Supplementary Figure 6: The E59K, V60M, P76L and G82R of DND1 disrupt the protein stability and protein-protein interaction. | Nature Cell Biology

Supplementary Figure 6: The E59K, V60M, P76L and G82R of DND1 disrupt the protein stability and protein-protein interaction.

From: CRISPR–Cas9-mediated base-editing screening in mice identifies DND1 amino acids that are critical for primordial germ cell development

Supplementary Figure 6

a, Sequence alignment of RRM domains from various proteins. G82 is well conserved in RRM domains. b, In vitro purification of wild-type and mutant RRM domains (1-217) of DND1 in E. coli. c, GST pull-down assay for the interactions of GST-NANOS2 or GST-CNOT1 (1-551) with DND1 and DND1 mutant proteins. d, Transcription analysis of Oct4, Sox2, Nanog, Dnd1, Nanos2 and Cnot1 genes in mouse ESCs (E14). The expression values were normalized to that of Gapdh. Data are shown as the average mean +/− s.d. (n = 3 three independent experiments). The source data can be found in Supplementary Table 13. e, Analysis of expression levels of DND1 and DND1 mutant proteins in ESCs by transfection of HA-tagged Dnd1 plasmids. f, g, Co-immunoprecipitation (Co-IP) analysis in mouse ESCs shows that the mutation of DND1 disrupt the interaction with NANOS2 but did not affect the binding with CNOT1. Data in b, c and e-g represent 3 independent experiments and their unprocessed images of gels are shown in Supplementary Fig. 7.

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