Figure 3: Cell-cell correlations within Dgcr8^−/− and Dgcr8^−/− transfected with either miR294 or let-7c.

(a) Density plots of distances (|1–|PCC|) between pairs of cells within each condition. Top plot is for all genes, whereas bottom plots is for CellNet pluripotency regulators as defined in ref. 14. Top plot inset shows PCC distribution between pairs of cells within each condition. It is worth noting that miR-294 increases homogeneity when considering all genes or pluripotency genes. (b) Heatmaps showing all within condition cell–cell PCC using all genes. Cells are ordered by hierarchical clustering and subpopulation of cells identified with Dynamic tree cut method (see Methods). Above each heatmaps the percentage of cells in each of the cell cycle phases is reported for each identified sub-population of cells as defined by Cyclone. (c) Inter-cluster distance distribution between clusters (that is, subpopulations) of cells shown in b. Inter-cluster distance for a cell x was defined as its average distance from all the other cells, expect the ones in the same subpopulation as cell x. Let-7c inter-cluster distance is significantly higher than Dgcr8^−/− inter-cluster distance (two tailed Wilcoxon test P=1.94e−8), meaning that let-7c is producing more distinct and better separated sub-populations of cell compared to the ones identified in Dgcr8^−/− cells. (d) Identification of pathways driving the formation of subpopultions of cells within let-7c or Dgcr8 knockdown conditions. Pathways are sorted according the corrected P-value (that is, FDR) returned by the ANOVA. Only top five significant pathways are shown. (e) Percentage of cells in each of the cell cycle phases for miRNA-transfected and Dgcr8^−/−-deficient cells. Exact Fisher’s test is used to the compare number of cells in each cell cycle stage between miRNA-transfected versus Dgcr8-knockdown cells. (f) Average expression of six pluripotency and ten differentiation markers across the three identified subpopulations of let-7c-transfected cells.