Figure 2. Atg7 deficiency has distinct consequences for tumor establishment and maintenance in Braf^V600E–driven lung tumors and is associated with the emergence of oncocytomas and failure of mitochondrial function.

A. Representative H&E staining of lung lobes at the indicated times post Cre. Scale bar= 3mm. Scanned H&E slides of the full time course are provided in Figure S2.

B. Quantification of tumor burden (two mice/group, all lobes analyzed) from H&E slides across the indicated time points via Matlab. Error bars indicate SEM. Scanned H&E slides of the full time course are provided in Figure S2.

C. Representative low power images of H&E slides 5, 7, 14 weeks post Cre. Scale bar= 100 μm. Higher magnification images are provided in the inset. Scale bar= 50 μm.

D. Representative high magnification images of H&E staining of tumor for the indicated time course. Note the increasing large cytoplasm in the Braf ^V600E/+; Atg7^−/− tumors indicative of oncocytoma. Scale bar=10 μm

E. Braf ^V600E/+; Atg7^−/− tumors have an accumulation of defective mitochondria. Representative images of Tom20 IHC, electron micrographs, and the enzymatic cytochrome c oxidase assay at the indicated time post Cre. For Tom20, Scale bar= 10μm. For EM, N=nucleus MT= mitochondria. Scale bar=500 nm. Arrows point to swollen mitochondria in Braf ^V600E/+; Atg7^−/− tumors. For cytochrome c oxidase, arrows point to tumor regions, indicated by dashed line, with defective mitochondria in the Braf ^V600E/+; Atg7^−/− tumors. Scale bar =100 μm.

F. Western blot of Cox-IV in tumor lysates of Braf ^V600E/+; Atg7^+/+ and Braf ^V600E/+; Atg7^−/− mice 10 weeks post Cre. Numbers refer to individual mice. A blank lane was intentionally included between the normal lung and tumor lysates. An additional Atg7 null tumor lysate was spliced out for aesthetic reasons.

G. Kaplan Meier analysis of overall survival of Braf ^V600E/+; Atg7^+/+ and Braf ^V600E/+; Atg7^−/− mice post Cre. Numbers of animals per group and median survival are indicated.

H. Kaplan Meier analysis of overall survival of Braf ^V600E/+; Beclin1^+/+ and Braf ^V600E/+; Beclin1^+/− mice post Cre. Numbers of animals per group and median survival are indicated.

I. Representative IHC of Ki67, p53 and p21 from Braf ^V600E/+; Atg7^+/+ and Braf ^V600E/+; Atg7^−/− mice 14 weeks post Cre. Scale bar=10 μm. Quantification of IHC at 5, 7, 14 weeks post Cre is shown below. Images for entire time course are provided in Figure S4.