Extended Data Fig. 1: Comparative analysis of different grazoprevir-inducible CRISPRa strategies.
From: Programmable solid-state condensates for spatiotemporal control of mammalian gene expression
For grazoprevir-repressible condensates formation, HEK-293 cells were co-transfected with constitutive expression vectors for NLS-PB1-AG-NS3a(H1) (pYK164), dCas9-VPR-4xANR (pYK237) and NLS-MCP-mCherry-2xANR (pYK231). For grazoprevir-triggered dCas9-VPR reconstitution, HEK-293 cells were co-transfected with constitutive expression vectors for dCas9-NS3a(H1) (pYK464) and GNCR-VPR (pYK465). For grazoprevir-triggered SAM assembly, HEK-293 cells were co-transfected with constitutive expression vectors for GNCR-VPR (pYK465), MCP-NS3a(H1) (pSL546) and dCas9 (pYK466). For grazoprevir-inhibited transactivator self-cleavage, HEK-293 cells were transfected with a constitutive dCas9-StaPLd-VPR expression vector (pYK480). All cells were further co-transfected with a minimal promoter (PhCMVmin)-driven SEAP expression vector (pWS107) and a constitutive expression vector for PhCMVmin-specific gRNA containing MS2-box motifs (pWS137). Total transfection amounts were kept constant (580ng) using pcDNA3.1(+) as a filler plasmid. SEAP levels in culture supernatants were quantified at 36h after addition of 1µM of grazoprevir or DMSO (vehicle control). All data are shown as the mean ± s.d. of n=3 independent experiments. Statistical significance was assessed using two-side t-tests. *, p<0.05; **, p<0.01; ***, p<0.001; ****, p<0.0001; ns, not significant. Exact P values are provided in source data. Filled circles show individual results (n).
