ATM kinase enables the functional axis of YAP,PML and p53 to ameliorate loss of Werner protein-mediated oncogenic senescence

Werner syndrome (WS) results from dysfunction of the WRN protein, and is associated with premature aging and early death. Here we report that loss of WRN function elicits accumulation of the Yes-associated protein (YAP protein), a major effector of the Hippo tumor suppressor pathway, both experimentally and in WS-derived fibroblasts. YAP upregulation correlates with slower cell proliferation and accelerated senescence, which are partially mediated by the formation of a complex between YAP and the PML protein, whose activity promotes p53 activation. The ATM kinase is necessary for YAP and PML accumulation in WRN-depleted cells. Notably, the depletion of either YAP or PML partially impairs the induction of senescence following WRN loss. Altogether, our findings reveal that loss of WRN activity triggers the activation of an ATM-YAP-PML-p53 axis, thereby accelerating cellular senescence. The latter has features of SASP (senescence-associated secretory phenotype), whose protumorigenic properties are potentiated by YAP, PML and p53 depletion.     

Effects of carnitine palmitoyltransferases on cancer cellular senescence

The carnitine palmitoyltransferase (CPT) family is essential for fatty acid oxidation. Recently, we found that CPT1C, one of the CPT1 isoforms, plays a vital role in cancer cellular senescence. However, it is unclear whether other isoforms (CPT1A, CPT1B, and CPT2) have the same effect on cellular senescence. This study illustrates the different effects of CPT knockdown on PANC-1 cell proliferation and senescence and MDA-MB-231 cell proliferation and senescence, as demonstrated by cell cycle kinetics, Bromodeoxyuridine incorporation, senescence-associated β-galactosidase activity, colony formation, and messenger RNA (mRNA) expression of key senescence-associated secretory phenotype factors. CPT1C exhibits the most substantial effect on cell senescence. Lipidomics analysis was performed to further reveal that the knockdown of CPTs changed the contents of lipids involved in mitochondrial function, and lipid accumulation was induced. Moreover, the different effects of the isoform deficiencies on mitochondrial function were measured and compared by the level of radical oxygen species, mitochondrial transmembrane potential, and the respiratory capacity, and the expression of the genes involved in mitochondrial function were determined at the mRNA level. In summary, CPT1C exerts the most significant effect on mitochondrial dysfunction-associated tumor cellular senescence among the members of the CPT family, which further supports the crucial role of CPT1C in cellular senescence and suggests that inhibition of CPT1C may represent as a new strategy for cancer treatment through the induction of tumor senescence.     

Downregulation of Wnt signaling is a trigger for formation of facultative heterochromatin and onset of cell senescence in primary human cells

Cellular senescence is an irreversible proliferation arrest of primary cells and an important tumor suppression process. Senescence is often characterized by domains of facultative heterochromatin, called senescence-associated heterochromatin foci (SAHF), which repress expression of proliferation-promoting genes. Formation of SAHF is driven by a complex of histone chaperones, HIRA and ASF1a, and depends upon prior localization of HIRA to PML nuclear bodies. However, how the SAHF assembly pathway is activated in senescent cells is not known. Here we show that expression of the canonical Wnt2 ligand and downstream canonical Wnt signals are repressed in senescent human cells. Repression of Wnt2 occurs early in senescence and independently of the pRB and p53 tumor suppressor proteins and drives relocalization of HIRA to PML bodies, formation of SAHF and senescence, likely through GSK3beta-mediated phosphorylation of HIRA. These results have major implications for our understanding of both Wnt signaling and senescence in tissue homeostasis and cancer progression.     

CK2 mediates phosphorylation and ubiquitin-mediated degradation of the PML tumor suppressor

The PML tumor suppressor controls growth suppression, induction of apoptosis, and cellular senescence. PML loss occurs frequently in hematopoietic and solid tumors. PML loss often correlates with tumor progression. Casein kinase 2 (CK2) is a stress-activated serine/threonine protein kinase that is oncogenic and frequently overexpressed in human tumor of multiple histological origins. In addition, CK2 overexpression due to gene amplification has been reported to be an adverse prognostic factor in non-small cell lung cancer. At the 5th International Conference on Protein Kinase CK2 in Padova, Italy, we reviewed our recent findings that PML undergoes ubiquitin/proteasome-mediated degradation in immortalized and tumor derived cell lines. PML degradation depends on direct CK2 phosphorylation of PML Ser517. PML mutants that are resistant to CK2 phosphorylation display increased tumor suppressive functions in assays measuring apoptosis, replicative senescence, and in xenograft models. More significantly, CK2 pharmacological inhibition enhances PML tumor suppressive property. These data identify a key post-translational mechanism that controls PML protein levels in cancer cells and suggest that CK2 inhibitors may be beneficial anti-cancer drugs.     

Definition of pRB- and p53-dependent and -independent steps in HIRA/ASF1a-mediated formation of senescence-associated heterochromatin foci

Cellular senescence is an irreversible proliferation arrest triggered by short chromosome telomeres, activated oncogenes, and cell stress and mediated by the pRB and p53 tumor suppressor pathways. One of the earliest steps in the senescence program is translocation of a histone chaperone, HIRA, into promyelocytic leukemia (PML) nuclear bodies. This relocalization precedes other markers of senescence, including the appearance of specialized domains of facultative heterochromatin called senescence-associated heterochromatin foci (SAHF) and cell cycle exit. SAHF represses expression of proliferation-promoting genes, thereby driving exit from the cell cycle. HIRA bound to another histone chaperone, ASF1a, drives formation of SAHF. Here, we show that HIRA's translocation to PML bodies occurs in response to all senescence triggers tested. Dominant negative HIRA mutants that block HIRA's localization to PML bodies prevent formation of SAHF, as does a PML-RARalpha fusion protein which disrupts PML bodies, directly supporting the idea that localization of HIRA to PML bodies is required for formation of SAHF. Significantly, translocation of HIRA to PML bodies occurs in the absence of functional pRB and p53 tumor suppressor pathways. However, our evidence indicates that downstream of HIRA's localization to PML bodies, the HIRA/ASF1a pathway cooperates with pRB and p53 to make SAHF, with the HIRA/ASF1a and pRB pathways acting in parallel. We present evidence that convergence of the HIRA/ASF1a and pRB pathways occurs through a DNAJ-domain protein, DNAJA2.     

Interleukin 6 signaling regulates promyelocytic leukemia protein gene expression in human normal and cancer cells

Tumor suppressor PML is induced under viral and genotoxic stresses by interferons and JAK-STAT signaling. However, the mechanism responsible for its cell type-specific regulation under non-stimulated conditions is poorly understood. To analyze the variation of PML expression, we utilized three human cell types, BJ fibroblasts and HeLa and U2OS cell lines, each with a distinct PML expression pattern. Analysis of JAK-STAT signaling in the three cell lines revealed differences in levels of activated STAT3 but not STAT1 correlating with PML mRNA and protein levels. RNAi-mediated knockdown of STAT3 decreased PML expression; both STAT3 level/activity and PML expression relied on IL6 secreted into culture media. We mapped the IL6-responsive sequence to an ISRE(-595/-628) element of the PML promoter. The PI3K/Akt/NFκB branch of IL6 signaling showed also cell-type dependence, being highest in BJ, intermediate in HeLa, and lowest in U2OS cells and correlated with IL6 secretion. RNAi-mediated knockdown of NEMO (NF-κ-B essential modulator), a key component of NFκB activation, suppressed NFκB targets LMP2 and IRF1 together with STAT3 and PML. Combined knockdown of STAT3 and NEMO did not further promote PML suppression, and it can be bypassed by exogenous IL6, indicating the NF-κB pathway acts upstream of JAK-STAT3 through induction of IL6. Our results indicate that the cell type-specific activity of IL6 signaling pathways governs PML expression under unperturbed growth conditions. As IL6 is induced in response to various viral and genotoxic stresses, this cytokine may regulate autocrine/paracrine induction of PML under these pathophysiological states as part of tissue adaptation to local stress.     

PML control of cytokine signaling

The promyelocytic leukemia (PML) protein is a tumor suppressor acting as the organizer of nuclear matrix-associated structures named nuclear bodies (NBs). The involvement of PML in various cell processes, including cell death, senescence or antiviral defense underlines the multiple functions of PML due to its ability to interact with various partners either in the cytoplasm or in the nucleus. The importance of paracrine signaling in the regulation of PML expression is well established. More recently, a growing body of evidence also supports PML as a key regulator of cytokine signaling. These findings shed light on unsuspected biological functions of PML such as immune response, inflammation and cytokine-induced apoptosis. Here we review the current understanding of the pleiotropic activities of PML on cytokine-induced signaling.     

综述: 从分子水平看c-Myc在细胞衰老中的作用

细胞衰老是指细胞生长永久阻滞于细胞周期的G1期,出现形态、生化及表观遗传的变化特性.细胞衰老由端粒缩短、DNA损伤、缺氧或癌基因失调等因素引起,它是抵抗肿瘤发生的主要壁垒.原癌基因c-myc编码转录因子,可调控很多基因,进而影响细胞周期演进、衰老、凋亡、代谢等生物学过程.c-Myc蛋白与细胞衰老密切相关,它可影响hTERT、p16、p53、Bmi-1和p27等衰老相关基因转录.c-Myc不仅可抑制复制性衰老,也能抑制癌基因诱发的衰老.c-Myc抑制ras诱导的细胞衰老取决于CDK2.c-Myc失活不仅能够诱导非恶性细胞(如人成纤维细胞)衰老,而且在许多肿瘤细胞中也可诱导衰老.然而,与ras基因类似,在特定条件下,c-Myc也可诱导细胞衰老,并可促进维氏综合症(Werner syndrome,WRN)缺失细胞的衰老.