# Functional and structural analyses of the Vitis rotundifolia genomic region containing Rpv2, a resistance gene to downy mildew

Laurie Marsan, Marie-Christine Piron, Camille Rustenholz, Pere Mestre

► To cite this version:

Laurie Marsan, Marie-Christine Piron, Camille Rustenholz, Pere Mestre. Functional and structural analyses of the Vitis rotundifolia genomic region containing Rpv2, a resistance gene to downy mildew. Plant Biology Europe 2023, Jul 2023, Marseille, France. hal-05282448

HAL Id: hal-05282448

https://hal.inrae.fr/hal-05282448v1

Submitted on 25 Sep 2025

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## Vitis rotundifolia QTL resistance

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Vitis vinifera, the cultivated European grapevine, is highly susceptible to *Plasmopara viticola*, the pathogen responsible of downy mildew. *V. rotundifolia*, an American wild grapevine, is totally resistant to many pathogens including *P. viticola*.

2 major resistance QTLs were identified in *V. rotundifolia* :

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*Rpv1* on chromosome 12, a partial resistance factor due to a TIR-NBS-LRR gene

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*Rpv2* a **total resistance** factor, still uncharacterized, located on chromosome 18

## 18 *Rpv2* locus boundaries

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Genotyping and phenotyping of *V. vinifera* X *V. rotundifolia* recombinants of the introgression of *Rpv2* allowed us to narrow down the boundaries of the locus to **780kb on haplotype 1 of cv Trayshed.**

## What is the molecular basis of Rpv2-mediated resistance?

### Selection of candidate genes

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Gene annotation, determination of their putative function, verification of protein integrity and gene expression allowed us to select some candidates for *Rpv2* resistance including TNL g231970 which has its expression induced in presence of *P. viticola*.

### Functional validation of candidate genes : methodological development

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**Preliminary results**

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**Method limitations:**

* Locations of agroinfiltration spots hard to track along the experiment

* Imprecise inoculations on agroinfiltration spots

* Different contact time with zoospores

**To be tested:**

* Work on leaf disks to limit the surface area to be inoculated and infiltrated?

* Inoculation before agroinfiltration

* Quantitative approach on downy mildew hyphae

## Identification of AvrRpv2 via transient expression of effector candidates

**HR test results**

Since two candidate genes for *Rpv2* resistance are TIR-NBS-LRR, it will be interesting for us to investigate on the avirulence gene recognised by this resistance factor.

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*P. viticola* effectors were selected via in silico analysis on the presence of RLXR motif and/or WY domain. 377 putative effectors were identified.

The candidate avirulence gene should induce a hypersensitive response (HR) in plant carrying *Rpv2*. Each effector is thus agroinfiltrated into *Rpv2* plants, then cell-death induction is assessed at 6 dpi. Plants lacking *Rpv2* (Syrah) are used as a negative control.

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## Conclusion and perspectives

Using *in silico* methods, we managed to identify one interesting candidate for the *Rpv2* resistance. To validate this gene in grapevine we choose a transient expression approach, owing to the lack of an efficient and rapid protocol for stable transformation. However the transient expression protocol in grapevine needs to be improved to overcome the limitations we identified. At the same time, we're trying to identify the avirulence protein recognised by the TIR-NBS-LRR candidate. Until now we managed to test 50 effectors up to then on *Rpv2* plants. In order to select new effectors for functional analyses, we will perform modelling to predict protein-protein interactions between all putative effectors and the TNL candidate.