Supplementary Figure 4: RSL1D1 knockdown-induced senescence requires the RB pathway.

(a) SA-β-gal of IMR90 cells expressing the indicated shRNA expression vectors in combination with the expression of E6 alone, E7 alone, E6/E7 together or an empty control vector (Vect) at day 20 post-infection. Data were quantified from >5-independent cell counts up to a total of at least 150 cells and are presented as the mean percentage of positive cells, (1 out of 2 independent experiments with similar results). shNTC: non-targeting shRNA; shR-A or -B: shRNAs against RSL1D1. (b-e) QPCR for the indicated genes performed with total RNA extracted from cells as in (a) at day 20 post-infection. Data were normalized over TBP and HMBS and presented as mean relative to vector/shNTC infected cells, (n = 1). (f) Growth curves of IMR90 cells expressing the indicated shRNA expression vectors in combination with the expression of E7(Δ6-10), E7(Δ21-24), E7(Δ79-83) or an empty control vector (Vect). Data are presented as mean normalized to day 0 of each condition. (1 out of 2 independent experiments with similar results). The percent of SA-β-gal positive cells for each condition at day 20 post-infection is indicated under each growth curve. Data were quantified from >5-independent cell counts up to a total of at least 150 cells and are presented as the mean percentage of positive cells, (1 out of 2 independent experiments with similar results). shNTC: non-targeting shRNA; shR-A or -B: shRNAs against RSL1D1. (g-j) QPCR for the indicated genes performed on reverse transcribed total RNA extracted from cells as in (f) at day 20 post-infection. Data were normalized over TBP and HMBS and presented as mean relative to vector/shNTC infected cells, (n = 1).