Identification of YABBY Transcription Factors and Their Function in ABA and Salinity Response in Nelumbo nucifera

Abstract

The plant-specific transcription factor family YABBY plays important roles in plant responses to biotic and abiotic stresses. Although the function of YABBY has been identified in many species, systematic analysis in lotus (Nelumbo nucifera) is still relatively lacking. The present study aimed to characterize all of the YABBY genes in lotus and obtain better insights into NnYABBYs in response to salt stress by depending on ABA signaling. Here, we identified nine YABBY genes by searching the whole lotus genome based on the conserved YABBY domain. Further analysis showed that these members were distributed on six different chromosomes and named from YABBY1 to YABBY9, which were divided into five subgroups, including YAB1, YAB2, YAB5, INO, and CRC. The analysis of cis-elements in promotors revealed that NnYABBYs could be involved in plant hormone signaling and plant responses to abiotic stresses. Quantitative real-time PCR (qRT-PCR) showed that NnYABBYs could be up-regulated or down-regulated by ABA, fluridone, and salt treatment. Subcellular localization indicated that NnYABBY4, NnYABBY5, and NnYABBY6 were mainly localized in the cell membrane and cytoplasm. In addition, the intrinsic trans-activity of NnYABBY was tested by a Y2H assay, which revealed that NnYABBY4, NnYABBY5, and NnYABBY6 are deprived of such a property. This study provided a theoretical basis and reference for the functional research of YABBY for the molecular breeding of lotus.

Keywords: ABA; Nelumbo nucifera; YABBY; salt stress; transcription factor.

Figure 1

Phylogenetic tree of interspecific YABBY relationships involving lotus, potato, soybean, rice, wheat, and Arabidopsis.

Figure 2

Gene structures and motif analyses of NnYABBYs. (A) Gene structures of NnYABBYs; (B) Conserved motif analysis of NnYABBYs. Motifs were analyzed by using the MEME website.

Figure 3

Chromosomal locations of NnYABBY genes in lotus. The yellow represents the chromosome and the chromosome number is on top of the bars. There is no correlation between the length of the bars and the size of the chromosomes. The map distance is on the left side of the chromosomes and its unit is megabases (Mb).

Figure 4

Collinearity analysis of the NnYABBY gene family in the lotus. (A) Intra-specific collinearity analysis. (B) Collinearity analysis between Nelumbo nucifera Gaertn and Arabidopsis thaliana. Gene pairs are represented by the red lines.

Figure 5

Tertiary structure of NnYABBY proteins. Through the protein sequence query, the default parameter Settings in SWISS are used to map the tertiary structures.

Figure 6

Interaction network of NnYABBY in lotus. The network type was a full STRING network. Different line colors indicate the type of interaction evidence.

Figure 7

cis-acting regulatory elements analysis of NnYABBY genes in lotus. The upstream 2000 bp of each NnYABBY member was used as the promoter sequence to query and analyze the cis-acting regulatory elements in PlantCARE. Different colors indicate different cis-acting regulatory elements.

Figure 8

qRT-PCR analysis of NnYABBY in lotus leaves under different hormone treatments. (A) Expression levels of NnYABBYs in lotus leaves under ABA stress. (B) Expression levels of NnYABBYs in leaves sprayed with ABA and FL. (C) Expression levels of NnYABBYs in lotus leaves under NaCl stress. (*) represents p values of <0.05 and is considered statistically significant. (**) represents p values of <0.01 and is considered highly statistically significant.

Figure 9

Subcellular localization of the YABBY proteins in lotus. The picture above is the control and that below shows NnYABBY5, NnYABBY6, and NnYABBY7. Scale bar = 20 μm.

Figure 10

Transcription activation analysis of the three selected NnYABBY proteins. (A) Schematic representation of the construction of the fusion plasmid. (B) Transcription activation analysis of the 3 selected NnYABBY proteins. pGBKT7 was used as the control.