The Strawberry FaWRKY1 Transcription Factor Negatively Regulates Resistance to Colletotrichum acutatum in Fruit Upon Infection

Abstract

Strawberry ( Fragaria × ananassa ) is a major food crop worldwide, due to the flavor, aroma and health benefits of the fruit, but its productivity and quality are seriously limited by a large variety of phytopathogens, including Colletotrichum spp. So far, key factors regulating strawberry immune response remain unknown. The FaWRKY1 gene has been previously proposed as an important element mediating defense responses in strawberry to Colletotrichum acutatum . To get further insight into the functional role that FaWRKY1 plays in the defense mechanism, Agrobacterium -mediated transient transformation was used both to silence and overexpress the FaWRKY1 gene in strawberry fruits ( Fragaria × ananassa cv. Primoris), which were later analyzed upon C. acutatum inoculation. Susceptibility tests were performed after pathogen infection comparing the severity of disease between the two agroinfiltrated opposite halves of the same fruit, one half bearing a construct either for FaWRKY1 overexpression or RNAi-mediated silencing and the other half bearing the empty vector, as control. The severity of tissue damage was monitored and found to be visibly reduced at five days after pathogen inoculation in the fruit half where FaWRKY1 was transiently silenced compared to that of the opposite control half and statistical analysis corroborated a significant reduction in disease susceptibility. Contrarily, a similar level of susceptibility was found when FaWRKY1 overexpression and control fruit samples, was compared. These results unravel a negative regulatory role of FaWRKY1 in resistance to the phytopathogenic fungus C. acutatum in strawberry fruit and contrast with the previous role described for this gene in Arabidopsis as positive regulator of resistance against the bacteria Pseudomonas syringae . Based on previous results, a tentative working model for WRKY75 like genes after pathogen infection is proposed and the expression pattern of potential downstream FaWRKY1 target genes was also analyzed in strawberry fruit upon C. acutatum infection. Our results highlight that FaWRKY1 might display different function according to species, plant tissue and/or type of pathogen and underline the intricate FaWRKY1 responsive defense regulatory mechanism taking place in strawberry against this important crop pathogen.

Genome-Wide Analysis of Members of the WRKY Gene Family and Their Cold Stress Response in Prunus mume

“…A total of 58 PpWRKY genes have been identified in the genome of Prunus persica [16]. WRKY proteins have been verified to be essential in the defense response to various biotic and abiotic stresses, including low or high temperature, mechanical damage, and pathogenic infection [17,18,19,20,21,22,23]. In addition, there is a great deal of definite evidence indicating that the regulation of plant development and metabolism also requires WRKYs.…”

Strawberry FaWRKY25 Transcription Factor Negatively Regulated the Resistance of Strawberry Fruits to Botrytis cinerea

“…Studies have shown that the WRKY gene plays an important role in strawberry disease resistance. According to previous research, we know that FaWRKY1 is related to strawberry C. acutatum [ 34 , 35 ], and that FvWRKY42 is a positive regulator of the resistance of wild strawberries against powdery mildew [ 37 ]. In 2016, 59 WRKY genes at different stages of fruit development were found in forest strawberry by transcriptome sequencing [ 31 ].…”

“…The separated strains were cultured in a culture medium with potato dextrose agar at 20 °C and a photoperiod of 16/8. Before the strains were inoculated in the strawberries, we cultured them in a medium with strawberry agar (500 g/L of ground strawberries and 1.5% of bacterial agar) to enhance their invasiveness [ 35 ]. Mycelia were scraped from the surface of the strains after 2 weeks of culturing, and conidia suspension was prepared in sterile distilled water with 0.03% ( v/v ) of Tween-80 at a concentration of 10 6 spores per milliliter.…”

“…Currently, 59 WRKY genes at different fruit growth stages of forest strawberries and 62 WRKY genes responding to biotic and abiotic stress have been identified [ 31 , 32 ]; moreover, 47 WRKY genes have been identified in octoploid strawberries [ 33 ]. FaWRKY1 was the first strawberry WRKY identified as a mediator of defense response against Colletotrichum acutatum in cultivated strawberry [ 34 , 35 ]. FaWRKY1 encodes an AtWRKY75 -like transcription factor type IIc, which is upregulated after Colletotrichum infection and responds to defense-related hormones such as SA, JA, ABA, and wounding, with its expression being dependent on strawberry cultivar and tissue [ 35 , 36 ].…”

“…FaWRKY1 was the first strawberry WRKY identified as a mediator of defense response against Colletotrichum acutatum in cultivated strawberry [ 34 , 35 ]. FaWRKY1 encodes an AtWRKY75 -like transcription factor type IIc, which is upregulated after Colletotrichum infection and responds to defense-related hormones such as SA, JA, ABA, and wounding, with its expression being dependent on strawberry cultivar and tissue [ 35 , 36 ]. A recent study has shown that FvWRKY42 is a positive regulator of the resistance of wild strawberries against powdery mildew [ 37 ].…”