Extended Data Figure 6: KBTBD8 specifies neural crest fate through TCOF1 and NOLC1.
From: Cell-fate determination by ubiquitin-dependent regulation of translation
a, mRNA levels of KBTBD8, NOLC1 and TCOF1 were determined in hESCs or differentiating cells transduced with lentiviruses expressing the indicated shRNAs by qRT–PCR (mean of 3 technical replicates ± s.e.m.). b, hESCs stably depleted of KBTBD8 and reconstituted with either wild-type KBTBD8, KBTBD8(W579A), or KBTBD8(Y74A) were subjected to neural conversion (9 days) and analysed for the expression of marker proteins by qRT–PCR (mean of 3 technical replicates ± s.e.m.). c, hESCs stably depleted of KBTBD8, TCOF1, or NOLC1 were subjected to neural conversion (9 days) and analysed for marker expression by qRT–PCR (mean of 3 technical replicates ± s.e.m.). d, Depletion of TCOF1 or NOCL1 from hESCs results in loss of neural crest cells, as determined by triple staining immunofluorescence against the neural crest markers HNK1, TFAP2 and p75 (n > 200 cells, mean of 3 biological replicates ± s.d.). Scale bar, 10 μm. e, hESCs were transduced with lentiviruses expressing control or BRD2 shRNAs, subjected to puromycin selection for 7 days, and analysed by western blot analysis. f, Depletion efficiency for shRNAs against various KBTBD8 binding partners, as determined by qRT–PCR (mean of 3 technical replicates ± s.e.m.).
