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. 2010 Oct 20;85(1):243–253. doi: 10.1128/JVI.01749-10

FIG. 9.

FIG. 9.

Role of RNase MRP in TBSV RNA degradation and RNA recombination in plants. (A) (Left) Phenotype of 7-2 RNA/NME1 knockdown N. benthamiana plants 14 days after agroinfiltration with TRV silencing vectors. (Right) Effect of knockdown of 7-2 RNA/NME1 on symptoms caused by CNV infection in N. benthamiana 8 and 10 days after the second agroinfiltration. VIGS was performed via agroinfiltration of vector (TRV) carrying the 7-2 RNA/NME1 sequence or the TRV empty vector (as a control). Coagroinfiltration to express TBSV DI-72 repRNA together with CNV gRNA was done 9 days after silencing of 7-2 RNA/NME1 expression by agroinfiltration. (B) Northern blot analysis of CNV gRNA accumulation in the agroinfiltrated leaves of 7-2 RNA/NME1 knockdown or control N. benthamiana plants 4 days after the second agroinfiltration. rRNA (not shown) was used as a loading control. The plant leaf samples were obtained from 5 separate experiments (total of 5 × 48 samples). (C) Decreased accumulation of TBSV degRNA-RI in 7-2 RNA/NME1 knockdown N. benthamiana plants compared to that in the control plants, which were agroinfiltrated with the TRV empty vector, 4 days after the second agroinfiltration. The accumulation level of degRNA-RI was normalized to the level of DI-72 repRNA. Note that the RI probe was specific for the TBSV repRNA and that the helper CNV gRNA or its degradation products were not detected. (D) Northern blot analysis of TBSV repRNA and newly formed recRNAs in agroinfiltrated leaves of 7-2 RNA/NME1 XRN4 double-knockdown or XRN4 single-knockdown control N. benthamiana plants. The numbers at the bottom of the panel show percentages of recRNA2 and recRNA3 accumulation normalized to the level of DI-72 repRNA (100% in each sample). rRNA (not shown) was used as a loading control. Note that the RIV probe was specific for the TBSV repRNA and that the helper CNV gRNA or its degradation products were not detected. (E) Decreased accumulation of TBSV degRNA-RI in 7-2 RNA/NME1 XRN4 double-knockdown plants compared to that in the control XRN4 single-knockdown N. benthamiana plants. See further details above as described for panel C.