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. 2019 Dec 11;10(12):947. doi: 10.1038/s41419-019-2148-2

Fig. 3. ODIR1 physically interacts with E3 ligases and histone markers.

Fig. 3

a The hUC-MSCs were differentiated for 14 days and transfected with vector and ODIR1 plasmid. After 24 h transfection, RT-qPCR analysis for ODIR1, OSX, RUNX2, and OPN RNA levels, western blotting analysis for OSX, RUNX2, and OPN proteins levels. b The hUC-MSCs were transfected with NC and ODIR1 siRNAs, RT-qPCR analysis for ODIR1, OSX, RUNX2, and OPN RNA levels, western blotting analysis for OSX, RUNX2, and OPN proteins levels. c Biotin-labeled ODIR1 sense and anti-sense chains were incubated with hUC-MSCs lysates, and enriched products were collected and subjected to SDS-PAGE polyacrylamide gel electrophoresis and silver staining. Differential bands were identified by LC-MS analysis. d ODIR1 associated with FBXO25, BARD1 and CUL3, histone proteins including H2A, H2B, H3, and H4 as shown by RNA pull-down and western blotting. e ODIR1 was enriched by FBXO25, BARD1, and CUL3 in hUC-MSCs lysates. Anti-IgG were used as negative control. The fold enrichment values were normalized to that of Input. f The hUC-MSCs were transfected with siNC and ODIR1 siRNAs, and the RNA levels of ODIR1 was measured by RT-qPCR, the proteins levels of H2AK119ub, H2BK120ub and H3K4me3 were analyzed by western blotting assay. g The hUC-MSCs were transfected with vector and ODIR plasmid, and the RNA levels of ODIR1, OSX, and FBXO25 was measured by RT-qPCR, the proteins levels of H2AK119ub, H2BK120ub, and H3K4me3 were analyzed by western blotting assay.