The sucrose synthase gene family in blueberry (Vaccinium darrowii): functional insights into the role of VdSUS4 in salt stress tolerance

Abstract

Introduction: The sucrose synthase (SUS), a crucial enzyme in the sucrose metabolism, is encoded by a multigene family in plant kingdom.

Methods: In our study, we utilized bioinformatics tools to identify and characterize the members of the SUS gene family within the blueberry genome. Our analysis encompassed the physicochemical properties, gene structures, conserved motifs, promoter cis-acting elements, chromosomal locations, evolutionary relationships and expression profiles of these family members, allowing us to predict their potential functions.

Results: We identified seven distinct SUS genes, mapped across six chromosomes, showcasing the complexity of this gene family in blueberries. Phylogenetic analysis, constructed through a multi-species phylogenetic tree, revealed that the SUS gene family can be categorized into three subfamilies: SUS I, SUS II and SUS III. Notable variations were observed among the VdSUS gene family members, particularly in the number of amino acids, molecular weight, isoelectric point, and hydrophobicity of the encoded proteins. Intriguingly, our predictive analysis of the promoter regions of VdSUS genes uncovered a wealth of cis-acting elements linked to light response, hormonal regulation, and stress responses, suggesting a role in adaptive mechanisms. Expression studies indicated that VdSUS genes were highly expressed in fruit tissues, with the application of exogenous sucrose leading to significant downregulation of VdSUS2, VdSUS3 and VdSUS6. Furthermore, the expression of VdSUS genes was found to be responsive to abiotic stresses, such as salt, drought, and low temperatures, with varying degrees of upregulation or downregulation observed. Most notably, the overexpression of VdSUS4 in Arabidopsis thaliana resulted in enhanced tolerance to salt stress.

Discussion: These findings have shed new light on the multifaceted roles of VdSUS gene family members in the complex physiological processes of blueberries, highlighting their potential in the context of stress adaptation and fruit development.

Keywords: bioinformatics; blueberry; salt stress; sucrose synthase; tissue-specific expression.

Figure 1

Phylogenetic analysis of the SUS gene family. VdSUSs were labeled with red stars.

Figure 2

Characteristics of VdSUS proteins. (A) Chromosomal localization. (B) Collinearity relationship between blueberry and Arabidopsis SUS genes. (C) Collinearity relationship between blueberry and rice SUS genes. (D) Collinearity relationship between blueberry and grape.

Figure 3

Structural features of VdSUS sequences. (A) Phylogenetic tree of the VdSUS protein family, where I represents SUS I, II represents SUS II, and III represents SUS III. (B) Gene structure features of the VdSUS genes family. (C) Motif analysis of the VdSUS proteins. (D) Conserved motif sequences.

Figure 4

Analysis of cis-regulatory elements in the promoter regions of SUS genes in blueberry. (A) Phylogenetic tree of the VdSUS protein family in blueberry. (B) The positions of cis-regulatory elements in the promoter regions of VdSUS genes in blueberry. (C) Statistical analysis of the number of cis-regulatory elements in the promoter regions of VdSUS genes in blueberry.

Figure 5

Spatial expression patterns analysis of VdSUS genes. (A) Phenotypes of extracted blueberry samples in different organs, tissues and developmental stages. YFl, young flowers; MFl, mature flowers; EGF, early green fruits; LGF, late green fruits; MF, mature fruits; YS, young stems; MS, mature stems; YL, young leaves; ML, mature leaves; R, roots. (B) Expression levels of VdSUS family genes.

Figure 6

Expression patterns of VdSUS genes in response to abiotic stress treatment. (A) Phenotypic charts before and after drought treatment. (B) Expression levels of VdSUS genes under drought treatment conditions. (C) Phenotypic charts before and after NaCl treatment. (D) Expression levels of VdSUS genes under NaCl treatment conditions. (E) Phenotypic charts before and after low temperature treatment. (F) Expression levels of VdSUS genes under low temperature treatment conditions. Values are the average ± standard deviation of three biological replicates. The transcription levels of VdSUS genes at 0 days and 0 hours were set as “1”. P-values < 0.05, 0.01 and 0.001 are denoted by “*”, “**” and “***” respectively (Student’s t-test).

Figure 7

Overexpression of VdSUS4 enhanced salt stress tolerance in Arabidopsis. (A) Germination of WT and VdSUS4 overexpressing lines on media with or without 100 mM NaCl, photographed after 15 d. (B) Root length measurement of 15-day-old WT and VdSUS4 overexpressing lines. (C) Fresh weight measurement of 15-day-old WT and VdSUS4 overexpressing lines. Values are the mean ± standard deviation of three biological replicates. p-values < 0.05 are represented by “*” (Student’s t-test).

Figure 8

VdSUS4 enhances salt tolerance by strengthening the plant antioxidant system. (A, B) Germination rate of WT and transgenic seeds. (C) Phenotypic comparison of WT and transgenic lines after treatment with 150 mM NaCl for 8 d. (D) POD activity. (E) SOD activity. (F) CAT activity. (G) MDA content. Values are the mean ± standard deviation of three biological replicates. p-values < 0.01 are represented by “**” (Student’s t-test). p-values < 0.05 are represented by "*".