Activation of the c-Jun N-terminal kinase (JNK) signaling pathway is essential during PBOX-6-induced apoptosis in chronic myelogenous leukemia (CML) cells

The mitogen-activated protein (MAP) kinase family is activated in response to a wide variety of external stress signals such as UV irradiation, heat shock, and many chemotherapeutic drugs and leads to the induction of apoptosis. A novel series of pyrrolo-1,5-benzoxazepines have been shown to potently induce apoptosis in chronic myelogenous leukemia (CML) cells, which are resistant to many chemotherapeutic agents. In this study we have delineated part of the mechanism by which a representative compound known as PBOX-6 induces apoptosis. We have investigated whether PBOX-6 induces activation of MAP kinase signaling pathways in CML cells. Treatment of K562 cells with PBOX-6 resulted in the transient activation of two JNK isoforms, JNK1 and JNK2. In contrast, PBOX-6 did not activate the extracellular signal-regulated kinase (ERK) or p38. Apoptosis was found to occur independently of the small GTPases Ras, Rac, and Cdc42 but involved phosphorylation of the JNK substrates, c-Jun and ATF-2. Pretreatment of K562 cells with the JNK inhibitor, dicoumarol, abolished PBOX-6-induced phosphorylation of c-Jun and ATF-2 and inhibited the induced apoptosis, suggesting that JNK activation is an essential component of the apoptotic pathway induced by PBOX-6. Consistent with this finding, transfection of K562 cells with the JNK scaffold protein, JIP-1, inhibited JNK activity and apoptosis induced by PBOX-6. JIP-1 specifically scaffolds JNK, MKK7, and members of the mixed-lineage kinase (MLK) family, implicating these kinases upstream of JNK in the apoptotic pathway induced by PBOX-6 in K562 cells.

Cloning and Characterization of Two Potent Kunitz Type Protease Inhibitors from Echinococcus granulosus

The tapeworm Echinococcus granulosus is responsible for cystic echinococcosis (CE), a cosmopolitan disease which imposes a significant burden on the health and economy of affected communities. Little is known about the molecular mechanisms whereby E. granulosus is able to survive in the hostile mammalian host environment, avoiding attack by host enzymes and evading immune responses, but protease inhibitors released by the parasite are likely implicated. We identified two nucleotide sequences corresponding to secreted single domain Kunitz type protease inhibitors (EgKIs) in the E. granulosus genome, and their cDNAs were cloned, bacterially expressed and purified. EgKI-1 is highly expressed in the oncosphere (egg) stage and is a potent chymotrypsin and neutrophil elastase inhibitor that binds calcium and reduced neutrophil infiltration in a local inflammation model. EgKI-2 is highly expressed in adult worms and is a potent inhibitor of trypsin. As powerful inhibitors of mammalian intestinal proteases, the EgKIs may play a pivotal protective role in preventing proteolytic enzyme attack thereby ensuring survival of E. granulosus within its mammalian hosts. EgKI-1 may also be involved in the oncosphere in host immune evasion by inhibiting neutrophil elastase and cathepsin G once this stage is exposed to the mammalian blood system. In light of their key roles in protecting E. granulosus from host enzymatic attack, the EgKI proteins represent potential intervention targets to control CE. This is important as new public health measures against CE are required, given the inefficiencies of available drugs and the current difficulties in its treatment and control. In addition, being a small sized highly potent serine protease inhibitor, and an inhibitor of neutrophil chemotaxis, EgKI-1 may have clinical potential as a novel anti-inflammatory therapeutic.

Identification of different specificity requirements between SGK1 and PKB alpha

NDRG1 is phosphorylated by SGK1 (but not PKB) in vivo at three residues each contained within three nonapeptide repeats. Here, we demonstrate that this nonapeptide, like the NDRG1 protein, is phosphorylated by SGK1, but not by PKBalpha or RSK1 in vitro. The inability of PKBa and RSK1 to phosphorylate the nonapeptide was traced to residues n + 1, n + 2 and n - 4 (where n is the phosphorylation site). Changing them from Ser, Glu and Ser to Phe, Ala and Pro, respectively, transformed the nonapeptide into an excellent substrate for PKBalpha and RSK1. Our results identify a specific substrate for SGK1 and may facilitate detection of additional physiological substrates for this enzyme. (C) 2005 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

Ultralow Silicon Substrate Noise Crosstalk Using Metal Faraday Cages in an SOI Technology

In mixed signal integrated circuits noise from the digital circuitry can upset the sensitive analogue circuitry. The Faraday cage structure reported here is based on the unique ground plane SOI technology developed some of the authors. The suppression of crosstalk achieved is an order of magnitude greater than that previously published for frequencies up to 10 GHz. The significance of the technology will be even greater as the operating frequency is increased. This collaborative EPSRC project was judge as tending to outstanding.

2D-drop model applied to the calculation of step formation energies on a (111) substrate

A model is presented for obtaining the step formation energy for metallic islands on (1 1 1) surfaces from Monte Carlo simulations. This model is applied to homo (Cu/Cu(1 1 1), Ag/Ag(1 1 1)) and heteroepitaxy (Ag/Pt(1 1 1)) systems. The embedded atom method is used to represent the interaction between the particles of the system, but any other type of potential could be used as well. The formulation can also be employed to consider the case of other single crystal surfaces, since the higher barriers for atom motion on other surfaces are not a hindrance for the simulation scheme proposed.