Promyelocytic leukemia (PML) plays a tumor suppressive role by inducing cellular senescence in response to oncogenic stress. However, tumor cell lines fail to engage in complete senescence upon PML activation. In this study, we investigated the mechanisms underlying resistance to PML-induced senescence. Here, we report that activation of the cyclin-dependent kinases CDK4 and CDK6 are essential and sufficient to impair senescence induced by PML expression. Disrupting CDK function by RNA interference or pharmacological inhibition restored senescence in tumor cells and diminished their tumorigenic potential in mouse xenograft models. Complete senescence correlated with an increase in autophagy, repression of E2F target genes, and an gene expression signature of blocked DNA methylation. Accordingly, treatment of tumor cells with inhibitors of DNA methylation reversed resistance to PMLinduced senescence. Further, CDK inhibition with palbociclib promoted autophagy-dependent degradation of the DNA methyltransferase DNMT1. Lastly, we found that CDK4 interacted with and phosphorylated DNMT1 in vitro, suggesting that CDK activity is required for its stabilization. Taken together, our findings highlight a potentially valuable feature of CDK4/6 inhibitors as epigenetic modulators to facilitate activation of senescence programs in tumor cells. Cancer Res; 76(11); 3252-64. Ó2016 AACR.

Expression of the suppressor of cytokine signaling-1 (SOCS1) is inactivated in hematopoietic and solid cancers by promoter methylation, miRNA-mediated silencing, and mutations. Paradoxically, SOCS1 is also overexpressed in many human cancers. We report here that the ability of SOCS1 to interact with p53 and regulate cellular senescence depends on a structural motif that includes tyrosine (Y)80 in the SH2 domain of SOCS1. Mutations in this motif are found at low frequency in some human cancers, and substitution of Y80 by a phosphomimetic residue inhibits p53-SOCS1 interaction and its functional consequences, including stimulation of p53 transcriptional activity, growth arrest, and cellular senescence. Mass spectrometry confirmed SOCS1 Y80 phosphorylation in cells, and a new mAb was generated to detect its presence in tissues by IHC. A tyrosine kinase library screen identified the SRC family as Y80-SOCS1 kinases. SRC family kinase inhibitors potentiated the SOCS1-p53 pathway and reinforced SOCS1-induced senescence. Samples from human lymphomas that often overexpress SOCS1 also displayed SRC family kinase activation, constitutive phosphorylation of SOCS1 on Y80, and SOCS1 cytoplasmic localization. Collectively, these results reveal a mechanism that inactivates the SOCS1-p53 senescence pathway and suggest that inhibition of SRC family kinases as personalized treatment in patients with lymphomas may be successful.Significance: These findings show that SOCS1 phosphorylation by the SRC family inhibits its tumor-suppressive activity, indicating that patients with increased SOCS1 phosphorylation may benefit from SRC family kinase inhibitors.

Several regulators of SUMOylation have been previously linked to senescence but most targets of this modification in senescent cells remain unidentified. Using a two-step purification of a modified SUMO3, we profiled the SUMO proteome of senescent cells in a site-specific manner. We identified 25 SUMO sites on 23 proteins that were significantly regulated during senescence. Of note, most of these proteins were PML nuclear body (PML-NB) associated, which correlates with the increased number and size of PML-NBs observed in senescent cells. Interestingly, the sole SUMO E2 enzyme, UBC9, was more SUMOylated during senescence on its Lys-49. Functional studies of a UBC9 mutant at Lys-49 showed a decreased association to PML-NBs and the loss of UBC9’s ability to delay senescence. We thus propose both pro- and anti-senescence functions of protein SUMOylation.

BACKGROUNDDespite the increasing frequency of adenocarcinomas of the uterine cervix, little is known regarding inactivation of tumor suppressor genes (TSGs) in this tumor type. The authors analyzed loss of heterozygosity (LOH) in 36 carcinomas of the cervix with glandular differentiation, and 5 adenocarcinoma in situ in 40 patients.METHODSThe authors analyzed samples using laser capture microdissection from archival material and DNA amplified with microsatellite markers on the following loci: 3p14.2 (D3S1234, D3S1300), 3p21.3 (D3S1029, D3S1447), 3p22‐24 (D3S1537, D3S1351), 6q21‐23.3 (D6S250), 6q25.1 (ESR), 6q25.2 (D6S255), 8p21 (D8S136, D8S1820), 13q12.3 (D13S220, D13S267), 17q21 (D17S579, D17S855). Eight additional markers spanning the short arm of chromosome 3 (3p12‐p25) and six spanning the long arm of chromosome 6 (6q11‐q27) were studied in the cases showing LOH to further define the deletion intervals.RESULTSThe frequency of allelic loss in cancers was chromosome 3p: 49% (p14.2: 35%, p21.3: 23%, p22‐24: 41%), 6q: 48% (q21‐23.1: 39%, q25.1: 45%, q25.2: 7%), 13q: 22%, 17q: 6%, and 8p: 18%. On chromosome arm 3p, the authors' data suggest at least two discrete areas of deletion: a proximal area between markers D3S1234 (p12) and D3S1766 (p14.2‐14.3), and a second distal interval, telomeric from marker D3S4623 (p21.3). On chromosome 6q, the deletion area is between marker D6S300 (q22) and D6S255 (q25.2). Two of five preneoplastic lesions showed LOH on chromosome arm 3p, and two five showed allelic loss on chromosome arm on 6q, suggesting the genes might be inactivated early in cervical tumorigenesis.CONCLUSIONSThe authors have identified three chromosomal regions that may harbor TSGs involved in the development/progression of adenocarcinomas of the uterine cervix, 3p12‐14.2, 3p21.3‐pter, and 6q22‐25.2. Deletions also were detected in adenocarcinoma in situ, suggesting the genes may be inactivated early in cervical tumorigenesis. Cancer 2002;94:793–802. © 2002 American Cancer Society.DOI 10.1002/cncr.10275

Antiviral innate immune response to RNA virus infection is supported by Pattern-Recognition Receptors (PRR) including RIG-I-Like Receptors (RLR), which lead to type I interferons (IFNs) and IFN-stimulated genes (ISG) production. Upon sensing of viral RNA, the E3 ubiquitin ligase TNF Receptor-Associated Factor-3 (TRAF3) is recruited along with its substrate TANK-Binding Kinase (TBK1), to MAVS-containing subcellular compartments, including mitochondria, peroxisomes, and the mitochondria-associated endoplasmic reticulum membrane (MAM). However, the regulation of such events remains largely unresolved. Here, we identify TRK-Fused Gene (TFG), a protein involved in the transport of newly synthesized proteins to the endomembrane system via the Coat Protein complex II (COPII) transport vesicles, as a new TRAF3-interacting protein allowing the efficient recruitment of TRAF3 to MAVS and TBK1 following Sendai virus (SeV) infection. Using siRNA and shRNA approaches, we show that TFG is required for virus-induced TBK1 activation resulting in C-terminal IRF3 phosphorylation and dimerization. We further show that the ability of the TRAF3-TFG complex to engage mTOR following SeV infection allows TBK1 to phosphorylate mTOR on serine 2159, a post-translational modification shown to promote mTORC1 signaling. We demonstrate that the activation of mTORC1 signaling during SeV infection plays a positive role in the expression of Viperin, IRF7 and IFN-induced proteins with tetratricopeptide repeats (IFITs) proteins, and that depleting TFG resulted in a compromised antiviral state. Our study, therefore, identifies TFG as an essential component of the RLR-dependent type I IFN antiviral response.