Most of the plant shoot originates from a small group of stem cells, which in Arabidopsis are specified by WUSCHEL (WUS). It is unknown whether these cells have an intrinsic potential to generate shoot tissues, or whether differentiation is guided by signals from more mature tissues. Here we show that WUS expression in the root induced shoot stem cell identity and leaf development (without additional cues), floral development (together with LEAFY), or embryogenesis (in response to increased auxin). Thus, WUS establishes stem cells with intrinsic shoot identity and responsive to developmental inputs that normally do not change root identity.

Summary: Amino acid substitutions in the eukaryotic translation initiation factor 4E (eIF4E) result in recessive resistance to potyviruses in a range of plant species, including Capsicum spp. Correspondingly, amino acid changes in the central part of the viral genome‐linked protein (VPg) are responsible for the potyvirus’s ability to overcome eIF4E‐mediated resistance. A key observation was that physical interaction between eIF4E and the VPg is required for viral infection, and eIF4E mutations that cause resistance prevent VPg binding and inhibit the viral cycle. In this study, polymorphism analysis of the pvr2‐eIF4E coding sequence in a worldwide sample of 25 C. annuum accessions identified 10 allelic variants with exclusively non‐synonymous variations clustered in two surface loops of eIF4E. Resistance and genetic complementation assays demonstrated that pvr2 variants, each with signature amino acid changes, corresponded to potyvirus resistance alleles. Systematic analysis of the interactions between eIF4E proteins encoded by the 10 pvr2 alleles and VPgs of virulent and avirulent potato virus Y (PVY) and tobacco etch virus (TEV) strains demonstrated that resistance phenotypes arose from disruption of the interaction between eIF4E and VPg, and that viral adaptation to eIF4E‐mediated resistance resulted from restored interaction with the resistance protein. Complementation of an eIF4E knockout yeast strain by C. annuum eIF4E proteins further shows that amino acid changes did not impede essential eIF4E functions. Altogether, these results argue in favour of a co‐evolutionary ‘arms race’ between Capsicum eIF4E and potyviral VPg.

Summary: In many crop species, natural variation in eIF4E proteins confers resistance to potyviruses. Gene editing offers new opportunities to transfer genetic resistance to crops that seem to lack natural eIF4E alleles. However, because eIF4E are physiologically important proteins, any introduced modification for virus resistance must not bring adverse phenotype effects. In this study, we assessed the role of amino acid substitutions encoded by a Pisum sativum eIF4E virus‐resistance allele (W69L, T80D S81D, S84A, G114R and N176K) by introducing them independently into the Arabidopsis thaliana eIF4E1 gene, a susceptibility factor to the Clover yellow vein virus (ClYVV). Results show that most mutations were sufficient to prevent ClYVV accumulation in plants without affecting plant growth. In addition, two of these engineered resistance alleles can be combined with a loss‐of‐function eIFiso4E to expand the resistance spectrum to other potyviruses. Finally, we use CRISPR‐nCas9‐cytidine deaminase technology to convert the Arabidopsis eIF4E1 susceptibility allele into a resistance allele by introducing the N176K mutation with a single‐point mutation through C‐to‐G base editing to generate resistant plants. This study shows how combining knowledge on pathogen susceptibility factors with precise genome‐editing technologies offers a feasible solution for engineering transgene‐free genetic resistance in plants, even across species barriers.

Capsicumresistance toPepper veinal mottle virus(PVMV) results from complementation between thepvr2andpvr6resistance genes: recessive alleles at these two loci are necessary for resistance, whereas any dominant allele confers susceptibility. In line with previous results showing thatpvr2resistance alleles encode mutated versions of the eukaryotic translation initiation factor 4E (eIF4E), the involvement of other members of theeIF4Emultigenic family in PVMV resistance was investigated. It was demonstrated thatpvr6corresponds to aneIF(iso)4Egene, predicted to encode the second cap-binding isoform identified in plants. Comparative genetic mapping in pepper and tomato indicated thateIF(iso)4Emaps in the same genomic region aspvr6. Sequence analysis revealed an 82 nt deletion ineIF(iso)4EcDNAs from genotypes with thepvr6resistance allele, leading to a truncated protein. This deletion was shown to co-segregate withpvr6in doubled haploid and F₂progeny. Transient expression in a PVMV-resistant genotype ofeIF(iso)4Ederived from a genotype with thepvr6⁺susceptibility allele resulted in loss of resistance to subsequent PVMV inoculation, confirming thatpvr6encodes the translation factor eIF(iso)4E. Similarly, transient expression ofeIF4Efrom a genotype with thepvr2⁺-eIF4Esusceptibility allele also resulted in loss of resistance, demonstrating that wild-type eIF4E and eIF(iso)4E are susceptibility factors for PVMV and that resistance results from the combined effect of mutations in the two cap-binding isoforms. Thus, whilst most potyviruses specifically require one eIF4E isoform to perform their replication cycle, PVMV uses either eIF4E or eIF(iso)4E for infection of pepper.