Journal: OncoTargets and therapy

Article Title: Ubiquitin-specific peptidase 28 enhances STAT3 signaling and promotes cell growth in non-small-cell lung cancer

doi: 10.2147/OTT.S194917

Figure Lengend Snippet: USP28 interacts with STAT3 and decreases its polyubiquitination level. Notes: ( A , B ) Myc-USP28 or Flag-STAT3 plasmids were transfected into HEK293T for 36 hours, and then whole cell lysates were prepared for reciprocal Co-IP and immunoblotting against specific antibodies as indicated. ( C ) H1299 cells were lysed for Co-IP by using anti-STAT3 antibody, followed by immunoblotting against USP28 and STAT3. ( D ) HEK293T cells were transfected with Myc-USP28 or EV along with HA-Ub plasmids for 36 hours. Cell lysates were then prepared for IP using anti-IgG or anti-STAT3 antibody, followed by immunoblotting against STAT3 and Ub-K48 (using K48-linkage specific polyubiquitin antibody). In addition, whole cell lysates were also prepared for immunoblotting against STAT3, Myc and GAPDH. Abbreviations: EV, empty vector; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; HA-Ub, HA-ubiquitin; IB, immunoblotting; IP, immunoprecipitation; n.s., non-sense; STAT3, signal transducer and activator of transcription 3; USP, ubiquitin-specific protease.

Article Snippet: Anti-p-STAT3, anti-STAT3, anti-immunoglobulin G (IgG), and anti-K48-linkage specific polyubiquitin antibodies were purchased from Cell Signaling Technology (Danvers, MA, USA).

Techniques: Transfection, Co-Immunoprecipitation Assay, Western Blot, Plasmid Preparation, Immunoprecipitation