Poly(ADP-ribose) polymerase family member 14 (PARP14) is a novel effector of the JNK2-dependent pro-survival signal in multiple myeloma

Copyright @ 2013 Macmillan Publishers Limited. This is the author's accepted manuscript. The final published article is available from the link below.Regulation of cell survival is a key part of the pathogenesis of multiple myeloma (MM). Jun N-terminal kinase (JNK) signaling has been implicated in MM pathogenesis, but its function is unclear. To elucidate the role of JNK in MM, we evaluated the specific functions of the two major JNK proteins, JNK1 and JNK2. We show here that JNK2 is constitutively activated in a panel of MM cell lines and primary tumors. Using loss-of-function studies, we demonstrate that JNK2 is required for the survival of myeloma cells and constitutively suppresses JNK1-mediated apoptosis by affecting expression of poly(ADP-ribose) polymerase (PARP)14, a key regulator of B-cell survival. Strikingly, we found that PARP14 is highly expressed in myeloma plasma cells and associated with disease progression and poor survival. Overexpression of PARP14 completely rescued myeloma cells from apoptosis induced by JNK2 knockdown, indicating that PARP14 is critically involved in JNK2-dependent survival. Mechanistically, PARP14 was found to promote the survival of myeloma cells by binding and inhibiting JNK1. Moreover, inhibition of PARP14 enhances the sensitization of MM cells to anti-myeloma agents. Our findings reveal a novel regulatory pathway in myeloma cells through which JNK2 signals cell survival via PARP14, and identify PARP14 as a potential therapeutic target in myeloma.Kay Kendall Leukemia Fund, NIH, Cancer Research UK, Italian Association for Cancer Research and the Foundation for Liver Research

JNK signalling in cancer: In need of new, smarter therapeutic targets

Copyright © 2013 The British Pharmacological Society. This is the accepted version of the following article: Bubici, C. and Papa, S. (2014), JNK signalling in cancer: in need of new, smarter therapeutic targets. British Journal of Pharmacology, 171: 24–37, which has been published in final form at http://onlinelibrary.wiley.com/doi/10.1111/bph.12432/abstract.The JNKs are master protein kinases that regulate many physiological processes, including inflammatory responses, morphogenesis, cell proliferation, differentiation, survival and death. It is increasingly apparent that persistent activation of JNKs is involved in cancer development and progression. Therefore, JNKs represent attractive targets for therapeutic intervention with small molecule kinase inhibitors. However, evidence supportive of a tumour suppressor role for the JNK proteins has also been documented. Recent studies showed that the two major JNK proteins, JNK1 and JNK2, have distinct or even opposing functions in different types of cancer. As such, close consideration of which JNK proteins are beneficial targets and, more importantly, what effect small molecule inhibitors of JNKs have on physiological processes, are essential. A number of ATP-competitive and ATP-non-competitive JNK inhibitors have been developed, but have several limitations such as a lack of specificity and cellular toxicity. In this review, we summarize the accumulating evidence supporting a role for the JNK proteins in the pathogenesis of different solid and haematological malignancies, and discuss many challenges and scientific opportunities in the targeting of JNKs in cancer.Kay Kendall Leukemia Fund, Italian Association for Cancer Research and Foundation for Liver Research

Phosphorylation of proteins and apoptosis induced by c-Jun N-terminal kinase1 activation in rat cardiomyocytes by H2O2 stimulation

AbstractCytokines and various cellular stresses are known to activate c-Jun N-terminal kinase-1 (JNK1), which is involved in physiological function. Here, we investigate the activation of JNK1 by oxidative stress in H9c2 cells derived from rat cardiomyocytes. H2O2 (100 μM) significantly induces the tyrosine phosphorylation of JNK1 with a peak 25 min after the stimulation. The amount of JNK1 protein remains almost constant during stimulation. Immunocytochemical observation shows that JNK1 staining in the nucleus is enhanced after H2O2 stimulation. To clarify the physiological role of JNK1 activation under these conditions, we transfected antisense JNK1 DNA into H9c2 cells. The antisense DNA (2 μM) inhibits JNK1 expression by 80% as compared with expression in the presence of the sense DNA, and significantly blocks H2O2-induced cell death. Consistent with the decrease in cell number, we detected condensation of the nuclei, a hallmark of apoptosis, 3 h after H2O2 stimulation in the presence of the sense DNA for JNK1. The antisense DNA of JNK1 inhibits the condensation of nuclei by H2O2. Under these conditions, the H2O2-induced phosphorylation of proteins with molecular masses of 55, 72, and 78 kDa is blocked by treatment with the antisense DNA for JNK1 as compared with the sense DNA for JNK1. These findings suggest that JNK1 induces apoptotic cell death in response to H2O2, and that the cell death may be involved in the phosphorylations of 55, 72, and 78 kDa proteins induced by JNK1 activation

c-Jun N-Terminal Kinase in Inflammation and Rheumatic Diseases.

The c-Jun N-terminal kinases (JNKs) are members of the mitogen-activated protein kinase (MAPK) family and are activated by environmental stress. JNK is also activated by proinflammatory cytokines, such as TNF and IL-1, and Toll-like receptor ligands. This pathway, therefore, can act as a critical convergence point in immune system signaling for both adaptive and innate responses. Like other MAPKs, the JNKs are activated via the sequential activation of protein kinases that includes two dual-specificity MAP kinase kinases (MKK4 and MKK7) and multiple MAP kinase kinase kinases. MAPKs, including JNKs, can be deactivated by a specialized group of phosphatases, called MAP kinase phosphatases. JNK phosphorylates and regulates the activity of transcription factors other than c-Jun, including ATF2, Elk-1, p53 and c-Myc and non-transcription factors, such as members of the Bcl-2 family. The pathway plays a critical role in cell proliferation, apoptosis, angiogenesis and migration. In this review, an overview of the functions that are related to rheumatic diseases is presented. In addition, some diseases in which JNK participates will be highlighted

Spinal release of tumour necrosis factor activates c-Jun N-terminal kinase and mediates inflammation-induced hypersensitivity.

BackgroundMounting evidence points to individual contributions of tumour necrosis factor-alpha (TNF) and the c-Jun N-terminal kinase (JNK) pathway to the induction and maintenance of various pain states. Here we explore the role of spinal TNF and JNK in carrageenan-induced hypersensitivity. As links between TNF and JNK have been demonstrated in vitro, we investigated if TNF regulates spinal JNK activity in vivo.MethodsTNF levels in lumbar cerebrospinal fluid (CSF) were measured by enzyme-linked immunosorbent assay, spinal TNF gene expression by real-time polymerase chain reaction and TNF protein expression, JNK and c-Jun phosphorylation by western blotting. The role of spinal TNF and JNK in inflammation-induced mechanical and thermal hypersensitivity was assessed by injecting the TNF inhibitor etanercept and the JNK inhibitors SP600125 and JIP-1 intrathecally (i.t.). TNF-mediated regulation of JNK activity was examined by assessing the effect of i.t. etanercept on inflammation-induced spinal JNK activity.ResultsTNF levels were increased in CSF and spinal cord following carrageenan-induced inflammation. While JNK phosphorylation followed the same temporal pattern as TNF, c-jun was only activated at later time points. Intrathecal injection of TNF and JNK inhibitors attenuated carrageenan-induced mechanical and thermal hypersensitivity. TNF stimulation induced JNK phosphorylation in cultured spinal astrocytes and blocking the spinal actions of TNF in vivo by i.t. injection of etanercept reduced inflammation-induced spinal JNK activity.ConclusionsHere we show that spinal JNK activity is dependent on TNF and that both TNF and the JNK signalling pathways modulate pain-like behaviour induced by peripheral inflammation

Differential Requirement for c-Jun N-terminal Kinase 1 in Lung Inflammation and Host Defense

The c-Jun N-terminal kinase (JNK) - 1 pathway has been implicated in the cellular response to stress in many tissues and models. JNK1 is known to play a role in a variety of signaling cascades, including those involved in lung disease pathogenesis. Recently, a role for JNK1 signaling in immune cell function has emerged. The goal of the present study was to determine the role of JNK1 in host defense against both bacterial and viral pneumonia, as well as the impact of JNK1 signaling on IL-17 mediated immunity. Wild type (WT) and JNK1 −/− mice were challenged with Escherichia coli, Staphylococcus aureus, or Influenza A. In addition, WT and JNK1 −/− mice and epithelial cells were stimulated with IL-17A. The impact of JNK1 deletion on pathogen clearance, inflammation, and histopathology was assessed. JNK1 was required for clearance of E. coli, inflammatory cell recruitment, and cytokine production. Interestingly, JNK1 deletion had only a small impact on the host response to S. aureus. JNK1 −/− mice had decreased Influenza A burden in viral pneumonia, yet displayed worsened morbidity. Finally, JNK1 was required for IL-17A mediated induction of inflammatory cytokines and antimicrobial peptides both in epithelial cells and the lung. These data identify JNK1 as an important signaling molecule in host defense and demonstrate a pathogen specific role in disease. Manipulation of the JNK1 pathway may represent a novel therapeutic target in pneumonia

Interplay between FGFR2b-induced autophagy and phagocytosis: role of PLCγ-mediated signalling

Signalling of the epithelial splicing variant of the fibroblast growth factor receptor 2 (FGFR2b) induces both autophagy and phagocytosis in human keratinocytes. Here, we investigated, in the cell model of HaCaT keratinocytes, whether the two processes might be related and the possible involvement of PLCγ signalling. Using fluorescence and electron microscopy, we demonstrated that the FGFR2b-induced phagocytosis and autophagy involve converging autophagosomal and phagosomal compartments. Moreover, the forced expression of FGFR2b signalling mutants and the use of specific inhibitors of FGFR2b substrates showed that the receptor-triggered autophagy requires PLCγ signalling, which in turn activates JNK1 via PKCδ. Finally, we found that in primary human keratinocytes derived from light or dark pigmented skin and expressing different levels of FGFR2b, the rate of phagocytosis and autophagy and the convergence of the two intracellular pathways are dependent on the level of receptor expression, suggesting that FGFR2b signalling would control in vivo the number of melanosomes in keratinocytes, determining skin pigmentation

JNK1 activation predicts the prognostic outcome of the human hepatocellular carcinoma

BACKGROUND: Hepatocellular carcinoma (HCC) is one of the most common cancers worldwide with an extremely poor prognosis. The classification of HCC based on the molecular signature is not well-established. RESULTS: In the present study, we reported HCC signature genes based on the JNK1 activation status in 31 HCC specimens relative to the matched distal noncancerous liver tissue from 31 patients. The HCCs with high JNK1 (H-JNK1) and low JNK1 (L-JNK1) were sub-grouped. Two different signature gene sets for both H-JNK1 and L-JNK1 HCC were identified through gene expression profiling. A striking overlap of signature genes was observed between the H-JNK1 HCC and the hepatoblastoma or hepatoblastoma-type HCC. Many established biomarkers for hepatic progenitor cells were over-expressed in H-JNK1 HCC, including AFP, TACSTD1, KRT19, KRT7, THY1, and PROM1. In addition, the majority of the most up-regulated genes were those associated with metastasis and earlier recurrence, whereas the genes for normal liver function were substantially down-regulated in H-JNK1 HCC tissue. A Kaplan-Meier plot demonstrated that the survival of the patients with H-JNK1 HCC was severely impaired. CONCLUSION: Accordingly, we believe that the H-JNK1 HCC may originate from hepatic progenitor cells and is associated with poorer prognosis. The status of JNK1 activation in HCC tissue, thus, might be a new biomarker for HCC prognosis and therapeutic targeting

Tyrosine dephosphorylation of H2AX modulates apoptosis and survival decisions.

Life and death fate decisions allow cells to avoid massive apoptotic death in response to genotoxic stress. Although the regulatory mechanisms and signalling pathways controlling DNA repair and apoptosis are well characterized, the precise molecular strategies that determine the ultimate choice of DNA repair and survival or apoptotic cell death remain incompletely understood. Here we report that a protein tyrosine phosphatase, EYA, is involved in promoting efficient DNA repair rather than apoptosis in response to genotoxic stress in mammalian embryonic kidney cells by executing a damage-signal-dependent dephosphorylation of an H2AX carboxy-terminal tyrosine phosphate (Y142). This post-translational modification determines the relative recruitment of either DNA repair or pro-apoptotic factors to the tail of serine phosphorylated histone H2AX (gamma-H2AX) and allows it to function as an active determinant of repair/survival versus apoptotic responses to DNA damage, revealing an additional phosphorylation-dependent mechanism that modulates survival/apoptotic decisions during mammalian organogenesis