Molecular Characterization and Expression Analysis of YABBY Genes in Chenopodium quinoa

Abstract

Plant-specific YABBY transcription factors play an important role in lateral organ development and abiotic stress responses. However, the functions of the YABBY genes in quinoa remain elusive. In this study, twelve YABBY (CqYAB) genes were identified in the quinoa genome, and they were distributed on nine chromosomes. They were classified into FIL/YAB3, YAB2, YAB5, INO, and CRC clades. All CqYAB genes consist of six or seven exons, and their proteins contain both N-terminal C2C2 zinc finger motifs and C-terminal YABBY domains. Ninety-three cis-regulatory elements were revealed in CqYAB gene promoters, and they were divided into six groups, such as cis-elements involved in light response, hormone response, development, and stress response. Six CqYAB genes were significantly upregulated by salt stress, while one was downregulated. Nine CqYAB genes were upregulated under drought stress, whereas six CqYAB genes were downregulated under cadmium treatment. Tissue expression profiles showed that nine CqYAB genes were expressed in seedlings, leaves, and flowers, seven in seeds, and two specifically in flowers, but no CqYAB expression was detected in roots. Furthermore, CqYAB4 could rescue the ino mutant phenotype in Arabidopsis but not CqYAB10, a paralog of CqYAB4, indicative of functional conservation and divergence among these YABBY genes. Taken together, these results lay a foundation for further functional analysis of CqYAB genes in quinoa growth, development, and abiotic stress responses.

Keywords: YABBY genes; abiotic stress; cis-elements; gene expression; quinoa.

Figure 1

Chromosomal distribution of CqYAB genes. Chromosome numbers are shown at the top of each chromosome. The scale bar represents chromosome size.

Figure 2

Phylogenetic tree of the YABBY family proteins from Arabidopsis, quinoa, spinach, beet, and rice. Gene names in red represent YABBYs from the quinoa genome. The five clades are indicated by differently colored backgrounds. At, Arabidopsis (Arabidopsis thaliana); Cq, quinoa (Chenopodium quinoa); So, spinach (Spinacia oleracea); Bv, beetroot (Beta vulgaris); and Os, rice (Oryza sativa).

Figure 3

Phylogenetic relationships, gene structures, protein motifs, and domains of CqYABs. (A) Phylogenetic tree constructed through the neighbor-joining method. (B) Gene structures of CqYAB genes. Blue boxes indicate exons, and black lines indicate introns. The scale bar represents the length of exon or intron. The double slash depicts an intron length longer than 5 kb. (C) Motif composition of CqYAB proteins. Ten motifs were organized, each with a different color. (D) Conserved N-terminal C2C2 zinc finger and C-terminal YABBY domains of CqYAB proteins. Stars indicate the conserved amino acids in the domains.

Figure 4

Cis-regulatory elements in the 3 kb promoter regions of CqYAB genes. The identified cis-elements were classified into six types. (A) Light-response elements. (B) Elements related to plant growth and development. (C) Phytohormone-response elements. (D) Stress-response elements. (E) Promoter-related elements. (F) Elements with unknown function. The names of cis-elements are listed on the left, and the numbers in different colors indicate the number of elements.

Figure 5

Expression responses of CqYAB genes under salt, drought, and Cd treatments. Two-week-old seedlings were treated with three stresses for 2 h and sampled for qRT-PCR. (A) Treatment with 200 mM NaCl. (B) Treatment with 15% PEG6000. (C) Treatment with 100 µM CdCl₂ (Cd). The relative expression levels of genes were calculated using the 2^−∆∆CT method. The expression level of the control (CK) was arbitrarily set to 1. Error bars indicate standard deviations of the mean value from three biological replicates and three technical replicates. Asterisks indicate significant differences in the expression between control and stress samples (* p < 0.05; ** p < 0.01; and *** p < 0.001).

Figure 6

Expression patterns of CqYAB genes under normal growth conditions. The expression levels of the 12 CqYAB genes were determined in different tissues at different developmental stages through qRT-PCR, and a fold change in the expression level was shown for each CqYAB. The expression level in flowers was arbitrarily set to 1. CqACTIN2 was used as an internal control. Different letters indicate significant differences in expression levels.

Figure 7

Complementation of the ino mutant phenotype by CqYAB4. (A) Silique morphology of different genotypes. (B) Dissected siliques in different genotypes, showing the developing seeds in siliques. (C) Silique length of different genotypes. (D) Seed number per silique of different genotypes. Capital letters indicate a significant difference (p < 0.01). The scale bar in the top left panel is 1 mm, whereas that in the top right panel is 0.5 mm.