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. 2017 Aug 28;36(50):6850–6862. doi: 10.1038/onc.2017.295

Figure 3.

Figure 3

D285 structurally affects the activity and function of Vps34. (a) Cleavage of Vps34 at D285 weakened the interaction of Vps34 with Beclin1 (Bcn 1). Immunoblots of Myc-Beclin1 in M2 immunoprecipitates from HEK293T cells transfected as indicated. (b) Effect of blocking the cleavage on the in vitro lipid kinase activity of Vps34; bars, s.d. *P<0.05; **P<0.01. (c) and Supplementary Figure S1 Expression levels of EMT indicators and autophagic flux in parental cells or MCF-7 cells stably expressing Vps34, Vps34-D1, Vps34-ΔD1 and Vps34-Mt (D285E). (d) In vitro invasion of MCF-7 parental cells or MCF-7 cells stably expressing Vps34, Vps34-D1, Vps34-ΔD1 and Vps34-Mt (D285E). Column, average data of three independent experiments; bars, s.d. *P<0.05. (e) Cleavage of Vps34 at D285 diminishes its competence to promote colony formation. Clonogenic images of parental cells, MCF-7/Vps34, MCF-7/Vps34-ΔD1 and MCF-7/vsp34-Mt (D285E) photographed on day 14. (f) Colony numbers of cells as shown in Figure 3e were counted. Values plotted were means of three experiments plus or minus s.d. **P<0.01.