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. 2021 Jun 25;12:3965. doi: 10.1038/s41467-021-24046-3

Fig. 6. UPF1 accumulates with NMD factors in SMG5-SMG7 depleted cells.

Fig. 6

a Overview of the TurboID-mediated proximity labeling of UPF1 binding partners. Transient UPF1 interactors are marked with biotin via TurboID catalysis. Biotinylated proteins are subsequently enriched with streptavidin beads. b Western blot and TCE (2,2,2-trichloroethanol)-mediated detection of input or streptavidin-enriched protein samples after proximity labeling in the indicated conditions (n = 3 biologically independent samples). c Heatmap of mass spectrometry-based analysis of streptavidin-enriched biotinylated NMD and selected other proteins in the respective comparison of conditions (n = 3 biologically independent samples). Colored points indicate the log2 fold change (log2FC) and point size corresponds to the adjusted p-value (adj. p-value; from two-sided Welch‘s t-test). Multiple testing between FLAG-TurboID-UPF1 in WT, SMG7 KO, and SMG7 KO + SMG5 KD was performed by ANOVA. d, e Volcano plots of mass spectrometry-based analysis of streptavidin-enriched biotinylated proteins in the respective comparison of conditions (n = 3 biologically independent samples). d FLAG-TurboID-UPF1 against FLAG-TurboID control in SMG7 KO cells, (e) FLAG-TurboID-UPF1 against FLAG-TurboID control in SMG7 KO + SMG5 KD cells. The yellow color labeling indicates targets that are significant in the respective comparisons after two-sided Welch’s t-testing (log2 fold change (log2FC) >1 or |log2FC| >1; and adj. p-value <0.05). Points labeled in blue indicate other proteins of interest; points labeled in red indicate NMD factors. Highlighted proteins that were not significant in the respective comparisons are labeled with gray text.