Fig. 6.

USP39 promotes the ADAM9 pre‐mRNA splicing. (A) qRT‐PCR analysis exhibiting relative mRNA levels of mature and precursor ADAM9 RNA transcripts and the ratio in U251 cells treated with USP39 siRNA relative to controls. (B) Western blotting analysis of USP39 and ADAM9 expression levels in U251 cells infected with Control‐shRNA‐expressing lentiviruses (shNC) or USP39‐shRNA‐expressing lentiviruses (shUSP39). (C) qRT‐PCR analysis showing relative mRNA levels of mature and precursor ADAM9 RNA transcripts and the ratio in U87 cells transfected with USP39 overexpressed plasmid relative to controls. (D) Western blotting analyzed USP39 and ADAM9 expression in U87 cells transfected with control plasmid (NC) or USP39 overexpressed plasmid (ov‐USP39). (E) RIP assay was conducted using anti‐USP39 (with IgG as control), and RT‐PCR were used to determine the mRNA level of ADAM9 in the immunoprecipitated complex. (F) Schematic model (top) representation of a region of ADAM9 gene used to prepare DNA template to synthesize pre‐mRNA substrate for in vitro splicing assay. Bottom: PCR fragment amplified with 5′ and 3′ primers using the DNA template from spliced product using nuclear extract from HEK293T cells carrying control vector or USP39 overexpressed plasmid. All data are presented as means ± SD. Significance calculated using Student’s t‐test. **P < 0.01, ***P < 0.001. Representative data are from three independent experiments.