List of contributors Volume 153

Authors Index

List of contributors Volume 153     

Induction of interferon in hybrid clones: Vero 153--mouse myeloma and Vero 153--human lymphocyte cells

A Vero cell line (Vero 153) resistant to 8-azaguanine and unresponsive to viral induction of interferon was isolated. This primate (African green) cell line was fused with mouse myeloma (S194/5) and normal human lymphocytes from peripheral blood. All Vero 153--mouse hybrids, 8 primary and 12 secondary clones, produced virus-induced mouse but not primate interferon. This occurred even in cultures where greater than 90% of primate chromosomes were retained. Similarly 7 primary and 3 secondary Vero 153--human clones synthesized virus-induced interferon. This could be neutralized by anti-human fibroblast (beta) but not by anti-human leukocyte (alpha) interferon antisera. The unresponsive nature of Vero 153 cells to interferon induction by viruses was not changed by the presence of interferon producing genomes from other cells. However, despite the inability to produce interferon, the Vero cell was able to play a role in the determination of the type of interferon made in the hybrid cell.     

Curcumin-induced GADD153 upregulation: modulation by glutathione

As we reported previously, GADD153 is upregulated in colon cancer cells exposed to curcumin. In the present study, we ascertained the involvement of glutathione and certain sulfhydryl enzymes associated with signal transduction in mediating the effect of curcumin on GADD153. Curcumin-induced GADD153 gene upregulation was attenuated by reduced glutathione (GSH) or N-acetylcysteine (NAC) and potentiated by the glutathione synthesis inhibitor, L-buthionine-(S,R)-sulfoximine (BSO). Additionally, GSH and NAC decreased the intracellular content of curcumin. Conversely, curcumin decreased intracellular glutathione and also increased the formation of reactive oxygen species (ROS) in cells, but either GSH or NAC prevented both of these effects of curcumin. In affecting the thiol redox status, curcumin caused activation of certain sulfhydryl enzymes involved in signal transduction linked to GADD153 expression. Curcumin increased the expression of the phosphorylated forms of PTK, PDK1, and PKC-delta, which was attenuated by either GSH or NAC and potentiated by BSO. Furthermore, selective inhibitors of PI3K and PKC-delta attenuated curcumin-induced GADD153 upregulation. Collectively, these findings suggest that a regulatory thiol redox-sensitive signaling cascade exists in the molecular pathway leading to induction of GADD153 expression as caused by curcumin.     

Estimated human absorbed dose of a new 153Sm bone seeking agent based on biodistribution data in mice: Comparison with 153Sm-EDTMP

PurposeThe main goal in radiotherapy is to deliver the absorbed dose within the target organs in highest possible amount, while the absorbed dose of the other organs, especially the critical organs, should be kept as low as possible. In this work, the absorbed dose to human organs for a new ¹⁵³Sm bone-seeking agent was investigated.Methods¹⁵³Sm-(4-{[(bis(phosphonomethyl))carbamoyl]methyl}-7,10-bis(carboxymethyl)-1,4,7,10-tetraazacyclododec-1-yl) acetic acid (¹⁵³Sm-BPAMD) complex was successfully prepared. The biodistribution of the complex was investigated in male Syrian mice up to 48 h post injection. The human absorbed dose of the complex was estimated based on the biodistribution data of the mice by radiation absorbed dose assessment resource (RADAR) method. The target to non-target absorbed dose ratios for ¹⁵³Sm-BPAMD were compared with these ratios for ¹⁵³Sm-EDTMP.ResultsThe highest absorbed dose for ¹⁵³Sm-BPAMD was observed in bone surface with 5.828 mGy/MBq. The dose ratios of the bone surface to the red marrow and to the total body for ¹⁵³Sm-BPAMD were 5.3 and 20.0, respectively, while these ratios for ¹⁵³Sm-EDTMP were 4.4 and 18.3, respectively. This means, for a given dose to the bone surface as the target organ, the red marrow (as the main critical organ) and the total body would receive lesser absorbed dose in the case of ¹⁵³Sm-BPAMD.ConclusionsGenerally, the human absorbed dose estimation of ¹⁵³Sm-BPAMD indicated that all other tissues approximately received insignificant absorbed dose in comparison with bone surface and therefore can be regarded as a new potential agent for bone pain palliation therapy.     

Single molecule study of the intrinsically disordered FG-repeat nucleoporin 153

Nucleoporins (Nups), which are intrinsically disordered, form a selectivity filter inside the nuclear pore complex, taking a central role in the vital nucleocytoplasmic transport mechanism. These Nups display a complex and nonrandom amino-acid architecture of phenylalanine glycine (FG)-repeat clusters and intra-FG linkers. How such heterogeneous sequence composition relates to function and could give rise to a transport mechanism is still unclear. Here we describe a combined chemical biology and single-molecule fluorescence approach to study the large human Nup153 FG-domain. In order to obtain insights into the properties of this domain beyond the average behavior, we probed the end-to-end distance (R_E) of several ∼50-residues long FG-repeat clusters in the context of the whole protein domain. Despite the sequence heterogeneity of these FG-clusters, we detected a reoccurring and consistent compaction from a relaxed coil behavior under denaturing conditions (R_E/R_E,RC = 0.99 ± 0.15 with R_E,RC corresponding to ideal relaxed coil behavior) to a collapsed state under native conditions (R_E/R_E,RC = 0.79 ± 0.09). We then analyzed the properties of this protein on the supramolecular level, and determined that this human FG-domain was in fact able to form a hydrogel with physiological permeability barrier properties.     

Roles of CHOP/GADD153 in endoplasmic reticulum stress

Endoplasmic reticulum (ER) is the site of synthesis and folding of secretory proteins. Perturbations of ER homeostasis affect protein folding and cause ER stress. ER can sense the stress and respond to it through translational attenuation, upregulation of the genes for ER chaperones and related proteins, and degradation of unfolded proteins by a quality-control system. However, when the ER function is severely impaired, the organelle elicits apoptotic signals. ER stress has been implicated in a variety of common diseases such as diabetes, ischemia and neurodegenerative disorders. One of the components of the ER stress-mediated apoptosis pathway is C/EBP homologous protein (CHOP), also known as growth arrest- and DNA damage-inducible gene 153 (GADD153). Here, we summarize the current understanding of the roles of CHOP/GADD153 in ER stress-mediated apoptosis and in diseases including diabetes, brain ischemia and neurodegenerative disease.     

Molecular characterization of the Ran-binding zinc finger domain of Nup153

The nuclear pore complex is the gateway for selective traffic between the nucleus and cytoplasm. To learn how building blocks of the pore can create specific docking sites for transport receptors and regulatory factors, we have studied a zinc finger module present in multiple copies within the nuclear pores of higher eukaryotes. All four zinc fingers of human Nup153 were found to bind the small GTPase Ran with dissociation constants ranging between 5 and 40 mum. In addition a fragment of Nup153 encompassing the four tandem zinc fingers was found to bind Ran with similar affinity. NMR structural studies revealed that a representative Nup153 zinc finger adopts the same zinc ribbon structure as the previously characterized Npl4 NZF module. Ran binding was mediated by a three-amino acid motif (Leu(13)/Val(14)/Asn(25)) located within the two zinc coordination loops. Nup153 ZnFs bound GDP and GTP forms of Ran with similar affinities, indicating that this interaction is not influenced by a nucleotide-dependent conformational switch. Taken together, these studies elucidate the Ran-binding interface on Nup153 and, more broadly, provide insight into the versatility of this zinc finger binding module.     

Oxidative stress mislocalizes and retains transport factor importin-alpha and nucleoporins Nup153 and Nup88 in nuclei where they generate high molecular mass complexes

Nuclear trafficking of proteins requires the cooperation between soluble transport components and nucleoporins. As such, classical nuclear import depends on the dimeric carrier importin-alpha/beta1, and CAS, a member of the importin-beta family, which exports importin-alpha to the cytoplasm. Here we analyzed the effect of oxidative stress elicited by diethyl maleate (DEM) on classical nuclear transport. Under conditions that do not induce death in the majority of cells, DEM has little effect on the nucleocytoplasmic concentration gradient of Ran, but interferes with the nuclear accumulation of several reporter proteins. Moreover, DEM treatment alters the distribution of soluble transport factors and several nucleoporins in growing cells. We identified nuclear retention of importin-alpha, CAS as well as nucleoporins Nup153 and Nup88 as a mechanism that contributes to the nuclear concentration of these proteins. Both nucleoporins, but not CAS, associate with importin-alpha in the nuclei of growing cells and in vitro. Importin-alpha generates high molecular mass complexes in the nucleus that contain Nup153 and Nup88, whereas CAS was not detected. The formation of high molecular mass complexes containing importin-alpha, Nup153 and Nup88 is increased upon oxidant treatment, suggesting that complex formation contributes to the anchoring of importin-alpha in nuclei. Taken together, our studies link oxidative stress to the proper localization of soluble transport factors and nucleoporins and to changes in the interactions between these proteins.     

Genome sequence of the probiotic bacterium Sporolactobacillus vineae SL153T

The novel Sporolactobacillus vineae SL153(T) strain has excellent intestinal adherence and growth inhibitory effect on pathogenic microorganisms, including Vibrio genus microorganisms, and therefore can be effectively used for the prevention and treatment of disease caused by pathogenic microorganisms. Here, we first report the draft genome sequence of a novel species in the genus Sporolactobacillus.