Fig. 5: Proteomic sequencing identifies SUMO2 as a key RAB5IF-dependent protein.
From: Endothelial RAB5IF is required for pathological and developmental retinal angiogenesis
A–F Proteomic sequencing was performed on hRMECs with shRAB5IF (shRAB5IF#3) or shC, with volcano plots shown differentially proteins (A) (n = 3 biological repeats/group); GO functional analysis for the upregulated proteins (B) and downregulated proteins (C). Volcano plots showing differentially expressed proteins involved in ribosomal subunit synthesis (D). Expression of listed ribosomal proteins was also tested (n = 4 biological repeats, 15 μg protein/lane) (E). Scatter graph displaying SUMO2 as the top downregulated protein following RAB5IF silencing in hRMECs (F), with the center line marking the median (n = 2 biological repeats, a replicate was excluded due to non-detection of the SUMO2 protein). G Western blotting analysis of SUMO1 and SUMO2 protein expression following RAB5IF knockdown (shRAB5IF) or RAB5IF knockout (koRAB5IF) in hRMECs (n = 5 biological repeats, 15 μg protein/lane for WB). H Western blotting analysis of SUMO1 and SUMO2 protein expression in mRMECs extracted from RAB5IF-eKD and shC groups in P17 OIR mice (pooled from 4 mice/sample, n = 5 samples/group, 15 μg protein/lane). I qRT-PCR analysis of SUMO1 mRNA expression following RAB5IF knockdown (by shRAB5IF#3) or RAB5IF knockout (by koRAB5IF#1) in hRMECs (n = 5 biological repeats). J Polysome profiling of shRAB5IF or shC. The x-axis represents the sucrose concentration gradient, and the y-axis shows OD260, reflecting mRNA levels (n = 5 biological repeats). K Polysome PCR analysis of SUMO1 and SUMO2 mRNA expression in shRAB5IF or shC hRMECs. The P/non-P ratio indicates overall translation activity (n = 5 biological repeats). Data are presented as means ± S.D; Fisher’s exact test (B, C), One-way ANOVA with Bonferroni’s post hoc test (E) and Two-tailed Student’s t-test (D, G, K). Source data are provided as a Source Data file.
