Changes in soluble sugars and the expression of sugar transporter protein genes in strawberry crowns responding to Colletotrichum fructicola infection

Abstract

Strawberry (Fragaria × ananassa) production has been greatly hampered by anthracnose crown rot caused by Colletotrichum fructicola. Crown, the modified stem of strawberry, is a sink organ involved in sugar allocation. Some Sugar Transport Proteins (STPs) are involved in competition for sugars between pathogen and host. However, the chemical nature and involvement of strawberry STPs (FaSTPs) in crown rot development is largely elusive. To reveal how strawberry alters soluble sugars and upregulates STPs in responses to C. fructicola, high performance liquid chromatograph and FaSTP expression analysis were performed in the crowns of three strawberry varieties, following a genome-wide identification of FaSTPs. Both C. fructicola and mock treatment/control changed glucose, fructose and sucrose accumulation in strawberry crowns. With increasing infection duration, the hexose/sucrose ratio increased in all varieties; no such trend was clearly visible in mock-treated plants. A total of 56 FaSTP loci scattered across four subgenomes were identified in octoploid strawberry, and most of the protein products of these genes had a preferential location on plasma membrane. Putative fungal elicitor responsive cis-elements were identified in the promoters of more than half FaSTPs. At least eight members were upregulated in strawberry crowns during C. fructicola invasion. Of them, FaSTP8 expression was markedly enhanced in three varieties at all time points except for 3 dpi in 'Jiuxiang'. RNAseq data retrieval further validated the expression responses of FaSTPs to Colletotrichum spp. In summary, this work identified several FaSTP candidate genes responsive to Colletotrichum fructicola invasion, demonstrated changes in soluble sugar levels in strawberry crowns as a result of infection, and laid the groundwork for future efforts to engineer strawberry resistance to Colletotrichum spp.

Supplementary information: The online version contains supplementary material available at 10.1007/s12298-024-01523-9.

Keywords: Anthracnose crown rot (ACR); Colletotrichum fructicola; Fragaria × ananassa; Soluble sugar; Sugar transport protein (STP).

Fig. 1

Crown rot development in strawberry upon C. fructicola invasion. A Typical symptoms in crown transections and whole plants under 25 °C. B The percentage of disease grade (%) derived from lesion size on crown transversal surface. C qPCR for the colonization of C. fructicola in strawberry crowns. Different lowercase letters indicated the significant differences between mock and treatment at distinct phases in certain genotype (P < 0.05). Different uppercase letters represented the significant differences among the effects of genotype × phase (two-way ANOVA, Tukey-test)

Fig. 2

Main soluble sugars in strawberry crowns revealed with HPLC. A Contents of fructose, glucose, sucrose and total soluble sugars in the crown tissues. B Composition of total soluble sugars in strawberry crowns post mock-treatment or Cf-infection. C Changes in H/S (hexose-to-sucrose ratio) and G/T (glucose-to-total soluble sugars in strawberry crowns. Six raw data were used in (A), and the average of six raw data was used in (B) and (C). Asterisks for the significant differences between mock and Cf-invasion (pairwise t-test); different lowercase and uppercase letters above the box charts indicating the differences in the dynamic and genotypic effects on sugar contents among mock or Cf-invasion, respectively (one-way ANOVA test, Waller–Duncan method)

Fig. 3

The phylogenetic relationships of Sugar transport protein (STP) proteins from Arabidopsis thaliana, diploid Fragaria vesca, and octoploid F. × ananassa strawberry. The tree consisting of 14 AtSTPs, 22 FvSTPs and 56 FaSTPs was clustered into four groups (I, II, III and IV). The length of branches indicates the relative phylogenetic relationship, and the bootstrap values near branches show confidence

Fig. 4

Chromosomal distribution of FaSTPs in the genome of octoploid strawberry F. × ananassa. The localization information was obtained from GDR, and visualized in MapChart software. The length of each chromosome was indicated below in megabases (Mb). Arrows and lines indicated the transcription orientation and the transcription starting site for each locus

Fig. 5

Cis-elements predicted in the 2500 bp promoter regions of STP genes in Fragaria vesca and F. × ananassa. The promoter sequences of 78 STP genes (56 from F. × ananassa and 22 from F. vesca) were analyzed at online PlantCare tool. Geometric shapes of different colors and shapes represent elements involved in different processes

Fig. 6

RT-qPCR analysis of strawberry FaSTP genes upon C. fructicola invasion in the crown tissues of cultivars ‘Hogyoku’, ‘Sweet Charlie’ and ‘Jiuxiang’. The relative transcript levels of STP genes were normalized with two reference genes EF1a and GAPDH2 (Amil-Ruiz et al. 2013) and reported as the mean of six replicates ± SE from two independent experiments. Pairwise T-test analysis was conducted for mock and C.f-invasion and indicated as asterisk (*P < 0.05; **P < 0.01; ***P < 0.001) for certain gene/genotype/phase. Distinct lowercase and uppercase letters indicated significant differences (P < 0.05) among relative mRNA levels during the dynamic process for mock and C.f-invasion, respectively

Fig. 7

RNA-seq data indicated the variations in the expression responses of STP family genes in octoploid strawberry upon Colletotrichum spp. infection. The heat map was produced with Log2-transformed RPKM (mapping readings per kilobytes) values for each transcript. A In leaves from cv. ‘Jiuxiang’ infected with C. fructicola (Zhang et al. 2018). B In leaves from cv. ‘Yanli’ and ‘Benihoppe’ with C. gloeosporioides (Wang et al. 2017). C In crowns from cv. ‘Elyana’ and ‘Festival’ infected with C. gloeosporioides (Chandra et al. 2021)