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Mol Cell Proteomics. 2017 Jul;16(7):1217-1232. doi: 10.1074/mcp.M115.050179. Epub 2015 Dec 10.

Lowering Endogenous Cathepsin D Abundance Results in Reactive Oxygen Species Accumulation and Cell Senescence.

Su S1,2, Zhu X1, Lin L3, Chen X1,2, Wang Y1,2, Zi J3, Dong Y4, Xie Y1,2, Zhu Y1,2, Zhang J1, Zhu J1, Xu D1, Xu N5, Lou X6, Liu S6,2.

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From the ‡CAS Key Laboratory of Genome Sciences and Information, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing, China, 100101.
§University of Chinese Academy of Sciences, Beijing, China, 100049.
¶Proteomics Division, BGI-Shenzhen, Shenzhen, Guangdong, China, 518083.
‖Beijing Protein Innovation, Beijing, China, 101318.
**Laboratory of Cell and Molecular Biology, Cancer Institute and Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China, 100021.
From the ‡CAS Key Laboratory of Genome Sciences and Information, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing, China, 100101;


Cathepsin D is reportedly to be closely associated with tumor development, migration, and invasion, but its pathological mechanism is not fully elucidated. We aimed to evaluate phenotypic changes and molecular events in response to cathepsin D knockdown. Lowering endogenous cathepsin D abundance (CR) induced senescence in HeLa cells, leading to reduced rate of cell proliferation and impaired tumorigenesis in a mouse model. Quantitative proteomics revealed that compared with control cells (EV), the abundances of several typical lysosomal proteases were decreased in the lysosomal fraction in CR cells. We further showed that cathepsin D knockdown caused increased permeability of lysosomal membrane and reactive oxygen species accumulation in CR cells, and the scavenging of reactive oxygen species by antioxidant was able to rescue cell senescence. Despite the increased reactive oxygen species, the proteomic data suggested a global reduction of redox-related proteins in CR cells. Subsequent analysis indicated that the transcriptional activity of nuclear factor erythroid-related factor 2 (Nrf2), which regulates the expression of groups of antioxidant enzymes, was down-regulated by cathepsin D knockdown. Importantly, Nrf2 overexpression significantly reduced cell senescence. Although transient oxidative stress promoted the accumulation of Nrf2 in the nucleus, we showed that the Nrf2 protein exited nucleus if oxidative stress persisted. In addition, when cathepsin D was transiently knocked down, the cathepsin-related events followed a sequential order, including lysosomal leakage during the early stage, followed by oxidative stress augmentation, and ultimately Nrf2 down-regulation and senescence. Our results suggest the roles of cathepsin D in cancer cells in maintaining lysosomal integrity, redox balance, and Nrf2 activity, thus promoting tumorigenesis. The MS Data are available via ProteomeXchange with identifier PXD002844.

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