Extended Data Fig. 7: DNMT3A and DNMT3L are essential to SSC plasticity.

a, Pseudotime trajectory of Dnmt3A mutants (mixed Dnmt3A^KO and Prdm1-Dnmt3A^cKO) germ cells from 10 dpp testes. The percentage of each cell type is indicated for every pseudotime segments. The two highest values are colored in purple. b, c, Pseudotime trajectory of germ cells from 10 dpp testes. (Left) Dnmt3L^WT germ cells (FACS-sorted). (Right) Dnmt3L^KO germ cells (FACS-sorted). In (b), cells were ordered from beginning (dark blue) to the end (light blue). In (c), cells were colored following germ cell cluster allocation, see key above. Patterns may differ from Dnmt3A mutant pseudotime (Fig. 4a, b), due to the use of different versions of the 10X Genomics scRNA-seq kit (V2 for Dnmt3L- versus V3 for Dnmt3A-related samples). RNA velocity could not be performed on Dnmt3L^KO, due to insufficient germ cells (1,031 WT and 1,182 Dnmt3L^KO) and the use of anterior version of the scRNA-seq kit. d, Representative FACS plots of live EpCAM-pos, β2M-neg gated testicular cells from 8 dpp Dnmt3A^KO and WT males. EpCAM-pos cells were divided into EpCAM^low and EpCAM^high populations. e, Morphological observation of germ cell clusters derived from EpCAM^low, β2M-neg cells of 8 dpp WT mice after 11 days of culture. f, Table showing the number of in vitro germ cell clusters forming from EpCAM^low and EpCAM^high spermatogonia at 8 dpp and EpCAM^total (= EpCAM ^{low + high}) spermatogonia at 21 dpp. At 8 dpp, the results come from one biological replicate (n = 1 Dnmt3A^KO and n = 1 WT). At 21 dpp, the results come from two biological replicates (n = 2 Dnmt3A^KO and n = 2 WT).