DNA Damage and Repair SIRT 2 Interacts with b-Catenin to Inhibit Wnt Signaling Output in Response to Radiation-Induced Stress

Wnt signaling is critical tomaintaining cellular homeostasis via regulation of cell division,mitigation of cell stress, and degradation. Aberrations in Wnt signaling contribute to carcinogenesis and metastasis, whereas sirtuins have purported roles in carcinogenesis, aging, and neurodegeneration. Therefore, the hypothesis that sirtuin 2 (SIRT2) directly interacts withb-catenin andwhether this interaction alters the expression ofWnt target genes to produce an altered cellular phenotype was tested. Coimmunoprecipitation studies, using mouse embryonic fibroblasts (MEF) from Sirt2wild-type and genomic knockoutmice, demonstrate thatb-catenin directly binds SIRT2.Moreover, this interaction increases in response to oxidative stress induced by ionizing radiation. In addition, this association inhibits the expression of important Wnt target genes such as survivin (BIRC5), cyclin D1 (CCND1), and c-myc (MYC). In Sirt2 null MEFs, an upregulation of matrix metalloproteinase 9 (MMP9) and decreased E-cadherin (CDH1) expression is observed that produces increased cellular migration and invasion. Together, these data demonstrate that SIRT2, a tumor suppressor lost in multiple cancers, inhibits the Wnt signaling pathway in nonmalignant cells by binding to b-catenin and that SIRT2 plays a critical role in the response to oxidative stress from radiation. Implications: Disruption of the SIRT2–b-catenin interaction represents an endogenous therapeutic target to prevent transformation and preserve the integrity of aging cells against exogenous stressors such as reactive oxygen species. Mol Cancer Res; 1–10. 2014 AACR. Introduction The Wnt signaling pathway is crucial in regulating cell proliferation, apoptosis, migration, and differentiation (1). b-Catenin is a critical coactivator of Wnt-mediated gene expression. When Wnt ligands are present, b-catenin accumulates in the cytoplasm and is transported to the nucleus whereupon it binds to the lymphocyte enhancer factor (LEF)/T-cell factor (TCF) complex to recruit chromatin remodeling complexes and activates gene expression (2–4). Disruptions of Wnt signaling are associated with accelerated aging (5, 6) and abnormal neural development (7) as well as several major diseases, including schizophrenia, autism, Alzheimer disease, and cardiovascular disease (3, 8, 9). In addition, aberrant regulation of the b-catenin pathway has been shown to promote tumorigenesis, including colon, breast, prostate, ovary, and melanoma cancers (10). Sirtuin 2 (SIRT2) belongs to the family of sirtuins that are NADþ-dependent class III protein deacetylases, and has been shown to play a major role in metabolism and longevity. SIRT2 expresses mainly in the brain, particularly the hippocampus and oligodendrocytes. SIRT2 is reported to deacetylate a-tubulin, thereby regulating microtubule dynamics. Consequently, SIRT2 is proposed to play a role in mitotic checkpoint to maintain genomic integrity (11– 13). It has also been observed that SIRT2may act as a tumor suppressor because SIRT2 is downregulated in gliomas (14), while high levels of SIRT2 in tumors confer resistance to chemotherapy (15). In the central nervous system, SIRT2 is believed to participate in oligodendrocyte differentiation (16), and SIRT2 has been found to accumulate in the aging rodent brain (17). In this study, we show that SIRT2 directly interacts with b-catenin, and the interaction increases following ionizing radiation exposure. Genetic deletion of Sirt2 leads to activation of Wnt target genes such as c-myc, cyclin D1, and survivin. Furthermore, we observe that SIRT2 is critical for regulating cell migration through induction of matrix metalloproteinase 9 (MMP-9) and downregulation of Ecadherin. These findings provide a mechanistic insight into SIRT2 as a potential tumor suppressor and in cellular response to ionizing radiation-induced oxidative stress in Radiation Oncology Branch, NCI, NIH, Bethesda, Maryland. Medical Oncology Branch, NCI, NIH, Bethesda, Maryland. Note: Supplementary data for this article are available at Molecular Cancer Research Online (http://mcr.aacrjournals.org/). Corresponding Author: DeeDee K. Smart, Radiation Oncology Branch, NCI, NIH, 9000 Rockville Pike, Bethesda, MD 20892. Phone: 301-4965457; Fax: 301-480-5439; E-mail: smartd@mail.nih. gov doi: 10.1158/1541-7786.MCR-14-0223-T 2014 American Association for Cancer Research. Molecular Cancer Research www.aacrjournals. org OF1 on June 27, 2017. © 2014 American Association for Cancer Research. mcr.aacrjournals. org Downloaded from Published OnlineFirst May 27, 2014; DOI: 10.1158/1541-7786.MCR-14-0223-T