Figure 8. KYP methylates Geminivirus chromatin as a host defense.
(A) Western blot analysis of kyp complementation lines expressing PKYP- or 35S-Flag-4Myc-KYP using anti-myc antibody. *, a cross-reaction band serves as a loading control. (B) Representative CaLCuV symptoms with different severities. (C) Time course of CaLCuV symptom development in kyp mutant and the complementation lines. The mean values were calculated with SD from at least three experiments (>30 plants/line). (D) CaLCuV symptom severity in Col-0, kyp mutant, and the complementation lines. The mean values were calculated with SD from at least three experiments (>30 plants/line). (E) ChIP-PCR of H3K9me2 marks in heterochromatic loci in kyp mutant and complementation lines inoculated with mock or CaLCuV. Note: CaLCuV infection largely removed H3K9me2 marks from heterochromatic loci. (F) Schematic linearized representation of the regions of viral genome A selected for ChIP assays. (G) ChIP-qPCR assays showed KYP-dependent enrichment of H3K9me2 in several tested loci in the viral chromatin. The relative value of histone methylation in each sample was normalized to that of wild-type control where the signal was arbitrarily assigned a value of 1 with standard deviation (SD) from at least three biological repeats. Note: the region defined by # Primer 7 serves as a negative control. (H) Western blot analysis to detect Flag-Myc4-KYP in the ChIP (IP) samples using anti-myc antibody. (I) ChIP-PCR assays showed that KYP binds to the viral minichromosome. The ChIP assays were done with a monoclonal anti-Flag antibody.
Figure 8—figure supplement 1. Virus chromatin contains H3K9me2 marks.
