ABC transporters alter plant-microbe-parasite interactions in the rhizosphere

Abstract

19Plants are master regulators of rhizosphere ecology, secreting a complex mixture of compounds into 20 the soil, collectively termed plant root exudate. Root exudate composition is highly dynamic and 21 functional, mediating interactions between plants and a wide range of beneficial / harmful soil 22 organisms. Exudate composition is under selective pressure to diversify in response to pathogen 23 perception, whilst maintaining interactions with beneficial organisms. However, crop domestication 24 has exerted significant and unintended changes to crop root exudate composition, and we know very 25 little about genotype -phenotype linkages that pertain to root exudates and rhizosphere interactions. 26 Better understanding could enable the modulation of root exudate composition for crop improvement 27 by promoting positive, and impeding negative, interactions. Root expressed transporters modulate 28 exudate composition and could be manipulated towards the rational design of beneficial root exudate 29 profiles. Using Virus Induced Gene silencing (VIGS), we demonstrate that knockdown of two root-30 expressed ABC transporter genes in tomato cv. Moneymaker, ABC-G33 and ABC-C6, alters the 31 composition of semi-volatile compounds in collected root exudates. Root exudate chemotaxis assays 32 demonstrate that knockdown of each transporter gene triggers the repulsion of economically relevant 33 Meloidogyne and Globodera spp. plant parasitic nematodes, which are attracted to control treatment 34 root exudates. Knockdown of ABC-C6 inhibits egg hatching of Meloidogyne and Globodera spp., 35 relative to controls. Knockdown of ABC-G33 has no impact on egg hatching of Meloidogyne spp. but 36 has a substantial inhibitory impact on egg hatching of G. pallida. ABC-C6 knockdown has no impact 37 on the attraction of the plant pathogen Agrobacterium tumefaciens, or the plant growth promoting 38 Bacillus subtilis, relative to controls. Silencing ABC-G33 induces a statistically significant reduction in 39 attraction of B. subtilis, with no impact on attraction of A. tumefaciens. ABC-C6 represents a promising 40 target for breeding or biotechnology intervention strategies as gene knockdown (-64.9%) leads to the 41 repulsion of economically important plant parasites and retains attraction of the beneficial 42 rhizobacterium B. subtilis. This study exposes the link between ABC transporters, root exudate 43 composition, and ex planta interactions with agriculturally and economically relevant rhizosphere 44 organisms, paving the way for an entirely new approach to rhizosphere engineering and crop 45 protection. 46 47 560 VIGS plasmids pTRV1 and pTRV2 were kindly provided by Prof. Ian Graham, University of York. A.

ETHYLENE RESPONSE FACTOR (ERF) genes modulate plant root exudate composition and the attraction of plant parasitic nematodes

“…The experimental workflow and approach mirrors that of Cox et al (2019a). 200 bp regions of ERF-E2 and ERF-E3 were synthesised individually, with a shared contiguous 200 bp region of the visual reporter gene, PDS.…”

“…pTRV1 and the pTRV2 variants were transformed into Agrobacterium tumefaciens strain LB4404, individually by electroporation. Tomato seedlings were inoculated by topical application of mixed pTRV1 and pTRV2 cultures (one of pTRV2-ERF-E2:PDS, pTRV2-ERF-E3:PDS, or pTRV2-PDS for each experimental mixture), following the methodology of Cox et al (2019a). All downstream assays were conducted using root exudates collected from the timepoint with the highest level of gene knockdown, equating to week three-post inoculation for ERF-E2, and week fourpost inoculation in the case of ERF-E3 (Figure 1).…”

“…We have observed similar variation between Meloidogyne spp. in response to exudates collected after ABC transport gene knockdown (Cox et al, 2019a). For this line of research to develop robust and durable new sources of PPN resistance, it will be important to establish the behavioural impact that such interventions have on geographically coincident species, as well as different populations of a given species (Cox et al, 2019b).…”

“…The plants were removed, and the ddH2O / root exudate solution was passed through a 0.22 µm filter by syringe and stored at 4 °C until use. All chemosensory assays were conducted in 60 mm Petri dishes with a 3 ml base of solidified 1.5% water agar and a surface layer of 3 ml 0.5% agar slurry, as in Cox et al (2019a). Autoclaved Whatman filter paper No.…”

“…The non-feeding second stage PPN juvenile (J2) hatches in the soil, and must find and invade a suitable host plant by following concentration gradients of water-soluble and volatile components of plant root exudate (Čepulytė et al, 2018;Murungi et al, 2018). We have previously demonstrated that the modulation of ABC transporter genes can alter plant root exudate composition, and parasite hatching and attraction ex planta (Cox et al, 2019a;Warnock et al, 2016). This represents an interesting approach to crop plant resistance, as it manipulates parasite behaviour in the rhizosphere, and limits host invasion events that can lead to secondary infection, and metabolically expensive defence responses.…”