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. Author manuscript; available in PMC: 2016 Oct 31.
Published in final edited form as: Nature. 2015 Jul 13;524(7565):361–365. doi: 10.1038/nature14587

Extended Data Figure 2. MiT/TFE-dependent regulation of autophagy-lysosome gene expression in PDA cell lines.

Extended Data Figure 2

a) Chromatin immunoprecipitation analysis of FLAG-MITF (left) and FLAG-TFE3 (right) binding to autophagy-lysosome genes in 8902 and Panc1 cells, respectively. Histograms show the amount of immunoprecipitated DNA detected by qPCR normalized to input and plotted as relative enrichment over mock control. Error bars indicate mean ± s.e.m for N = 3 independent experiments. * p < 0.05. b) siRNA mediated knockdown of MITF in HupT3 and PL18 cells causes a decrease in autophagy-lysosome gene expression, assayed 48 hrs post siRNA transfection. * p < 0.05. c) Knockdown of TFE3 in PSN1, Panc1 and 8988T cells, or of TFEB in 8902 cells, causes a decrease in autophagy-lysosome gene expression. * p < 0.05. d) HPDE, HPNE, and QGP1 control cells show minimal changes in autophagy-lysosome gene expression upon knockdown of the MiT/TFE genes. e) Decreased autophagy-lysosome gene expression following MITF knockdown (left; PL18 cells; * p < 0.05) or TFE3 knockdown (right; 8988T cells; * p < 0.01) is rescued by transient ectopic expression of MITF or TFE3. Cells were transfected with expression constructs for MITF or TFE3 24 hrs post-siRNA transfection. After 48 hrs, gene expression was assayed. f) Expression of MITF-DN in HupT3 cells causes a decrease in autophagy-lysosome gene expression compared to control cells (left panel; * p < 0.02). Similar results are seen in PSN1 cells expressing Doxycycline (Dox)-inducible MITF-DN upon addition of 1 μg/ml of Dox for 48 hrs (right panel; * p < 0.05). For all graphs error bars indicate mean ± s.d. for N = 3 independent experiments.