Figure 7
Set1 represses Ty1 retrotransposition post-transcriptionally. (a) Example of the Set1 and Set1YF-AA binding on Ty1-1 mRNA. The Set1 CRAC and ChIP profiles are shown. (b) Global expression of endogenous Ty1 retrotransposons in SET1, set1Δ or set1-YF/AA yeast cells, monitored by quantifying Ty1 mRNA levels by RT-qPCR (normalized to 25 S rRNA values; mean±s.d.; n=3). (c) Ty1 retrotransposition assay from a plasmid expressing a Ty1 element tagged with the his3AI reporter gene [33]. An intron is inserted in the HIS3 gene in an antisense orientation in a spliceable orientation in the Ty1 transcript resulting in a Ty1 complementary DNA (cDNA) bearing a functional HIS3 gene. The cDNA can then be integrated into the host genome. Cells that sustain a Ty1-HIS3 retrotransposition event give rise to His+ colonies [32]. The position of the qPCR amplicons used to amplify all the Ty1 mRNAs in (b) and the Ty1 reporter mRNA specifically expressed from the plasmid in (e) are indicated (Ty1 and Ty1-HIS3 amplicons, respectively). (d) Frequency of Ty1-his3AI retrotransposition in SET1, set1Δ or set1-YF/AA yeast cells (number of His+ prototrophs divided by the total number of cells; mean±s.d.; n=3). *p≤0.05 (Welch’s t-test). (e) Plasmid Ty1-his3AI expression in SET1, set1Δ or set1-YF/AA yeast cells monitored by quantifying Ty1-HIS3 mRNA levels by RT-qPCR (normalized to 25S rRNA values; mean±s.d.; n=3).
