Figure 5: Beclin 1/Mcl-1 axis modulates melanoma growth in vitro and in vivo.

(a) Beclin 1 and Mcl-1 levels in patient-derived melanomas compared with primary melanocytes. (b) Mcl-1 mRNA levels in patient-derived melanomas and primary melanocytes. Error bars indicate s.e.m. (n=3 for all graphs). (c) Mcl-1 ubiquitination and degradation on transient expression of Beclin 1 in patient-derived 1949 melanoma cells. Endogenous Mcl-1 was pulled down from GaLa 1949 transiently transfected with Beclin 1 or empty vector and the level of Mcl-1 ubiquitination as well as total protein levels was assessed by immunoblotting. (d) In vitro growth of GaLa1949 melanoma cells expressing either scrambled shRNA or Beclin 1 shRNA upon ablation of Mcl-1 with doxycycline-inducible shRNA. Results are representative of at least three independent experiments. Error bars represent s.e.m. (n=3 for all graphs). (e) In vivo growth of xenograft tumours derived from cells transduced with the indicated shRNA. Error bars indicate s.e.m. (n=8 per group). After establishment of xenografts, NSG mice were kept on 1 mg ml⁻¹ Dox supplemented in the drinking water to induce Mcl-1 shRNA. (f) Beclin 1 and Mcl-1 protein levels in xenografts isolated from animals in (c).