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Fig. 2 | Cell Communication and Signaling

Fig. 2

From: TRIM30 modulates Interleukin-22-regulated papillary thyroid Cancer cell migration and invasion by targeting Sox17 for K48-linked Polyubiquitination

Fig. 2

TRIM30 is involved in IL-22 regulated K48-linked polyubiquitination of Sox17. (a) TPC-1 cells were treated with 50 ng/ml rhIL-22 for the indicated time points. TRIM30 RNA levels were quantified using qRT-PCR (upper panel) and protein levels of TRIM30 and Sox17 were measured using western blot (lower panel). (b) TPC-1 cells were treated with rhIL-22 for 24 h at indicated concentrations. TRIM30 RNA levels were quantified using qRT-PCR (upper panel) and protein levels of TRIM30 and Sox17 were measured using western blot (lower panel). (c) TPC-1 cells were transfected with indicated plasmids for 24 h and treated with or without 50 ng/ml rhIL- 22 for 24 h prior to real-time RT-PCR (upper panel) and western blot (lower panel). (d) Experiments were performed as in (c) except cells were transfected with TRIM30-specific shRNA (shRNA-TRIM30). (e) TPC-1 cells were transfected with pCMV-TRIM30 or control vector for 48 h and treated with or without cycloheximide (CHX) for indicated times prior to western blot. (f) Overexpressed TRIM30 enhances wild-type and K48-linked polyubiquitination of Sox17. A total of 293 cells (2 × 106) were transfected with Sox17 or/and Flag-TRIM30 and the indicated ubiquitin plasmids for 24 h and treated with MG132 (100 μM) for 6 h. Cell lysates were immunoprecipitated with anti-Myc. The immunoprecipitates were analyzed by immunoblots with anti-HA, anti-Myc, or anti-Flag as indicated. (g) TPC-1 cells were treated with 50 ng/ml rhIL-22 for the indicated time points. Co-IP and immunoblot analyses were performed with the indicated antibodies. (h) TPC-1 cells were transfected with indicated plasmid for 24 h and treated with or without 50 ng/ml rhIL- 22 for 24 h. Co-IP and immunoblot analyses were performed with the indicated antibodies. (i) Experiments were performed as in (c) except cells were transfected with shRNA-TRIM30. All experiments were repeated at least three times. Bar graphs present means ± SD, n = 3 (**P < 0.01; *P < 0.05)

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