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Fig. 5 | Journal of Experimental & Clinical Cancer Research

Fig. 5

From: Cyclin G2 suppresses Wnt/β-catenin signaling and inhibits gastric cancer cell growth and migration through Dapper1

Fig. 5

Cyclin G2 inhibits Wnt/β-catenin signaling through DPR1. a Cell lysates from COS-7 cells were immunoprecipitated with Dpr1 or IgG control antibody, followed by western blot analysis with a cyclin G2 antibody (upper panel). Total cellular protein from COS-7 cells was immunoprecipitated with an anti-cyclin G2 or IgG control antibody, then blotted with anti-DPR1 and anti-Dvl2 antibody, respectively (middle panel). Western blot analysis of endogenous Dpr1 and cyclin G2 protein level in COS-7 cells (lower panel). b Direct interaction of Dpr1 and cyclin G2 was detected using Duolink in situ PLA. Red spots represent the interaction of Dpr1 and cyclin G2. The nuclei were stained using DAPI and are shown in blue. Images were acquired using a confocal microscopy with a 40× objective. c Western blot analysis of the Dvl2 protein levels in the control or cyclin G2-overexpressing COS-7 cells treated with 25 μM MG132, 1 μM bafilomycin A1 (BFA1), or DMSO as a negative vehicle control for 6 h followed. d Overexpression of cyclin G2 down-regulated β-catenin and Dvl2 protein expression levels in COS-7 cells, whereas knockdown of Dpr1 attenuated the effect of cyclin G2. cyclin G2 overexpressing vector or control vector (NC) was co-transfected with Dpr1-specific (shDACT1) or nonspecific shRNA vectors (shNC) into COS-7 cells for 48 h followed by western blot analysis. e The phosphorylation level of Dpr1 was suppressed by cyclin G2. Cell lysates from HEK-293 and SGC-7901 cells overexpressing cyclin G2 or GFP as a negative control was immunoprecipitated with anti-Dpr1 antibodies, then immunoblotted with anti-Phosphoserine/threonine antibody (upper panel). Western blot analysis of Dpr1 and cyclin G2 expression in total cellular protein (lower panel). f Cyclin G2 inhibited the CKI-induced phosphorylation of Dpr1. HEK-293 cells were transfected with expression vectors encoding CKI (pEGFP-CKI) and cyclin G2 (pCMV-3 × FLAG-G2) or transfected with negative control vectors (pCMV-3 × FLAG-BAP or pEGFP-N3) for 48 h. The cell lysates were immunoprecipitated with Dpr1 antibodies and immunoblotted with anti-Phosphoserine antibody (upper panel). Dpr1 and cyclin G2 expression in total cellular protein was analysed by western blot (lower panel)

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