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Journal: Protein & Cell
Article Title: NDFIP1 limits cellular TAZ accumulation via exosomal sorting to inhibit NSCLC proliferation
doi: 10.1093/procel/pwac017
Figure Lengend Snippet: NDFIP1 interacts with TAZ, and TAZ can be sorted into exosomes, which requires NDFIP1. (A) The binding of NDFIP1 to TAZ in SPC-A1 and SPC-A1-BM. (B) The converse binding of TAZ to NDFIP1 in SPC-A1. (C) The binding of NDFIP1 to TAZ in 293T. (D) The binding of NDFIP1 to TAZ WT, but not to the TAZ ΔWW in SPC-A1. (E) IF co-localization of NDFIP1 and TAZ in SPC-A1 and SPC-A1-BM. Scale bars: 5 μm. (F) TEM image of SPC-A1 exosomes. Scale bars: 100 nm. (G) The particle size distribution of exosomes from SPC-A1. (H) The protein levels of YAP, TAZ, exosome-specific proteins, ALIX, TSG101, CD63, a cell internal reference, GAPDH and a Golgi marker, GM130 in different components of SPC-A1, including the pellets of centrifugation at 500 × g , 2,000 × g , 10,000 × g , exosomes and cells. # represents longer exposure. (I) IF co-localization of CD63 and TAZ in SPC-A1 and SPC-A1-BM. Scale bars: 5 μm. (J–K) The average number of secreted exosomes per cell of (J) the control (sgNC) and NDFIP1 knockout (sgNDFIP1) and (K) the control (PCDH) and NDFIP1 overexpression (NDFIP1-OE) SPC-A1 cells. Bars represent the mean ± SD ( n = 3). (J) ns, P = 0.1977. (K) ns, P = 0.8157. (L–M) The TAZ and NDFIP1 protein levels in equal number of exosomes from the (L) control (sgNC) and NDFIP1 knockout (sgNDFIP1) SPC-A1 cells, and (M) from the control (PCDH) and NDFIP1 overexpression (NDFIP1-OE) SPC-A1 cells.
Article Snippet: Briefly, the cells were incubated with the complex of plasmids, P3000 and Lipofectamine 3000 reagent, and the plasmid transfection efficiency was assessed after 48 h. The
Techniques: Binding Assay, Marker, Centrifugation, Knock-Out, Over Expression