Fig. 3: Homodimeric OS9-SEL1L complex forms in vivo.
From: Structural basis and pathological implications of the dimeric OS9-SEL1L-HRD1 ERAD Core Complex
a, b OS9-SEL1L-HRD1 model shown as a transparent surface. Zoomed-in view of the dashed box shown in (b). Key residues (SEL1L-D439, M440, and OS9-D135) at the interface are shown, with N442 as a negative control. c Distance between OS9-D135 and SEL1L-D439, M440 and N442. d, e (Non-)Reducing and Western blot analysis of the native disulfide crosslinking assays in HEK293F cells transfected with HRD1-FLAG, SEL1L WT and mutants (D439C, M440C, N442C), and OS9-D135C mutations, with quantitation of the percentage of disulfide-linked OS9-SEL1L shown in (e) (three independent repeats. p value in OS9-SEL1L/OS9: D439C p < 0.0001, M440C p < 0.0001, N442C p = 0.8804; p value in OS9-SEL1L/SEL1L: D439C p < 0.0001, M440C p = 0.0007, N442C p = 0.958). Lysates were treated with 15 mM N-ethylmaleimide (NEM) to preserve native disulfide-linked species during sample preparation. f (Non-)Reducing SDS-PAGE analysis (Coomassie blue staining) of HRD1-FLAG IP in HEK293F cells expressing indicated OS9 and SEL1L mutations (two independent repeats). Three panels in (f) were from the same gel, with the irrelevant lanes in the middle cut off. Values, mean ± SEM. n.s., not significant, ***p < 0.001 and ****p < 0.0001 by One-way ANOVA analysis with comparison to WT. Source data of Fig. 3d–f are provided as a Source Data file.
