Identification and expression analysis of the cysteine synthase (CSase) gene family in Brassica napus L. under abiotic stress

Abstract

Cysteine is the first organic compound identified in plants that contains both sulfur and nitrogen. It serves as a precursor for sulfur-containing metabolites such as methionine, glutathione (GSH), and Fe-S clusters, all of which play crucial roles in plant growth, development, and stress responses. Cysteine synthase (CSase) catalyzes the final step in cysteine biosynthesis; therefore, studying the CSase gene family is essential for understanding its role in plant abiotic stress tolerance. Using the CSase protein sequences of Arabidopsis thaliana as seed sequences and integrating protein domain information, 69 members of the BnCSase gene family were identified from the whole genome of Brassica napus ZS11. These members were analyzed for their physicochemical properties, phylogenetic relationships, covariance relationships, protein-protein interaction (PPI) networks, associated miRNAs, and SNP variations. Based on transcriptome data, the expression patterns of BnCSase genes under different abiotic stress treatments were investigated. Furthermore, the relative expression levels of several BnCSase genes were analyzed under salt, alkali, low nitrogen, and drought stress treatments at 0, 6, 12, and 24 h using qRT-PCR to explore their roles in abiotic stress tolerance in B. napus. The results revealed distinct expression patterns of BnCSase genes in response to different abiotic stress signals, indicating stress-specific responses in B. napus. This study provides a theoretical basis for elucidating the functions and molecular genetic mechanisms of the BnCSase gene family in abiotic stress tolerance in rapeseed.

Keywords: Brassica napus L.; Abiotic stress; Cysteine synthase; Gene family.

Fig. 1

Chromosomal location of the BnCSases in the B. napus genome

Fig. 2

Phylogenetic tree of CSase proteins. (A) Phylogenetic tree of CSase proteins in B. napus and (A) thaliana. (B) Phylogenetic tree of CSase proteins in (B) napus, B. rapa, and B. oleracea

Fig. 3

Gene structure analysis of CSase family in B.napus. (A) Conserved motifs of BnCSase family proteins. (B) Pfam structure of BnCSase family proteins. (C) Promoter cis-acting element of BnCSases. (D) The mRNA structure encoded by the BnCSases

Fig. 4

Classification of Cis-acting Elements in the Members of the BnCSase Gene Family

Fig. 5

Collinearity of CSase genes in B. napus, B. rapa, and B. oleracea

Fig. 6

Collinearity of BnCSases. The circles in the figure from inside to outside represent the unknown base (a) N ratio, (b) gene density, (c) GC ratio, (d) GC skew, and (e) chromosome length of the B. napus genome

Fig. 7

The selective evolutionary pressure on BnCSases. Blue dots represent the Ka/Ks values within BnCSase genes, and red dots represent the corresponding Ka and Ks values of BnCSase genes within species

Fig. 8

BnCSase proteins interaction network analysis

Fig. 9

Analysis of the interaction between microRNA and BnCSases

Fig. 10

Analysis of expression patterns of BnCSases under abiotic stress treatments

Fig. 11

Statistics of CSase gene variation in B. napus

Fig. 12

SNP variations in the BnCSase48 frameshift mutant. (A) Germination shoot length under salt stress. (B) FPKM expression level of BnCSase48 in seeds

Fig. 13

The CT value distribution of three candidate reference genes under abiotic stress

Fig. 14

The relative expression of BnCSase1, BnCSase14, BnCSase17, BnCSase27, BnCSase30, BnCSase31, BnCSase32, BnCSase48, BnCSase52, BnCSase64, BnCSase67, BnCSase68, in leaves under 1.2% (w/v) NaCl, 0.2% (w/v) NaHCO₃, Low Nitrogen (Table 2), and 20% (w/v) PEG 6000 after 0, 6 h, 12 h and 24 h. Data represent the mean ± standard error for threebiological experiments. Statistical differences between treatment groups were determined using one-way ANOVA. *: significant differences between treatments at p ≤ 0.05. **: significant differences between treatments at p ≤ 0.01