analyses of four successive sub-apical vegetative internodes representing different stages of internode development of the bioenergy sorghum genotype R.07020.

Results: Stem internodes of sorghum genotype R.07020 are formed during the vegetative phase and their length is enhanced by environmental signals such as shade and floral induction in short days. During vegetative growth, the first visible and youngest sub-apical internode was ~0.7 cm in length, whereas the fourth fully expanded internode was ~5 cm in length. Microscopic analyses revealed that all internode tissue types including pith parenchyma and vascular bundles are present in the four successive internodes. Growth in the first two sub-apical internodes occurred primarily through an increase in cell number consistent with expression of genes involved in the cell cycle and DNA replication. Growth of the 3rd internode was associated with an increase in cell length and growth cessation in the 4th internode was associated with up-regulation of genes involved in secondary cell wall deposition. The expression of genes involved in hormone metabolism and signaling indicates that GA, BR, and CK activity decreased while ethylene, ABA, and JA increased in the 3rd/4th internodes. While the level of auxin appears to be increasing as indicated by the up-regulation of ARFs, down-regulation of TIR during development indicates that auxin signaling is also modified. The expression patterns of transcription factors are closely associated with their role during the development of the vegetative internodes.

Conclusions: Microscopic and transcriptome analyses of four successive sub-apical internodes characterized the developmental progression of vegetative stem internodes from initiation through full elongation in the sorghum genotype R.07020. Transcriptome profiling indicates that dynamic variation in the levels and action of GA, CK, IAA, BR, ethylene, ABA, and JA modulate gene expression and growth during internode growth and development. This study provides detailed microscopic and transcriptomic data useful for identifying genes and molecular pathways regulating internode elongation in response to various developmental and environmental signals.

Keywords: Bioenergy; Cell cycle; Hormone; Internode; Sorghum; Transcriptome.

Fig. 1

Microscopic cross sections of internodes of bioenergy sorghum inbred R.07020 at 60 DAP (days after planting). Internode1 (Int1) is the youngest sub-apical internode and internode 4 (Int4) is the oldest fully expanded internode. Stem sections were stained with safranin and alcian blue. Samples were from the middle section of each internode

Fig. 2

Microscopic longitudinal sections of internodes of bioenergy sorghum inbred R.07020 at 60 DAP (days after planting). Internode1 (Int1) is the youngest sub-apical internode, and internode 4 (Int4) is the oldest fully expanded internode. Stem sections were stained with safranin and alcian blue. Samples were from the middle section of each internode

Fig. 3

Venn diagram displaying genes expressed in four successive sub-apical internodes of bioenergy sorghum inbred R.07020. Int1 is the youngest sub-apical and Int4 is the oldest internode

Fig. 4

K-means clustering of genes differentially expressed between four sub-apical internodes of bioenergy sorghum inbred R.07020. Genes differentially expressed between at least any two of the four successive sub-apical internodes (Int1–Int4) were grouped into six clusters. Int1 is the youngest sub-apical and Int4 is the oldest internode. Expression level of each gene/transcript in RPKM was normalized prior clustering. Data are mean RPKM of three biological replicates. Bars represent standard deviation (SD)

Fig. 5

Heatmap of DNA and cell-related genes differentially expressed between four sub-apical internodes of bioenergy sorghum inbred R.07020. Int1 is the youngest sub-apical and Int4 is the oldest internode. Gene expression level in RPKM in each internode was normalized

Fig. 6

Heatmap of cell wall biosynthesis and modification genes differentially expressed between four sub-apical internodes of bioenergy sorghum inbred R.07020. Int1 is the youngest sub-apical and Int4 is the oldest internode. Gene expression level in RPKM in each internode was normalized

Fig. 7

Heatmap of secondary metabolism-related genes differentially expressed between four sub-apical internodes of bioenergy sorghum inbred R.07020. Int1 is the youngest sub-apical and Int4 is the oldest internode. Gene expression level in RPKM in each internode was normalized

Fig. 8

Heatmap of sugar transport, metabolism, and signaling genes differentially expressed between four sub-apical internodes of bioenergy sorghum inbred R.07020. Int1 is the youngest sub-apical and Int4 is the oldest internode. Gene expression level in RPKM in each internode was normalized

Fig. 9

Heatmap of genes encoding transcription factors differentially expressed between four sub-apical internodes of bioenergy sorghum inbred R.07020. Int1 is the youngest sub-apical and Int4 is the oldest internode. Gene expression level in RPKM in each internode was normalized

Fig. 10

Heatmap of plant hormone biosynthesis, transport, and signaling genes differentially expressed between four sub-apical internodes of bioenergy sorghum inbred R.07020. Int1 is the youngest sub-apical and Int4 is the oldest internode. Gene expression level in RPKM in each internode was normalized