Autophagy is an evolutionarily conserved catabolic process whereby cellular organelles and bulk cytoplasm are targeted to lysosomes for degradation. Autophagy is a survival pathway required for cell viability during starvation; however, if it proceeds to completion, autophagy can lead to cell death. Autophagy also plays a role in tumorigenesis, as the essential autophagy regulator beclin 1 is monoallelically deleted in many human ovarian, breast, and prostate cancers, and beclin 1+/- mice are tumor-prone. Thus, autophagy likely suppresses tumorigenesis, but the mechanism has been unknown. We recently showed that allelic loss of beclin 1 compromises the autophagy potential of immortalized mouse mammary epithelial cells (iMMECs) in vitro and in mammary tumors in vivo, sensitizes iMMECs to metabolic stress and accelerates lumen formation in mammary acini. Autophagy defects also activate the DNA damage response in vitro and in mammary tumors in vivo, promote gene amplification, and synergize with defective apoptosis to accelerate mammary tumorigenesis.
We investigated the mechanism by which autophagy mitigates metabolic stress and limits genome damage in mammary epithelial cells, so as to gain insight into how autophagy suppresses mammary tumorigenesis. A proteomic analysis was performed to determine how autophagy-competent mammary epithelial cells respond successfully to metabolic stress as compared to cells with autophagy defects. Protein extracts from apoptosis-defective beclin 1+/+ and beclin 1+/- iMMECs exposed to metabolic stress for 0, 4 and 7 days were analyzed by two-dimensional difference gel electrophoresis (2D-DIGE). More than a hundred proteins with differential expression under metabolic stress were sequenced and mostly represented members of the GRP chaperone system, cellular metabolism, mitochondrial and cytoskeletal proteins. The 2D-DIGE results were validated for a subset of the sequenced proteins on beclin 1+/+ and beclin 1+/- iMMECs under metabolic stress in vitro and on beclin 1+/+ and beclin 1+/- iMMEC-generated mammary tumors in vivo. Expression of several proteins upregulated under metabolic stress was also examined in 3D-morphogenesis assays. For most proteins, expression was higher in the central acinar cells, indicating that the acinar center is a physiologic area of increased metabolic stress. Differential protein expression between wild-type and autophagy-defective iMMECs under conditions of metabolic stress is currently being validated on mammary tissues from beclin 1+/+ and beclin 1+/- female mice.
Therefore, failure of autophagy-deficient mammary tumor cells to effectively manage metabolic stress is associated with increased unfolded protein load and endoplasmic reticulum (ER) stress, which may in turn play a pivotal role in promoting genome damage and instability, and thus cancer progression, especially when apoptosis is concurrently inactivated. These studies identify new therapeutic targets and provide valuable clues into how to best modulate autophagy for cancer prevention and treatment.
Citation Information: Cancer Prev Res 2008;1(7 Suppl):CN05-02.
- American Association for Cancer Research