The E1B 19K protein associates with lamins in vivo and its proper localization is required for inhibition of apoptosis

Expression of the E1B 19K protein is required to inhibit apoptosis induced by E1A during adenovirus infection and transformation. E1B 19K is homologous to Bcl-2 in function and the two proteins also share limited amino acid sequence homology. Consequently, the E1B 19K and Bcl-2 proteins bind to and inhibit the cellular death-inducing proteins Bax, Bak and Nbk/Bik. Both E1B 19K and Bcl-2 localize to membranes of the nucleus and the endoplasmic reticulum. In addition to membrane association, and unlike Bcl-2, the E1B 19K protein is found associated with intermediate filament proteins in the cytoplasm and the nuclear lamina and copurifies with the lamins both during infection and transformation. While a membrane targeting domain at the C-terminus of Bcl-2 ensures its proper localization, the mechanism by which the E1B 19K protein localizes is unknown. Not surprisingly, lamin A fragments were cloned from a yeast two-hybrid screen for E1B 19K-interacting proteins. The interaction was demonstrated in yeast and mammalian cells in vivo and in vitro and was unique and specific to E1B 19K, with no interaction evident between Bcl-2 and lamin A. Mutants of lamin A/C which localized inappropriately in the cytoplasm or nucleus but retained E1B 19K binding, interfered with the nuclear envelope and cytoplasmic membrane targeting of the E1B 19K protein. Improper localization impaired the ability of the E1B 19K protein to inhibit apoptosis. Thus, proper localization of the E1B 19K protein is required for its function and the interaction of the E1B 19K protein with lamin A/C may represent a means for nuclear envelope localization.     

E1B 19,000-molecular-weight protein interacts with and inhibits CED-4-dependent, FLICE-mediated apoptosis

Genetic studies of the nematode Caenorhabditis elegans (C. elegans) have identified several important components of the cell death pathway, most notably CED-3, CED-4, and CED-9. CED-4 directly interacts with the Bcl-2 homologue CED-9 (or the mammalian Bcl-2 family member Bcl-xL) and the caspase CED-3 (or the mammalian caspases ICE and FLICE). This trimolecular complex of CED-4, CED-3, and CED-9 is functional in that CED-9 inhibits CED-4 from activating CED-3 and thereby inhibits apoptosis in heterologous systems. The E1B 19,000-molecular weight protein (E1B 19K) is a potent apoptosis inhibitor and the adenovirus homologue of Bcl-2-related apoptosis inhibitors. Since E1B 19K and Bcl-xL have functional similarity, we determined if E1B 19K interacts with CED-4 and regulates CED-4-dependent caspase activation. Binding analysis indicated that E1B 19K interacts with CED-4 in a Saccharomyces cerevisiae two-hybrid assay, in vitro, and in mammalian cell lysates. The subcellular localization pattern of CED-4 was dramatically changed by E1B 19K, supporting the theory of a functional interaction between CED-4 and E1B 19K. Whereas expression of CED-4 alone could not induce cell death, coexpression of CED-4 and FLICE augmented cell death induction by FLICE, which was blocked by expression of E1B 19K. Even though E1B 19K did not prevent FLICE-induced apoptosis, it did inhibit CED-4-dependent, FLICE-mediated apoptosis, which suggested that CED-4 was required for E1B 19K to block FLICE activation. Thus, E1B 19K functions through interacting with CED-4, and presumably a mammalian homologue of CED-4, to inhibit caspase activation and apoptosis.     

E1B 55K sequesters WT1 along with p53 within a cytoplasmic body in adenovirus-transformed kidney cells

WT1 encodes a tumor suppressor that is expressed in cells of the developing kidney and is inactivated in Wilms tumor, a pediatric kidney cancer. The adenovirus E1B 55K gene product contributes to the transformation of primary baby rat kidney (BRK) cells by binding and inactivating the product of the p53 tumor suppressor. We have previously demonstrated that WT1 and p53 are present within a protein complex in vivo. We now show that WT1 is physically associated with E1B 55K in adenovirus-transformed cells, an interaction that is mediated by the first two zinc fingers of WT1. Immunodepletion of p53 abrogates the coimmunoprecipitation of E1B 55K and WT1, consistent with the presence of a trimeric protein complex containing these three proteins. In the presence of E1B 55K, WT1 which is normally localized in the nucleus, is retained within a very high molecular weight complex and sequestered in the characteristic perinuclear cytoplasmic body that contains E1B 55K and p53. Expression of E1B 55K in osteosarcoma cells that undergo apoptosis following expression of WT1 inhibits WT1-mediated cell death. We conclude that E1B 55K may target WT1 along with p53, resulting in the functional inactivation of both tumor suppressor gene products by this viral oncoprotein.     

硼替佐米作用不同时间对K562耐药细胞株核因子-κ B途径的影响

目的 观察硼替佐米(商品名:万珂,PS-341)对柔红霉素(DNR)诱导的K562耐药细胞株(K562/DNR)核因子-κ B(NF-κ B)、抑制蛋白κ B(I κ B)及P-糖蛋白(P-gp)表达的影响,探讨PS-341逆转耐药的分子机制.方法 以100μg/ml DNR单用或联合应用4μg/L PS-341分别作用于K562/DNR 12、24及36 h,检测不同时间点各组NF-κ B、Iκ B及P-gp表达情况,同时测定NF-κ B p65活性,检测各组细胞凋亡率.结果 Western blot结果显示:与阴性对照组相比,DNR可诱导NF-κ B表达上调及活性增强、I κ B表达下调、P-gp表达上凋;加用PS-341可显著抑制DNR诱导的NF-κ B及P-gp表达,使I κ B表达增加.加用PS-341后,NF-κ B活性12 h为(15.3±1.87)%[DNR组为(23.8±2.27)%],24 h为(10.2±1.69)%[DNR组为(25.4±1.98)%],36 h为(6.08±2.53)%[DNR组为(26.9±2.58)%],与相应单用DNR组相比均有明显下降,差异有统计学意义(P值均＜0.05).DNR与PS-341联用后,细胞凋亡率12 h为(35.23±5.15)%[DNR组为(15.56±4.12)%],24 h为(40.26±6.89)%[DNR组为(17.25±2.89)%],36 h为(43.58±7.69)%[DNR组为(22.47±4.58)%],与DNR组相比,细胞凋亡率均明显增加,差异具有统计学意义(P值均＜0.05).上述作用呈时间依赖性.结论 PS-341可减少K562/DNR细胞NF-κ B的活化,降低P-gp表达,逆转细胞耐药,促进细胞凋亡.     

Fex-C1-x/Si颗粒膜的巨磁电阻效应

利用激光脉冲沉积（PLD）方法制备了沉积于硅基片上的掺杂过渡金属的非晶碳膜结构Fex-C1-x/Si。Fex-C1-x/Si的磁电阻（MR）可正可负，随温度而变化。当温度T〈258K时，Fe0．011-C0.989／Si的MR为负值；当258K〈T〈340K时，该材料的MR为正值，在室温磁场为1T时，该材料的正MR可以大于20％。且在不同的温度范围中，该材料的MR和外加磁场的依存关系呈现出不同的特点：在T=280和300K时，当磁场小于1T时，MR随磁场的增加而快速增加，之后随磁场的继续增加MR增加开始变得缓慢；在T=350K时，MR近似以磁场的B^1.5。的规律变化；而在T=30K时，MR为负值且其大小随磁场的增加而减小。利用双通道模型对该MR效应进行了初步解释。     

E1B 19K inhibits Fas-mediated apoptosis through FADD-dependent sequestration of FLICE

E1B 19K, the adenovirus Bcl-2 homologue, is a potent inhibitor of apoptosis induced by various stimuli including Fas and tumor necrosis factor-alpha. Fas and TNFR-1 belong to a family of cytokine-activated receptors that share key components in their signaling pathways, Fas-associating protein with death domain (FADD) and FADD-like interleukin-1beta-converting enzyme (FLICE), to induce an apoptotic response. We demonstrate here that E1B 19K and Bcl-xL are able to inhibit apoptosis induced by FADD, but not FLICE. Surprisingly, apoptosis was abrogated by E1B 19K and Bcl-xL when FADD and FLICE were coexpressed. Immunofluorescence studies demonstrated that FADD expression produced large insoluble death effector filaments that may represent oligomerized FADD. E1B 19K expression disrupted FADD filament formation causing FADD and FLICE to relocalize to membrane and cytoskeletal structures where E1B 19K is normally localized. E1B 19K, however, does not detectably bind to FADD, nor does it inhibit FADD and FLICE from being recruited to the death-inducing signaling complex (DISC) when Fas is stimulated. Thus, E1B 19K may inhibit Fas-mediated cell death downstream of FADD recruitment of FLICE but upstream of FLICE activation by disrupting FADD oligomerization and sequestering an essential component of the DISC.     

Functional analysis of the amino- and carboxyl-termini of streptokinase

1. The classical ATP sensitive K+ (K(ATP)) channels are composed of a sulphonylurea receptor (SUR) and an inward rectifying K+ channel subunit (BIR/Kir6.2). They are the targets of vasorelaxant agents called K+ channel openers, such as pinacidil and nicorandil. 2. In order to examine the tissue selectivity of pinacidil and nicorandil, in vitro, we compared the effects of these agents on cardiac type (SUR2A/Kir6.2) and vascular smooth muscle type (SUR2B/Kir6.2) of the K(ATP) channels heterologously expressed in HEK293T cells, a human embryonic kidney cell line, by using the patch-clamp method. 3. In the cell-attached recordings (145 mM K+ in the pipette), pinacidil and nicorandil activated a weakly inwardly-rectifying, glibenclamide-sensitive 80 pS K+ channel in both the transfected cells. 4. In the whole-cell configuration, pinacidil showed a similar potency in activating the SUR2B/Kir6.2 and SUR2A/Kir6.2 channels (EC50 of approximately 2 and approximately 10 microM, respectively). On the other hand, nicorandil activated the SUR2B/Kir6.2 channel > 100 times more potently than the SUR2A/Kir6.2 (EC50 of approximately 10 microM and > 500 microM, respectively). 5. Thus, nicorandil, but not pinacidil, preferentially activates the K(ATP) channels containing SUR2B. Because SUR2A and SUR2B are diverse only in 42 amino acids at their C-terminal ends, it is strongly suggested that this short part of SUR2B may play a critical role in the action of nicorandil on the vascular type classical K(ATP) channel.     

Major histocompatibility complex class II expression and parathyroid autoantibodies in primary hyperparathyroidism

We evaluated whether kinins exert a protective action against the development of two-kidney, one clip (2K1C) hypertension, a model characterized by an activated renin-angiotensin system in the ischemic kidney and increased expression of the bradykinin (BK) B2 receptor in the contralateral kidney. BK B2-receptor knockout (B2-/-), wild-type (B2+/+), and heterozygous (B2+/-) mice underwent clipping of the left renal artery, with the other kidney remaining untouched. Basal systolic blood pressure (SBP, via tail-cuff plethysmography) was higher in B2-/- mice than in B2+/- or B2+/+ mice (121+/-2 versus 113+/-2 and 109+/-1 mm Hg; P<0.05 for both comparisons). SBP did not change from basal values after sham operation, but it increased in mice that underwent clipping. The increase in SBP was greater in 2K1C B2-/- mice than in B2+/- or B2+/+ mice (28+/-2 versus 14+/-2 and 14+/-2 mm Hg, respectively, at 2 weeks; P<0.05 for both comparisons). Blockade of the BK B2 receptor by Icatibant enhanced the pressure response to clipping in B2+/+ mice (29+/-2 mm Hg at 2 weeks). Intra-arterial mean blood pressure (MBP) was higher in 2K1C than in respective sham-operated mice, with the MBP difference being higher in B2-/- mice (32 and 38 mm Hg, at 2 and 4 weeks, respectively), and higher in B2+/+ mice given Icatibant (30 and 32 mm Hg) than in B2+/+ mice without Icatibant (17 and 18 mm Hg). At 4 weeks, acute injection of an angiotensin type 1 receptor antagonist normalized the MBP of 2K1C hypertensive mice. A tachycardic response was observed 1 week after clipping in B2-/- and B2+/- mice, but this effect was delayed in B2+/+ mice. However, the HR response to clipping in B2+/+ mice was enhanced by Icatibant. Within each strain, heart weight to body weight ratio was greater in 2K1C hypertensive mice than in sham-operated control animals (B2-/-: 5.7+/-0.1 versus 5.2+/-0.1; B2+/+: 5.1+/-0.1 versus 4.5+/-0.1; P<0.01 for both comparisons). The clipped kidney weight to nonclipped kidney weight ratio was consistently reduced in mice with 2K1C hypertension. Our results indicate that kinins acting on the BK B2 receptor exert a protective action against excessive blood pressure elevation during early phases of 2K1C hypertension.     

Temperature dependence of the Qy resonance Raman spectra of bacteriochlorophylls, the primary electron donor, and bacteriopheophytins in the bacterial photosynthetic reaction center

Qy-excited resonance Raman spectra of the accessory bacteriochlorophylls (B), the bacteriopheophytins (H), and the primary electron donor (P) in the bacterial photosynthetic reaction center (RC) of Rhodobacter sphaeroides have been obtained at 95 and 278 K. Frequency and intensity differences are observed in the low-frequency region of the P vibrational spectrum when the sample is cooled from 278 to 95 K. The B and H spectra exhibit minimal changes of frequencies and relative intensities as a function of temperature. The mode patterns in the Raman spectra of B and H differ very little from Raman spectra of the chromophores in vitro. The Raman scattering cross sections of B and H are 6-7 times larger than those for analogous modes of P at 278 K. The cross sections of B and of H are 3-4 times larger at 95 K than at 278 K, while the cross sections of P are approximately constant with temperature. The temperature dependence of the Raman cross sections for B and H suggests that pure dephasing arising from coupling to low-frequency solvent/protein modes is important in the damping of their excited states. The weak Raman cross sections of the special pair suggest that the excited state of P is damped by very rapid (<30 fs) electronic relaxation processes. These resonance Raman spectra provide information for developing multimode vibronic models of the excited-state structure and dynamics of the chromophores in the RC.     

A comparison of injectable anaesthetic regimens in mules

Three combinations of injectable anaesthetic agents were compared in nine adult mules. The combinations were xylazine/ketamine (X/K), xylazine/butorphanol/ketamine (X/B/K), and xylazine/tiletamine-zolazepam (X/T). Measured variables were heart rate, respiratory rate, systolic blood pressure, arterial blood pH, PCO2 and PO2, recumbency time and number of attempts to stand. Quality of induction and recovery, muscle relaxation and response to stimulus were evaluated subjectively. Recumbency time was significantly (P < 0.05) longer with X/B/K and X/T than with X/K. Mules required significantly more attempts to stand under the influence of X/T than X/K or X/B/K. No statistically significant (P < 0.05) differences in heart rate, respiratory rate, blood pressure or arterial pH, PCO2 and PO2 were detected between groups.     

Mass production of magnetocaloric LaFeMnSiB alloys with hydrogenation

LaFe_(11.39)Mn_(0.35)Si_(1.26)B_(0.1)Hxalloys were prepared by hydrogenation.Samples were annealed at 1343Kfor30-90 hto form the NaZn13 phase.La-rich andα-Fe secondary phases were also detected.Saturated hydrogenation at 553 Kand 0.15 MPa of H_2 pressure for 5hwas employed to improve the Curie temperature of the alloys to 279 K.The maximum magnetic entropy change,relative cooling power,and adiabatic temperature change of LaFe_(11.39)Mn_(0.35)Si_(1.26)B_(0.1)H_x annealed at 1343 Kfor 90hafter hydrogen absorption are 6.38J/(kg·K)(magnetic changesμ0ΔH =1.65T),100.1J/kg(μ0ΔH =1.65T),and 2.2 K(μ0ΔH =1.48T),respectively.Although the maximum magnetic entropy change of the LaFe_(11.39)Mn_(0.35)Si_(1.26)B_(0.1)H_x alloys is lower than those of similar alloys with high purity raw materials,the relative cooling power is nearly the same.The effect of impurities of the raw materials used was also discussed.It is assumed that the impurity of 0.2wt.% Al is responsible for the reduced entropy change of the resulted alloys.The LaFe_(11.39)Mn_(0.35)Si_(1.26)B_(0.1)H_x alloys prepared by this method could be a low cost alternative material for room temperature magnetic cooling applications.     

Associative training results in persistent reductions in a calcium-activated potassium current in Hermissenda Type B photoreceptors.

Type B photoreceptors from the eyes of associatively trained Hermissenda had significantly greater light responses and net input resistances than Type B cells from control Ss on retention days following conditioning. Differences were still apparent when the contribution of a rapidly inactivating K+ current (IA) to the photoresponse was minimized by either depolarization-induced inactivation or block by 4-aminopyridine ions. Training-produced differences in Type B cell light responses were abolished by treatments that reduced the contribution of a calcium-activated K+ current (IK-Ca) to the light response. Under voltage-clamp conditions in which IK-Ca was isolated from other components of outward current, it was selectively reduced by associative training. The associative reduction of IK-Ca could not be attributed to training-produced reductions in the voltage-dependent calcium current of Type B cells. ICa was enhanced by associative training. A slower component of light-induced inward current was greater for Type B cells from associatively trained Ss. Previous results suggest that this slower component may be due to light-induced reduction of a calcium-activated K+ current. Training-produced reductions in a calcium-activated K+ may be responsible for enhanced photoresponses of Type B cells from conditioned Ss. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Differential generation of class I H-2D- versus H-2K-restricted cytotoxicity against a demyelinating virus following central nervous system infection

Despite the fact that both H-2K and D molecules are up-regulated in the central nervous system (CNS) following Theiler's murine encephalomyelitis virus (TMEV) infection, resistance in this virus model of multiple sclerosis maps exclusively to D. To address this paradox, we examined the ability of the K and D molecules to present viral antigens to cytotoxic T lymphocytes (CTL). Whereas no virus-specific CTL were detected in the CNS of susceptible B10.Q and B10.S mice 7 days post-infection, D-restricted CTL were identified readily in the CNS of resistant B10 animals. There was no evidence of K-restricted CTL in the CNS of B10 mice at day 7 post-infection. The presence of both K- and D-restricted virus-specific CTL in the spleen of immunized B10 mice demonstrates that the exclusive use of D molecules by CTL in the CNS of mice 7 days post-infection is not due to the inability of the K molecules to present viral peptides to lymphocytes. We conclude that the prominent role of the D locus in determining resistance or susceptibility to TMEV-induced demyelination is determined by factors governing the regulation of the immune response, and not by the presence or absence of CTL precursors capable of recognizing viral peptides presented by the K and D antigen-presenting molecules, or by differences in the ability of the K and D molecules to present viral peptides.     

Kinetics and Mechanism of Low-Grade Manganese Ore Reduction by Natural Gas

Metallurgical and Materials Transactions B - The calcination and the reduction behaviors of a low-grade manganese ore by methane was studied at 973 K to 1273 K by several techniques. The onset...     

Aluminum Deoxidation Equilibrium of Fe-Ni Alloy at 1773 K and 1873 K

Metallurgical and Materials Transactions B - The aluminum deoxidation equilibrium in molten Fe-36 mass pct Ni and Fe-46 mass pct Ni alloys was experimentally determined at 1773 K and 1873 K to...     

A Comparison of Static and Cyclic Long-Term Oxidation of Two Nb-Cr-Mo-Si-B Alloys

Nb-25Cr-20Mo-15Si-10B (compositions in at pct) and Nb-25Cr-20Mo-15Si-15B alloys were exposed to air for a maximum period of 2 weeks under static and cyclic conditions to determine oxidation response. Oxidation was carried out at temperatures of 973 K, 1173 K, 1373 K, and 1573 K (700 °C, 900 °C, 1100 °C, and 1300 °C). Results of long-term cyclic oxidation show an increase in oxidation resistance with an increase in boron content. Pesting has been observed at 973 K (700 °C) in the 10B alloy in cyclic and static modes of oxidation. Comparative analysis of oxide formation is done by the weight gain per unit surface area method. The alloys and their oxides are characterized by X-ray diffraction, scanning electron microscopy, and X-ray mapping.     

Relationship of Coke Reactivity and Critical Coke Properties

Metallurgical and Materials Transactions B - The gasification reactions of five metallurgical cokes were studied using a thermogravimetric analyzer at 1273&nbsp;K and 1673&nbsp;K in...