Participation of tomato ERF30 in plant immunity against Pseudomonas syringae pv. tomato (Pst)

SIRVENT, EMILIA; POMBO, MARINA A.; ROSLI, HERNÁN GUILLERMO

Congreso; XXXIV Reunión de la Sociedad Argentina de Fisiología Vegetal; 2023

Plants are exposed to a wide variety of pathogens, and to defend themselves they have evolved a two-layered immune system. The first one, called PTI (pattern-triggered immunity), is activated upon the recognition of MAMPs (microbe-associated molecular patterns). Pathogens like Pst can deliver effector proteins into the cytoplasm to promote susceptibility. However, some plants have evolved a second layer of immunity called ETI (effector-triggered immunity) which requires resistance proteins (R) that can recognise these effectors and activate a localized response that culminates in a hypersensitive response (HR). Tomato Rio Grande Pto-R plants detect two Pst effectors (AvrPto and AvrPtoB) with a two-protein complex Pto/Prf.In this investigation we propose to identify and characterize new transcription factors (TF) implicated in tomato defense against Pst. We previously showed silencing ERF30 using virus induced gene silencing (VIGS) in Nicotiana benthamiana (Nb) lead to a compromised Pto/AvrPto-associated programmed cell death (PCD). In addition silenced Nb 35s::Pto plants challenged with P. s. pv. tabaci carrying AvrPto showed compromised Pto/Prf mediated resistance. We have now measured electrolyte leakage in plants infiltrated with this same strain and observed lower conductivity in ERF30-silencend plats compared to controls.CRISPR-Cas9 technique was used to develop ERF30 knockout tomato lines. The targeted genomic region of T0 generation was sequenced confirming a homozygous single base deletion in two events and from these plants we generated T1 seeds. Both, T0 and T1, were indistinguishable from wild type plants. PCR reactions amplifying two different regions of the cassette used in CRISPR technique, Cas9 and kanamycin resistance genes, confirmed two plants of each line as transgene free. From these plants we plan to obtain transgene-free homozygous plants in T2 generation to assess the functional status of the defense in the mutant plants.