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Biomedical subjects

Ze'ev Ronai

Publications and source records attributed to Ze'ev Ronai.

14 recordsLinked to original sources
PubMed2003

ERK and PI3K negatively regulate STAT-transcriptional activities in human melanoma cells: implications towards sensitization to apoptosis.

Signal transducers and activators of transcription (STAT) proteins nuclear translocation and transcriptional activity are regulated by diverse protein kinases in response to extracellular stimuli by cytokines, growth factors and stress. Using two melanoma-derived cell lines that exhibit marked differences in basal activities of MAPKs and PI3K-AKT, we studied changes both in STAT activities and in their sensitization to apoptosis. Activating mutations of B-RAF (T1796A) and impaired expression of PTEN are detected in LU1205, but not in FEMX melanoma cells, and are reflected in high basal levels of expression and activities of MAPKs and PI3K-AKT. Treatment with either PD98059 (PD) or LY294002 (LY), the pharmacological inhibitors of MEK-ERK and PI3K, respectively, markedly increased GAS-Luc activity in LU1205, but not in FEMX cells. Tyrosine phosphorylation of STAT3/5 and of JAK2 also increased upon treatment of LU1205 cells with either PD or LY, suggesting that constitutive active MAPK and PI3K signals inhibit tyrosine phosphorylation of JAK/STATs. Treatment of FEMX and LU1205 with PD sensitized the cells to apoptosis, albeit by TNFalpha and TRAIL death cascades, respectively, indicating that additional yet distinct targets are affected by each signaling pathway. Indeed, the combination of LY and PD treatment synergistically increased the apoptosis of LU1205 and FEMX cells. Overall, whereas PI3K and MAPK downregulate JAK-STAT signaling, additional targets are affected by these kinases and sensitizes melanoma to apoptosis via distinct death cascades.

Apoptosis↗
PubMed2003

Death receptors and melanoma resistance to apoptosis.

Impaired ability to undergo programmed cell death in response to a wide range of external stimuli acquires melanomas a selective advantage for progression and metastasis as well as their notorious resistance to therapy. Better understanding of mechanisms that govern apoptosis has enabled identification of diverse routes by which melanomas manage to escape stimuli of apoptosis. Changes at genomic, transcriptional and post-translational levels of G-proteins and protein kinases (Ras, B-Raf) and their transcription factor effectors (c-Jun, ATF2, Stat3 and NF-kappaB) affects TNF, Fas and TRAIL receptors, which play important roles in acquiring melanoma's resistance to apoptosis. Here, we summarize our current understanding of changes that alters the regulation of death receptors during melanoma development.

Activating Transcription Factor 2↗
PubMed2003

The cytoprotective aminothiol WR1065 activates p53 through a non-genotoxic signaling pathway involving c-Jun N-terminal kinase.

WR1065 is an aminothiol with selective cytoprotective effects in normal cells compared with cancer cells. In a previous study (North, S., El-Ghissassi, F., Pluquet, O., Verhaegh, G., and Hainaut, P. (2000) Oncogene 19, 1206-1214), we have shown that WR1065 activates wild-type p53 in cultured cells. Here we show that WR1065 induces p53 to accumulate through escape from proteasome-dependent degradation. This accumulation is not prevented by inhibitors of phosphatidylinositol 3-kinases and is not accompanied by phosphorylation of Ser-15, -20, or -37, which are common targets of the kinases activated in response to DNA damage. Furthermore, WR1065 activates the JNK (c-Jun N-terminal kinase), decreases complex formation between p53 and inactive JNK, and phosphorylates p53 at Thr-81, a known site of phosphorylation by JNK. A dominant negative form of JNK (JNK-APF) reduces by 50% the activation of p53 by WR1065. Thus, WR1065 activates p53 through a JNK-dependent signaling pathway. This pathway may prove useful for pharmacological modulation of p53 activity through non-genotoxic mechanisms.

3T3 Cells↗
PubMed2003

FAP-1 association with Fas (Apo-1) inhibits Fas expression on the cell surface.

As revealed by intracellular pools of nonactive Fas (Apo-1), export of Fas to the cell surface is often impaired in human tumors, thereby inactivating Fas ligand-mediated apoptosis. Here, we demonstrate that association with Fas-associated phosphatase 1 (FAP-1) attenuates Fas export to the cell surface. Forced expression of FAP-1 reduces cell surface Fas levels and increases the intracellular pool of Fas within the cytoskeleton network. Conversely, expression of dominant-negative forms of FAP-1, or inhibition of FAP-1 expression by short interfering RNA, efficiently up-regulates surface expression of Fas. Inhibition of Fas surface expression by FAP-1 depends on its association with the C terminus of Fas. Mutation within amino acid 275 results in decreased association with FAP-1 and greater export of Fas to the cell surface in melanomas, normal fibroblasts, or Fas null cells. Identifying the role of FAP-1 in binding to, and consequently inhibition of, Fas export to the cell surface provides novel insight into the mechanism underlying the regulation of Fas trafficking, which is commonly impaired in advanced tumors with FAP-1 overexpression.

Animals↗
PubMed2003

RNF5, a RING finger protein that regulates cell motility by targeting paxillin ubiquitination and altered localization.

RNF5 is a RING finger protein found to be important in the growth and development of Caenorhabditis elegans. The search for RNF5-associated proteins via a yeast two-hybrid screen identified a LIM-containing protein in C. elegans which shows homology with human paxillin. Here we demonstrate that the human homologue of RNF5 associates with the amino-terminal domain of paxillin, resulting in its ubiquitination. RNF5 requires intact RING and C-terminal domains to mediate paxillin ubiquitination. Whereas RNF5 mediates efficient ubiquitination of paxillin in vivo, protein extracts were required for in vitro ubiquitination, suggesting that additional modifications and/or an associated E3 ligase assist RNF5 targeting of paxillin ubiquitination. Mutant Ubc13 efficiently inhibits RNF5 ubiquitination, suggesting that RNF5 generates polychain ubiquitin of the K63 topology. Expression of RNF5 increases the cytoplasmic distribution of paxillin while decreasing its localization within focal adhesions, where it is primarily seen under normal growth. Concomitantly, RNF5 expression results in inhibition of cell motility. Via targeting of paxillin ubiquitination, which alters its localization, RNF5 emerges as a novel regulator of cell motility.

3T3 Cells↗
PubMed2002

Stress-induced decrease in TRAF2 stability is mediated by Siah2.

TRAF2 serves as a central regulator of the cellular response to stress and cytokines through the regulation of key stress-signaling cascades. Here we demonstrate that wild-type, but not RING mutant, Siah2 targets TRAF2 for ubiquitylation and degradation in vitro. Siah2 mediates equally efficient ubiquitylation of RING mutant TRAF2. In vivo, Siah2 primarily targets TRAF2 for degradation under stress conditions. Tumor necrosis factor-alpha (TNF-alpha) and actinomycin D treatment results in accelerated TRAF2 degradation in wild-type mouse embryo fibroblasts (MEFs), as compared with Siah2(-/-) cells. Similarly, TRAF2 half-life is prolonged in Siah2(-/-) compared with wild-type MEFs subjected to stress stimuli. Siah2 efficiently decreases TNF-alpha-dependent induction of JNK activity and transcriptional activation of NF-kappaB. Apoptosis induced by TNF-alpha and actinomycin D treatment is increased upon expression of Siah2, or attenuated upon expression of TRAF2 or RING mutant Siah2. Identifying Siah2 as a regulator of TRAF2 stability reveals its role in the regulation of TRAF2 signaling following exposure to stress.

Animals↗
PubMed2002

The prolyl isomerase Pin1 reveals a mechanism to control p53 functions after genotoxic insults.

The tumour suppressor p53 is important in the cell decision to either arrest cell cycle progression or induce apoptosis in response to a variety of stimuli. p53 post-translational modifications and association with other proteins have been implicated in the regulation of its stability and transcriptional activities. Here we report that, on DNA damage, p53 interacts with Pin1, a peptidyl-prolyl isomerase, which regulates the function of many proteins involved in cell cycle control and apoptosis. The interaction is strictly dependent on p53 phosphorylation, and requires Ser 33, Thr 81 and Ser 315. On binding, Pin1 generates conformational changes in p53, enhancing its transactivation activity. Stabilization of p53 is impaired in UV-treated Pin1(-/-) cells owing to its inability to efficiently dissociate from Mdm2. As a consequence, a reduced p53-dependent response was detected in Pin1(-/-) cells, and this correlates with a diminished transcriptional activation of some p53-regulated genes. Our results suggest that, following stress-induced phosphorylation, p53 needs to form a complex with Pin1 and to undergo a conformational change to fulfil its biological roles.

Amino Acid Motifs↗
PubMed2002

Regulation of c-myc stability by selective stress conditions and by MEKK1 requires aa 127-189 of c-myc.

C-myc availability is central for its ability to serve as a regulator of cell growth and death. Here we study the regulation of c-myc protein stability and identify domains of c-myc that are important for its stabilization in response to stress kinases activated following selective stress conditions. UV-irradiation elicited an increase in c-myc protein levels, which could be attenuated by inhibitors of stress kinases but also by actinomycin D-inhibitor of transcription. Inhibition of protein synthesis results in a noticeable decrease in c-myc levels, further pointing to the short half-life of the protein. However, in combination with tumor necrosis factor-alpha (TNF-alpha), cycloheximide efficiently increases steady-state levels of c-myc, suggesting that selective stress conditions are required to increase c-myc protein stability. Expression of MEKK1, an upstream regulator of protein kinases that has been implicated in mediating the response to diverse stress conditions, also results in an efficient increase in the half-life of c-myc protein. To map c-myc domains that are responsive to stress kinases, we monitored changes in the level of c-myc deletion mutants following MEKK1 expression. Of the seven c-myc deletion mutants analysed, the domain spanning amino acids 127-189 was found to be required for MEKK1-dependent increase in c-myc stability. In all, the present study identifies a novel domain that is important for the regulation of c-myc stability by stress kinases in response to selective stress conditions.

Antineoplastic Agents↗
PubMed2002

p53-Mdm2--the affair that never ends.

The p53-Mdm2 paradigm represents the best-studied relationship between a tumor suppressor gene which functions as a transcription factor and an oncogene, which functions primarily as an E3 protein ligase. The intimate relationship between these two partners has expanded to include almost every cellular biological system - from development, to growth control and programmed cell death. The affair between Mdm2 and p53 is closely controlled by a complex array of post-translational modifications, which in turn dictates the stability and activity of p53 and Mdm2. Functional diversity depends on the association with a large subset of partner proteins, which dictates the type of activity and corresponding selectivity. Here we summarize the current understanding of post-translational modifications and their effect on conformation-based functional relationship between Mdm2 and p53, as it pertains to their diverse cellular biological functions.

Humans↗
PubMed2002

An ATF2-derived peptide sensitizes melanomas to apoptosis and inhibits their growth and metastasis.

Melanomas are among the aggressive tumor types because of their notorious resistance to treatment and their high capacity to metastasize. ATF2 is among transcription factors implicated in the progression of melanoma and its resistance to treatment. Here we demonstrate that the expression of a peptide spanning amino acids 50-100 of ATF2 (ATF2(50-100)) reduces ATF2 transcriptional activities while increasing the expression and activity of c-Jun. Altering the balance of Jun/ATF2 transcriptional activities sensitized melanoma cells to apoptosis, an effect that could be attenuated by inhibiting c-Jun. Inhibition of ATF2 via RNA interference likewise increased c-Jun expression and primed melanoma cells to undergo apoptosis. Growth and metastasis of SW1 and B16F10 mouse melanomas were inhibited by ATF2(50-100) to varying degrees up to a complete regression, depending on the mode (inducible, constitutive, or adenoviral delivery) of its expression.

Activating Transcription Factor 2↗
PubMed2001

Regulation of Fas expression by STAT3 and c-Jun is mediated by phosphatidylinositol 3-kinase-AKT signaling.

Cooperation between STAT3 and c-Jun results in suppression of Fas Receptor (FasR) transcription, which is often seen in advanced human tumors. To identify requirements for STAT3-Jun cooperation, we elucidated the role of protein kinases that affect both transcription factors. The phosphatidylinositol 3-kinase (PI3K)-AKT signaling pathway was found capable of down-regulating both STAT3- and c-Jun-dependent transcription, resulting in derepression of FasR transcription. Conversely, inhibition of PI3K-AKT signaling via the specific pharmacological inhibitor LY294002 up-regulated AP1/Jun- and STAT-dependent transcriptional activities, resulting in suppression of the FasR promoter activities and decreased FasR surface expression. PI3K-AKT's ability to affect FasR transcription was not observed in c-jun null fibroblasts, suggesting that c-Jun is required for PI3K/AKT-mediated regulation of FasR transcription. Interestingly, the dominant negative form of Rac1 (RacN17) was also efficient in relieving FasR expression, suggesting that the increase in FasR expression following AKT stimuli could be mediated via AKT ability to elicit suppression of Rac1, which in turn decreases JNK activities and c-Jun phosphorylation. Overall, our findings demonstrate that through its negative effects on both c-Jun and STAT3, the PI3K-AKT pathway disrupts cooperation between c-Jun and STAT3, which is required for silencing the FasR promoter, resulting in increased expression of surface FasR and concomitant sensitization to FasL-mediated programmed cell death.

Apoptosis↗
PubMed1999

GST function in drug and stress response.

Glutathione (GSH) is the major non-protein thiol in cells and is a co-factor for many types of conjugation reactions involving glutathione S-transferases (GST). Because of the defined role of GST in drug metabolism, elevated expression of GSTpi either as a tumor marker or in drug resistance has automatically been linked to detoxification. Similarly, GSH has mostly been viewed as a passive cellular supply of nucleophilic sulfur. This mini-review highlights recent reports which advocate a novel, non-catalytic role for GSTpi as an integral determinant of cellular pathways of stress response, proliferation and apoptosis. Copyright 1999 Harcourt Publishers Ltd.

Journal Article↗
PubMedDate not supplied

SUMO in cancer--wrestlers wanted.

SUMO (small ubiquitin-related modifier) represents a class of ubiquitin-like proteins that is conjugated, like ubiquitin, by a set of enzymes to cellular regulatory proteins, including oncogenes and tumor suppressor genes, that play key roles in the control of cell growth, differentiation and apoptosis. SUMO conjugation affects substrates' subcellular localization and stability as well as transcriptional activities. Given the substrates involved, protein SUMOylation would be expected to be important in the course of tumorigenesis and, accordingly, altered in human cancer. Although evidence to support this notion is still scarce, this review summarizes the current knowledge of protein sumoylation and highlights the challenges to be addressed in the context of human cancer.

Cell Cycle↗