Heterologous Expression of the AtNPR1 Gene in Olive and Its Effects on Fungal Tolerance.

Isabel Narváez,Elena Palomo-Ríos,Carlos Lopez-Herrera,Clara Pliego Prieto,Jose L. Trapero-Casas,Rafael M. Jiménez-Díaz,Fernando Pliego-Alfaro,Louis Fresta,Jose A. Mercado,Juan M. Arjona-Lopez

doi:10.3389/fpls.2020.00308

Abstract

The NPR1 gene encodes a key component of systemic acquired resistance (SAR) signaling mediated by salicylic acid (SA). Overexpression of NPR1 confers resistance to biotrophic and hemibiotrophic fungi in several plant species. The NPR1 gene has also been shown to be involved in the crosstalk between SAR signaling and the jasmonic acid-ethylene (JA/Et) pathway, which is involved in the response to necrotrophic fungi. The aim of this research was to generate transgenic olive plants expressing the NPR1 gene from Arabidopsis thaliana to evaluate their differential response to the hemibiotrophic fungus Verticillium dahliae and the necrotroph Rosellinia necatrix. Three transgenic lines expressing the AtNPR1 gene under the control of the constitutive promoter CaMV35S were obtained using an embryogenic line derived from a seed of cv. Picual. After maturation and germination of the transgenic somatic embryos, the plants were micropropagated and acclimated to ex vitro conditions. The level of AtNPR1 expression in the transgenic materials varied greatly among the different lines and was higher in the NPR1-780 line. The expression of AtNPR1 did not alter the growth of transgenic plants either in vitro or in the greenhouse. Different levels of transgene expression also did not affect basal endochitinase activity in the leaves, which was similar to that of control plants. Response to the hemibiotrophic pathogen V. dahliae varied with pathotype. All plants died by 50 days after inoculation with defoliating (D) pathotype V-138, but the response to non-defoliating (ND) strains differed by race: following inoculation with the V-1242 strain (ND, race 2), symptoms appeared after 44–55 days, with line NPR1-780 showing the lowest disease severity index. This line also showed good performance when inoculated with the V-1558 strain (ND, race 1), although the differences from the control were not statistically significant. In response to the necrotroph R. necatrix, all the transgenic lines showed a slight delay in disease development, with mean area under the disease progress curve (AUDPC) values 7–15% lower than that of the control.

Highlights

Olive (Olea europaea) is a fruit crop widely cultivated in the countries of the Mediterranean Basin
A total of 1064 globular somatic embryos (SE) from the P1 line were inoculated with the A. tumefaciens AGL1 disarmed strain carrying the pK7WG2.0 binary vector with the AtNPR1 gene (Figure 1A)
The percentages of SE maturation were similar in the control and transgenic lines, approximately 30%, whereas the rates of shoot germination were slightly higher in the transgenic NPR1 lines, 42–53% vs. 23% in the control

Summary

Introduction

Olive (Olea europaea) is a fruit crop widely cultivated in the countries of the Mediterranean Basin. Populations of V. dahliae comprise two types of pathogenic variation, namely, defoliating (D) and non-defoliating (ND) pathotypes (i.e., symptom types) and pathogenic races 1 and 2 (Jiménez-Díaz et al, 2017). Isolates of the D pathotype cause defoliation of cotton, olive, and okra, whereas isolates of the ND pathotype do not defoliate these species (Jiménez-Díaz et al, 2017). Isolates of race 1, which is avirulent on tomato plants with the resistance gene Ve1 (de Jonge et al, 2012), belong to the ND pathotype, whereas the Ve1resistance breaking isolates of race 2 can be found with both D and ND pathotypes (Jiménez-Díaz et al, 2017). Race 2 lacks Ave and is pathogenic to cultivars with the Ve1 gene because a lack of recognition between the factors prevents the induction of defense mechanisms

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