No abstract

Mitogen-activated protein (MAP) kinase cascades are activated in response to various extracellular stimuli, including growth factors and environmental stresses. A MAP kinase kinase kinase (MAPKKK), termed ASK1, was identified that activated two different subgroups of MAP kinase kinases (MAPKK), SEK1 (or MKK4) and MKK3/MAPKK6 (or MKK6), which in turn activated stress-activated protein kinase (SAPK, also known as JNK; c-Jun amino-terminal kinase) and p38 subgroups of MAP kinases, respectively. Overexpression of ASK1 induced apoptotic cell death, and ASK1 was activated in cells treated with tumor necrosis factor-alpha (TNF-alpha). Moreover, TNF-alpha-induced apoptosis was inhibited by a catalytically inactive form of ASK1. ASK1 may be a key element in the mechanism of stress- and cytokine-induced apoptosis.

The bacterial macrolide rapamycin is an efficacious anticancer agent against solid tumors. In a hypoxic environment, the increase in mass of solid tumors is dependent on the recruitment of mitogens and nutrients. When nutrient concentrations change, particularly those of essential amino acids, the mammalian Target of Rapamycin (mTOR) functions in regulatory pathways that control ribosome biogenesis and cell growth. In bacteria, ribosome biogenesis is independently regulated by amino acids and adenosine triphosphate (ATP). Here we demonstrate that the mTOR pathway is influenced by the intracellular concentration of ATP, independent of the abundance of amino acids, and that mTOR itself is an ATP sensor.

Tumor necrosis factor (TNF)-induced activation of the c-jun N-terminal kinase (JNK, also known as SAPK; stress-activated protein kinase) requires TNF receptor-associated factor 2 (TRAF2). The apoptosis signal-regulating kinase 1 (ASK1) is activated by TNF and stimulates JNK activation. Here we show that ASK1 interacts with members of the TRAF family and is activated by TRAF2, TRAF5, and TRAF6 overexpression. A truncated derivative of TRAF2, which inhibits JNK activation by TNF, blocks TNF-induced ASK1 activation. A catalytically inactive mutant of ASK1 is a dominant-negative inhibitor of TNF- and TRAF2-induced JNK activation. In untransfected mammalian cells, ASK1 rapidly associates with TRAF2 in a TNF-dependent manner. Thus, ASK1 is a mediator of TRAF2-induced JNK activation.

Apoptosis signal-regulating kinase 1 (ASK1) is a MAPKKK family member which activates c-Jun N-terminal kinase (JNK) and p38. In non-stressed cells, ASK1 exists as an inactive complex with the reduced form of thioredoxin. Oxidative stress such as hydrogen peroxide (H2O2) disrupts the ASK1-thioredoxin complex by oxidization of thioredoxin and thereby activates ASK1. The precise mechanism by which ASK1 is activated after its release from thioredoxin is unknown. Here we show that phosphorylation of Thr845 at the activation loop is essential for ASK1 to be activated by H2O2. ASK1 appears to form a silent homo-oligomer through its C-terminal coiled-coil region in non-stressed cells. Following H2O2 treatment, pre-existing ASK1 oligomer undergoes conformational change and creates a new interface within an oligomer, which ultimately leads to trans-autophosphorylation of Thr845. Thus, direct interaction via the coiled-coil region is required for self-scaffolding but not sufficient for activation of ASK1. Importantly, Thr845 of ASK1 can also be trans-phosphorylated by an unidentified Thr845 kinase in response to H2O2 treatment. We propose that this potential Thr845 kinase may be an ignition kinase that triggers Thr845 phosphorylation in oligomerized and activation-competent forms of ASK1.

Epithelial-mesenchymal transition (EMT), a crucial event in cancer progression and embryonic development, is induced by transforming growth factor (TGF)-␤ in mouse mammary NMuMG epithelial cells. Id proteins have previously been reported to inhibit major features of TGF-␤-induced EMT.In this study, we show that expression of the ␦EF1 family proteins, ␦EF1 (ZEB1) and SIP1, is gradually increased by TGF-␤ with expression profiles reciprocal to that of E-cadherin. SIP1 and ␦EF1 each dramatically down-regulated the transcription of E-cadherin in NMuMG cells through direct binding to the E-cadherin promoter. Silencing of the expression of both SIP1 and ␦EF1, but not either alone, completely abolished TGF-␤-induced E-cadherin repression. However, expression of mesenchymal markers, including fibronectin, N-cadherin, and vimentin, was not affected by knockdown of SIP1 and ␦EF1. TGF-␤-induced the expression of Ets1, which in turn activated ␦EF1 promoter activity. Moreover, up-regulation of SIP1 and ␦EF1 expression by TGF-␤ was suppressed by knockdown of Ets1 expression. In addition, Id2 suppressed the TGF-␤-and Ets1-induced up-regulation of ␦EF1. Taken together, these findings suggest that the ␦EF1 family proteins, SIP1 and ␦EF1, are necessary, but not sufficient, for TGF-␤-induced EMT and that Ets1 induced by TGF-␤ may function as an upstream transcriptional regulator of SIP1 and ␦EF1. INTRODUCTIONTransforming growth factor (TGF)-␤, a prototypical member of the TGF-␤ family, regulates a broad range of cellular responses, including cell proliferation, differentiation, adhesion, migration, and apoptosis (Bierie and Moses, 2006). TGF-␤ and related factors exhibit their pleiotropic effects through binding to transmembrane serine-threonine kinase receptors type I (T␤R-I) and type II (T␤R-II). On ligand-induced heteromeric complex formation between T␤R-I and T␤R-II, T␤R-I is phosphorylated and activated by T␤R-II kinase and mediates specific intracellular signaling through phosphorylation of receptor-regulated Smads (R-Smads). Phosphorylated R-Smads interact with coSmad (Smad4) and translocate into the nucleus, where they regulate transcription of target genes in cooperation with various transcription factors and transcriptional coactivators or corepressors (Miyazawa et al., 2002;Miyazono et al., 2003;Shi and Massague, 2003).TGF-␤ has potent antiproliferative effects on a wide variety of cells, including epithelial cells, endothelial cells, and hematopoietic cells, although under certain conditions it promotes the proliferation of mesenchymal cells, including fibroblasts, chondrocytes, and osteoblasts. TGF-␤ also induces the deposition of extracellular matrix proteins. In early stages of tumorigenesis, TGF-␤ inhibits the growth of epithelial cells, and insensitivity to this growth-inhibitory effect is associated with progression of tumors Derynck et al., 2001). Transgenic mice expressing a dominant-negative T␤R-II in epidermis exhibit malignant conversion of epithelial cells and promotion of tumor formation (Gorska et al., 20...

The epithelial-mesenchymal transition (EMT) provides an outstanding example of cellular plasticity during embryonic development and cancer progression. During EMT in embryonic development, epithelial cells lose all vestiges of their epithelial origin and acquire a fully mesenchymal phenotype, known as complete EMT, which is typically characterized by a so-called cadherin switch. Conversely, during EMT in cancer progression, cancer cells that originate from epithelial cells exhibit both mesenchymal and epithelial characteristics, that is the hybrid E/M phenotype in a process known as partial EMT. Partial EMT in cancer cells is thought to enhance their invasive properties, generate circulating tumour cells and cancer stem cells, and promote resistance to anti-cancer drugs. These phenotypic changes are regulated by extracellular matrix components, exosomes and soluble factors, which regulate several transcription factors known as EMT transcription factors. In this review, I summarize our current understanding of the EMT program during cancer progression.

Apoptosis signal-regulating kinase 1 (ASK1) is a MAP kinase kinase kinase (MAPKKK) that activates the JNK and p38 MAP kinase cascades and is activated in response to oxidative stress such as hydrogen peroxide (H 2 O 2 ). A yeast two-hybrid screening identi®ed a serine/threonine protein phosphatase 5 (PP5) as a binding partner of ASK1. PP5 directly dephosphorylated an essential phospho-threonine residue within the kinase domain of ASK1 and thereby inactivated ASK1 activity in vitro and in vivo. The interaction between PP5 and ASK1 was induced by H 2 O 2 treatment and was followed by the decrease in ASK1 activity. PP5 inhibited not only H 2 O 2 -induced sustained activation of ASK1 but also ASK1-dependent apoptosis. Thus, PP5 appears to act as a physiological inhibitor of ASK1±JNK/p38 pathways by negative feedback.