Fig. 4.

Progenitors with higher SOX2 produce TRC-containing organoids, whereas organoids from progenitors expressing low levels of SOX2 comprise mostly non-taste epithelium. (A) Expression of type I, II and III TRC markers (Kcnj1, Gnat3 and Pkd2l1, respectively) is higher in SOX2^High or SOX2^HiMed organoids but all SOX2-derived organoids express these genes at lower levels than LGR5-derived organoids. Data are mean±s.e.m. *P<0.05, **P<0.01, ***P<0.001, ****P<0.0001 (one-way ANOVA). (B,C) SOX2^High and SOX2^HiMed organoids contain type I (NTPDase2, green), type II (gustducin, green) and type III (CAR4, green) TRCs (yellow arrows), and SOX2^High organoids contain more type III TRCs than other organoid types. The large majority of SOX2^MedLow and SOX2^Low organoids do not contain TRCs. n vales (in brackets) indicate total organoids quantified per condition from three independently derived organoid experiments. (D) SOX2^High and SOX2^HiMed organoids express high levels of a general TRC marker, Kcnq1, and moderate levels of Krt13 (non-taste epithelium). SOX2^MedLow and SOX2^Low organoids express limited Kcnq1 and highly elevated Krt13. Data are mean±s.e.m. ****P<0.0001 (one-way ANOVA). (E-H‴) KRT14⁺ (green) cells make up the external epithelium of all SOX2 organoids (E′,F′,G′,H′). SOX2^High and SOX2^HiMed organoids contain KRT8⁺ TRCs (magenta, yellow asterisks in E″,F″) interspersed among KRT13⁺ non-taste cells (cyan, yellow asterisks in E‴,F‴), while SOX2^MedLow and SOX2^Low organoids are predominantly composed of KRT13⁺ cells (cyan in G,G‴,H,H‴). SOX2^MedLow organoids have sparse KRT8⁺ cells that co-express KRT14 (magenta, red asterisk in G′-G‴). In A,D, n=9 (three samples each from three independently derived organoid experiments). Images in B are maximum projections of confocal z-stacks of whole organoids with DAPI nuclear counterstain (blue). Images in E-H‴ are optical sections of immunostained organoids; areas outlined in the left column are shown in more detail in the three right columns. DAPI (white) nuclear counterstain. Scale bars: 100 µm.