Genome Editing of eIF4E1 in Tomato Confers Resistance to Pepper Mottle Virus

Abstract

Many of the recessive virus-resistance genes in plants encode eukaryotic translation initiation factors (eIFs), including eIF4E, eIF4G, and related proteins. Notably, <i>eIF4E</i> and its isoform <i>eIF(iso)4E</i> are pivotal for viral infection and act as recessive resistance genes against various potyviruses in a wide range of plants. In this study, we used Clustered Regularly Interspaced Palindromic Repeats/CRISPR-associated protein 9 (CRISPR/Cas9)-mediated targeted mutagenesis to test whether novel sequence-specific mutations at <i>eIF4E1</i> in <i>Solanum lycopersicum</i> (tomato) cv. Micro-Tom could confer enhanced resistance to potyviruses. This approach produced heritable homozygous mutations in the transgene-free E₁ generation. Sequence analysis of <i>eIF4E1</i> from E₀ transgenic plants expressing <i>Cas9</i> and <i>eIF4E-sgRNA</i> transcripts identified chimeric deletions ranging from 11 to 43 bp. Genotype analysis of the <i>eIF4E1</i>-edited lines in E₀, E₁, and E₂ transgenic tomato plants showed that the mutations were transmitted to subsequent generations. When homozygous mutant lines were tested for resistance to potyviruses, they exhibited no resistance to tobacco etch virus (TEV). Notably, however, several mutant lines showed no accumulation of viral particles upon infection with pepper mottle virus (PepMoV). These results indicate that site-specific mutation of tomato <i>eIF4E1</i> successfully conferred enhanced resistance to PepMoV. Thus, this study demonstrates the feasibility of the use of CRISPR/Cas9 approach to accelerate breeding for trait improvement in tomato plants.