Fig. 1: An inner nuclear membrane complex composed of CTDNEP1-NEP1R1-MAN1 interacts with R-SMADs.
From: Suppression of TGF-β/SMAD signaling by an inner nuclear membrane phosphatase complex
A Proteins co-precipitating with CTDNEP1-FLAG as detected by mass spectrometry. The x-axis shows the log2 fold change of CTDNEP1-FLAG versus untagged control cell line; the y-axis shows the −log10 p-value estimated by the Significance B analysis71. CTDNEP1 partner NEP1R1 is labeled in light blue and MAN1 in orange. A two-sided t-test was performed, and the results plotted as a volcano plot in R (significance cut-off: −1 and 1 log2 fold change, –log10 adj.p-value = 1.3). B, C Immunoprecipitation of FLAG-tagged wild-type CTDNEP1 or phosphatase dead CTDNEP1D67E, D69T (B) or NEP1R1-HA (C) from detergent solubilized extracts of HeLa cells. Eluted proteins were analyzed by SDS-PAGE followed by immunoblotting with the indicated antibodies. Short and long exposures are shown and labeled SE and LE, respectively. D Proteins co-precipitating with V5-MAN1 as detected by mass spectrometry. The x-axis shows the log2 fold change of V5-MAN1 versus untagged control cell line; the y-axis shows the −log10 p-value estimated by the Significance B analysis71. CTDNEP1 and NEP1R1 are labeled in dark and light blue, respectively, and the R-SMADs are labeled in green. A two-sided t-test was performed, and the results plotted as a volcano plot in R (significance cut-off: −1 and 1 log2 fold change, –log10 adj.p-value = 1.3). E and F Immunoprecipitation of CTDNEP1 or CTDNEP1D67E, D69T-FLAG (E) or NEP1R1-HA (F) from detergent solubilized extracts of HeLa cells co-expressing V5-MAN1. Immunoprecipitations were performed in the absence or upon 1 hr treatment with 20 ng/ml of TGF-β3. Eluted proteins were analyzed by SDS-PAGE followed by immunoblotting with the indicated antibodies. The asterisk (*) indicates the light chain of the antibody used for immunoprecipitation.
