Expression of artificial microRNAs in transgenic Arabidopsis thaliana confers virus resistance

Plant microRNAs ( miRNAs) regulate the abundance of target mRNAs by guiding their cleavage at the sequence complementary region. We have modified an Arabidopsis thaliana miR159 precursor to express artificial miRNAs ( amiRNAs) targeting viral mRNA sequences encoding two gene silencing suppressors, P69 of turnip yellow mosaic virus ( TYMV) and HC- Pro of turnip mosaic virus ( TuMV). Production of these amiRNAs requires A. thaliana DICER- like protein 1. Transgenic A. thaliana plants expressing amiR- P69(159) and amiR- HC- Pro(159) are specifically resistant to TYMV and TuMV, respectively. Expression of amiR-TuCP(159) targeting TuMV coat protein sequences also confers specific TuMV resistance. However, transgenic plants that express both amiR- P69(159) and amiR- HC- Pro(159) from a dimeric pre- amiR- P69(159)/ amiR- HC- Pro(159) transgene are resistant to both viruses. The virus resistance trait is displayed at the cell level and is hereditable. More important, the resistance trait is maintained at 15 degrees C, a temperature that compromises small interfering RNA - mediated gene silencing. The amiRNA- mediated approach should have broad applicability for engineering multiple virus resistance in crop plants.