Supplementary Figure 8: Rbpj-deficient T-IPs at E11.5 show unaffected expression of Notch-related and lineage-affiliated genes.
From: Initial seeding of the embryonic thymus by immune-restricted lympho-myeloid progenitors
Related to main Figure 8. (a-d) Single cell gene expression analysis of E11.5 CD45+Lin–c-Kit+CD25–Flt3+ T-IPs from RbpjFl/Fl Vav-CreTg/+ Rag1-GFPTg/+ (n=46 cells from 6 embryos) and RbpjFl/Fl Vav-Cre+/+ Rag1-GFPTg/+ (n=36 cells from 8 embryos) littermate controls from 2 different litters. Only cells that amplified Flt3, c-Kit and Hprt (corresponding to 98% of total cells analyzed) were included. (a) Rbpj (b) Notch receptors and Notch target genes (c) frequency of single cells co-expressing GM and early lymphoid affiliated genes. (d) T cell related genes. (b-d) For additional gene expression analysis of Vav-Cre+ cells only cells in which Rbpj had been deleted were included. For panels (a-d) numbers indicate the frequencies of positive cells. For panels b-d no differences between Cre– and Cre+ cells were statistically significant (p<0.05). (e,f) Impact of Rbpj-deficiency on thymocyte progenitor development during early stages of embryonic development. Representative flow cytometry analysis of thymus rudiments from E13.5 (e) and E14.5 (f) RbpjFl/FlVav-CreTg/+Rag1-GFPTg/+ (Vav-CreTg/+; n=3 and n=4, respectively) and RbpjFl/FlVav-Cre+/+Rag1-GFPTg/+ (Vav-Cre+/+; n=7 and n=11, respectively) littermate control embryos. Numbers represent frequencies of total CD45+ cells. (g) In vitro T cell differentiation of E11.5 RbpjFl/FlVav-CreTg/+ (Vav-CreTg/+) and RbpjFl/FlVav-Cre+/+Rag1-GFPTg/+ (Vav-Cre+/+) T-IPs on OP9-DL1 stroma. Data are representative of 4 Vav-Cre+/+ and 6 Vav-CreTg/+ embryos analysed following 14 days of culture. Numbers represent frequencies of total CD45+ cells.
