Fig. 7: Rac1b function is required for the maintenance of a large subset of BCSCs expressing Rac1b.

A Representative dot plots for flow cytometry analysis of primary mammary tumor cells from Rac1b^+/−;MMTV-NIC and Rac1b^RFP/+;MMTV-NIC mice are shown for the single, alive, lineage-negative cell population. The observed range for RFP⁺ cell frequency in tumors of different animals is shown within the RFP⁺ gate (n = 6). B Mammosphere forming efficiency (%MFE) of RFP⁺ versus RFP^- cells sorted from Rac1b^RFP/+;MMTV-NIC tumors as shown in (A). Values represent the mean ± SEM of 6 animals. (***p < 0.005; paired t-test). C Representative confocal microscopy images of a primary mammosphere formed by Lin^-RFP⁺ tumor cells sorted from Rac1b^RFP/+;MMTV-NIC mammary tumors (n = 3 mice). The mammosphere was coimmunostained for CK18 and CK14 expression. The leftmost image shows the deconvoluted image for CK14 (green) and CK18(red). Individual images of confocal planes selected to represent different Z-stack positions are shown together with DAPI-staining (shown in blue) serving as a nuclear stain. Scale bars represent 20 um. D Pie chart representation of the distribution of CK-18 and/or CK-14 expressing Lin^-RFP⁺ tumor cells sorted from Rac1b^RFP/+;MMTV-NIC breast tumors. Values represent the mean data for 3 independent tumors analyzed. E Representative histogram of flow cytometry analysis for CD24 expression in Lin^-RFP⁺ cells sorted from Rac1b^RFP/+;MMTV-NIC tumors. The observed range for the CD24⁺ cell frequency in tumors of different animals is shown on the histogram (n = 3). F Representative dot plots of flow cytometry analysis for primary mammary tumor cells of Rac1b^RFP/+;MMTV-NIC and Rac1b^RFP/−;MMTV-NIC mice are shown for the single, alive, lineage-negative cell population. The observed range of RFP⁺ cell frequency in tumors of different animals with same genotype is shown within the RFP⁺ gate (n = 9). G Mammosphere forming efficiency (%MFE) of RFP⁺ and RFP^- cells sorted from Rac1b^RFP/+;MMTV-NIC and Rac1b^RFP/−;MMTV-NIC tumors as shown in (F). Values represent the mean ± SEM of 3 animals. (**p < 0.01, ns not significant; paired t-test).