Fig. 4

LncRNA DHRS4-AS1 binds DHX9 and accelerates its degradation. (A) FISH was used to detect DHX9 localization in GC, AGS, and HGC-27 cells. (B) LncRNA DHRS4-AS1 pull-down followed by western blotting to detect interactions with DHX9. (C) RIP assays were employed using DHX9 antibodies to determine if lncRNA DHRS4-AS1 co-precipitates with DHX9. IgG was used as a negative control that did not interact with DHRS4-AS1. (D) Western blot analysis showing that DHRS4-AS1 alters DHX9 expression. (E) qRT-PCR showed that DHRS4-AS1 has no effect on DHX9 mRNA levels. (F) DHX9 expression was measured in GC and AGS cells expressing Sh-DHRS4-AS1 or Sh-NC groups by western blotting after treatment with 50 µg/mL CHX. (G) DHX9 expression was measured in HGC-27 and MGD-803 cells overexpressing DHRS4-AS1 or empty vector using western blotting after treatment with 50 µg/mL CHX. (H) DHX9 expression in DHRS4-AS1-overexpressing and empty vector-expressing GC, HGC-27, and MGC-803 cells after treatment with 5 µM MG132 was evaluated using western blotting. (I) Western blot of DHX9 immunoprecipitated from DHRS4-AS-expressing GC cells to examine endogenous DHX9 ubiquitination.*p < 0.05, **p < 0.01,NS:Not significant