a, Expression of three short hairpin RNAs in NIH3T3 fibroblasts reduced Cfp1 messenger RNA levels to 15% compared with vector-only transfected control cells. Expression of Cfp1 relative to Gapdh in control cells is set to 1. The inset shows reduction of Cfp1 by western blotting. Error bars indicate s.d. (n = 3) b, c, Gross morphology (b) and growth rate (c) of Cfp1-depleted versus vector-only transformant cells. Cells were plated at low density and monitored at the times shown using a haemocytometer. Original magnification, ×200. d, ChIP qPCR using Cfp1, H3K4me3 and H3K27me3 antibodies at selected loci in vector-only control and Cfp1-depleted NIH3T3 cells. The results were replicated with an independent clone expressing the same shRNA combination (data not shown) and with each of two individual shRNA constructs (see Supplementary Fig. 3).