Fig. 1

CRISPRa induces upregulation of cell surface protein levels. a Schematics of the dCas9 fusion protein and gRNA-dependent recruitment of the VP64 and MS2-p65-HSF1 transcriptional activators to gene promoter regions (left), and transposon-based plasmids for CRISPR-based transcriptional activation and non-activating control (right). b Exemplar cytometry plots showing heterogenous upregulation of SEMA7A and ICAM1 at the surface of HEK293 cells 48 h after transduction at low MOI with a lentivirus-delivered pool of eight gRNAs and transfection with either activating (dCas9-VP64) or non-activating control (dCas9) plasmids. gRNA positive cells are represented by red dots, gRNA negative cells in gray. c Quantification of cell surface receptor protein upregulation on cells transduced with lentiviruses encoding gRNAs corresponding to the appropriate gene using five different dCas9 activator constructs relative to a non-activating dCas9 control. d qRT-PCR analysis of relative mRNA abundance of indicated target genes in cells 48 h post co-transfection with dCas9-VP64 and either targeting gRNA (+) or no gRNA control. Transcript abundance was normalized to CYPA expression; bars represent mean ± s.e.m.; n = 6. P values calculated using a Student’s t test, ns P > 0.05; **P ≤ 0.01; ***P ≤ 0.001. e Percentage of cells expressing the indicated cell surface receptors as determined by mAb staining after transduction of the cloned activator cell line, HEK293-V2M, with appropriate pooled gRNAs. Data points in c and e represent mean ± s.e.m.; n = 3