Extended Data Fig. 4: O-GlcNAcylation of YTHDF3 repressed target mRNA translation efficiency.
From: O-GlcNAcylation determines the translational regulation and phase separation of YTHDF proteins
(a, b) Expression analysis of N-WT-YTHDF1/3-λ and N-YTHDF1/3-Mut-λ in of Fig. 4b,c and Extended Data Fig. 4f-g. (c) Translation efficiency analysis of N-WT-YTHDF1-λ containing no, either or both mutation on O-GlcNAcylation sites. Error bars, mean ± s.d., n = 3 biologically independent repeats. Significance was calculated using two-sided t-test. (d) Translation efficiency analysis of N-WT-YTHDF1-λ and λ treated with different concentration of OGT inhibitor OSMI-1 in HEK293T. Error bars, mean ± s.d., n = 3 biologically independent repeats. Significance was calculated using two-sided t-test. (e) Translation efficiency analysis of N-WT-YTHDF1-λ and λ treated with different concentration of OGA inhibitor Thiamet G in HeLa. Error bars, mean ± s.d., n = 3 biologically independent repeats. Significance was calculated using two-sided t-test. (f) Translation efficiency analysis of N-WT-YTHDF3-λ, N-YTHDF3-Mut-λ and λ in HEK293T. Error bars, mean ± s.d., n = 3 biologically independent repeats. Significance was calculated using two-sided t-test. (g) Translation efficiency analysis of N-WT-YTHDF3-λ, N-YTHDF3-Mut-λ and λ in HeLa. Error bars, mean ± s.d., n = 3 biologically independent repeats. Significance was calculated using two-sided t-test. (h) RNA abundance analysis of N-WT-YTHDF3-λ, N-YTHDF3-Mut-λ and λ in HEK293T by qPCR. Error bars, mean ± s.d., n = 3 biologically independent repeats. Significance was calculated using two-sided t-test. (i) Translation efficiency analysis of N-WT-YTHDF3-λ containing no, either or both mutation on O-GlcNAcylation sites. Error bars, mean ± s.d., n = 3 biologically independent repeats. Significance was calculated using two-sided t-test. (j) Comparison of fold change of label-free quantification of immunoprecipitated translation related proteins interacting with WT-YTHDF1/3 and YTHDF1/3-Mut in HEK293T and HeLa by LC–MS/MS. The red lines denoted the mean fold change of the proteins. Mean fold change in HEK293T is lower than in HeLa, which might be due to the low O-GlcNAcylation level of YTHDF1/3 in HeLa.
