Genome-wide characterization of cys-tathionine-β-synthase domain-containing proteins in sugarcane reveals their role in defense responses under multiple stressors

Abstract

Cys-tathionine-β-synthase (CBS) domain-containing proteins (CDCPs) are essential for regulating plant responses to various biotic and abiotic stressors. This study describes the systematic identification and characterization of CDCP family genes in Saccharum spontaneum. A total of 95 SsCDCP genes and eight phylogenetic groups were identified that were distributed over 29 chromosomes of the AP85-441 genome. Most (78/95) SsCDCPs underwent fragment duplication events, and 64 gene pairs were located in synteny blocks. Expression profiling of nine ShCDCPs was also carried out in the Saccharum spp. cultivars ROC22 and MT11-611 that are resistant and susceptible to red stripe, respectively, in response to: (i) Infection by the bacterial pathogen Acidovorax avenue subsp. avenae (Aaa); (ii) abiotic stressors (drought and salinity); and (iii) exogenous salicylic acid (SA) treatment. Members of one gene pair (ShCBSD-PB1-5A and ShCBSD-PB1-7A-1) with a fragment duplication event acted as negative regulators in sugarcane under four stresses, as supported by the significantly decreased expression levels of ShCBSD-PB1-5A (23-83%) and ShCBSD-PB1-7A-1 (15-75%) at all-time points, suggesting that they have functional redundancy. Genes in another pair, ShCBS-4C and ShCBS-4D-1, which have a fragment duplication event, play opposing regulatory roles in sugarcane exposed to multiple stresses, particularly Aaa and NaCl treatments. ShCBS-4C expression was significantly decreased by 32-77%, but ShCBS-4D-1 expression was dramatically upregulated by 1.2-6.2-fold in response to Aaa treatment of both cultivars across all-time points. This result suggested that both genes exhibited functional divergence. Meanwhile, the expression of SsCBSDCBS-5A was significantly upregulated in ROC22 by 1.4-4.6-fold in response to the four stressors. These findings provide important clues for further elucidating the function of ShCDCP genes in sugarcane responding to a diverse range of stresses.

Keywords: Acidovorax avenue subsp. avenae; CBS domain containing proteins (CDCPs); abiotic stress; defense response; gene expression; sugarcane.

FIGURE 1

Unrooted phylogenetic tree of SsCDCPs identified in S. spontaneum AP85-441. The tree was conducted using the maximum likelihood (ML) method with 1,000 bootstrap replicates.

FIGURE 2

Gene structure (A) and conserved motif (B) of SsCDCPs in S. spontaneum. (A) Untranslated 5′- and 3′-regions, CDS and conserved domains are displayed with different colored boxes. (B) Motifs 1–10 are displayed as different colored boxes. Gene and protein lengths can be estimated using the scales at the bottom.

FIGURE 3

Heatmap of cis-regulatory elements in promoter sequences (2 kb) of SsCDCPs from S. spontaneum.

FIGURE 4

Chromosomal localization and gene replication relationship of SsCDCPs in S. spontaneum. Ribbon links (Red lines) indicate segmental duplication events between genes. Chromosome numbers are indicated inside the outer circle. The gene names on each chromosome are indicated on the outer circle.

FIGURE 5

Heatmap of transcript expression profiles of ShCDCPs in two cultivars, ROC22 (resistant to red stripe, R) and MT11-610 (susceptible to red stripe, S) based on RNA-Seq data. The gene names are indicated in the outer circle and the expression values mapped to a color gradient from low (blue) to high (red) expression level are shown inside the outer circle.

FIGURE 6

Expression levels of nine ShCDCPs in the cultivars ROC22 and MT11-610 with Aaa infection based on RT-qPCR data. Means ± standard errors are shown. The same letters atop the bars indicate no significant difference at the 5% level.

FIGURE 7

Expression levels of nine ShCDCPs in ROC22 and MT11-610 cultivars treated with NaCl, PEG6000 or exogenous SA treatment based on RT-qPCR data. Means ± standard errors are shown. The same letters atop the bars indicate no significant difference at the 5% level. Rows show the same gene and columns correspond to stressor. hpt, h post treatment.