Fig. 5: LRPPRC interacts with SNHG17 and promotes G1/S transition by increasing the stability of c-Myc protein. | Cell Death & Disease

Fig. 5: LRPPRC interacts with SNHG17 and promotes G1/S transition by increasing the stability of c-Myc protein.

From: LncRNA SNHG17 interacts with LRPPRC to stabilize c-Myc protein and promote G1/S transition and cell proliferation

Fig. 5

A Identification of SNHG17-associated proteins by RNA pulldown and mass spectrometry assays. Biotin-labeled SNHG17 or its antisense RNA (SNHG17-AS, negative control) was incubated with lysates from HepG2 cells to pull down the SNHG17-associated proteins, followed by silver staining. A specific fragment (indicated by an arrow), pulled down by SNHG17 but not SNHG17-AS (left panel), was subjected to mass spectrometry. The candidate proteins are listed (right panel). B LRPPRC but not DHX9 or UPF1 was enriched in the protein complexes pulled down by SNHG17. Cellular proteins were pulled down with biotin-labeled SNHG17 or SNHG17-AS and then subjected to Western blotting. GAPDH, negative control. C RNA immunoprecipitation assays revealed an interaction between SNHG17 and LRPPRC in vivo. The RNAs that were precipitated by anti-LRPPRC antibody or isotype-matched control IgG were analyzed by qPCR, using primers for SNHG17 or U6. The enrichment was normalized to the IgG control. U6 was used as a negative control. D Silencing LRPPRC decreased the protein level of c-Myc. E The siLRPPRC-induced c-Myc reduction was abrogated by proteasome inhibitor. NC- or siLRPPRC-transfectants without (−) or with (+) 10 μg/mL MG132 treatment for 3 h were subjected to Western blotting. SE, short exposure; LE, long exposure. F Silencing LRPPRC increased the level of ubiquitinated c-Myc. NC- or siLRPPRC-transfectants without (−) or with (+) 10 μg/mL MG132 treatment for 3 h were subjected to immunoprecipitation (IP) using anti-c-Myc antibody and then Western blotting using anti-Ubiquitin antibody (Ub). G Silencing LRPPRC decreased the stability of c-Myc protein. NC- or siLRPPRC-transfected cells were treated with 25 μg/mL cycloheximide for the indicated time periods before Western blotting. H Ectopic expression of c-Myc antagonized the roles of siLRPPRC in decreasing DNA-replicating cells (left panel) and cell growth (right panel). Cells stably expressing c-Myc and the control cells (Ctrl) were transfected with NC or siLRPPRC and then subjected to EdU incorporation assays (left panel) and cell counting (right panel). For C and H, error bars represent mean ± SEM from three independent experiments. P values were assessed by unpaired Student’s t test. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ns, not significant.

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