Geminivirus VIGS of endogenous genes requires SGS2/SDE1 and SGS3 and defines a new branch in the genetic pathway for silencing in plants

Virus-induced gene silencing (VIGS) is a sequence-specific RNA degradation process that can be used to downregulate plant gene expression. Both RNA and DNA viruses have been used for VIGS, but they differ in their mode of replication, gene expression, and cellular location. This study examined silencing mediated by a DNA virus, cabbage leaf curl virus (CaLCuV), in several silencing-deficient Arabidopsis mutants. A DNA VIGS vector derived from CaLCuV, which silenced chlorata42 (ChlI) needed for chlorophyll formation, was used to test endogenous gene silencing responses in suppressor of gene silencing (sgs)1, sgs2, sgs3, and Argonaute (ago)1 mutants defective in sense transgene-mediated post-transcriptional silencing (S-PTGS). SGS2/silencing defective (SDE)1, SGS3, and AGO1 are each dispensable for silencing mediated by transgenes containing inverted repeats (IR-PTGS), and SGS2/SDE1 is dispensable for RNA VIGS. We show that DNA VIGS requires both SGS2/SDE1 and SGS3, regardless of the orientation of 362 nt ChlI transcripts produced from the viral DNA promoter. Viral DNA accumulation is slightly higher, and viral symptoms increase in sgs2 and sgs3, whereas overexpression of SGS2/SDE1 mRNA results in decreased viral symptoms. Mutants affected in SGS1 and AGO1 function are only delayed in the onset of silencing, and have a small effect on chlorophyll accumulation. DNA VIGS is unaffected in defective DNA methylation (ddm)1/somniferous (som)8 and maintenance of methylation (mom)1 mutants, impaired for TGS. These results demonstrate that SGS2/SDE1 and SGS3 are needed for endogenous gene silencing from DNA viruses, and suggest that SGS2/SDE1 may reduce geminivirus symptoms by targeting viral mRNAs.