Subcellular distribution and phosphorylation of the nuclear localization signal binding protein, NBP60

We previously purified a nuclear localization signal binding protein, NBP60, from rat liver (1993, J. Biochem. 113, 308-313). In this study, the subcellular localization of NBP60 was examined using anti-NBP60. Most NBP60 was found to be localized in the nuclear envelope fraction of rat liver obtained on cell fractionation followed by immunoblotting. Staining of the nuclei of cultured cells by the antibody was observed on immunofluorescence microscopy. NBP60 was widely detected in rat nuclear fractions prepared from other tissues and also in nuclei of cultured cells derived from other species. It was shown by immunoelectron microscopy that most NBP60 is present in the nuclear envelope and at least some of that is present on nuclear pore complexes. Although NBP60 was localized in the nuclear envelope in interphase cells, it diffused into the cytoplasm in the mitotic phase. The purified NBP60 was highly phosphorylated by a cdc2 mitotic kinase, whereas nuclear pore proteins p144, p62, p60, and p54 were not phosphorylated by the kinase directly. NBP60 was also phosphorylated by protein kinase A, calmodulin-dependent protein kinase II, and casein kinase II. The phosphorylation of NBP60 by cdc2 kinase and/or the other kinases may be related to the change in the protein's location during the mitotic phase.     

Sensitivity degradation in polarization diversity receivers forlightwave systems

The effect of weighting circuit unidealities on the sensitivity degradation of diversity receivers is examined for synchronous and nonsynchronous detectors. A novel model is developed in which the weighting ratio for each polarization diversity receiver branch can be calculated using practical values. For synchronous detectors, it is shown analytically that the sensitivity degradation is relatively independent of the weighting ratio deviation from the optimum value. The same is also true for nonsynchronous detectors. Results of experiments on a 600 Mb/s single-filter frequency-shift keying (FSK) lightwave system, a 400 Mb/s continuous-phase FSK (CPFSK) lightwave system, and an amplitude-shift keying (ASK) synchronous detection network simulated with a 600 Mb/s baseband system generally support the theoretical results     

System reliability of suspension bridges

Provisions for the design of existing suspension bridges often rely on a deterministic basis. Consequently, the reliability of these bridges cannot be assessed if current provisions are applied. In order to develop cost-effective design and maintenance strategies for suspension bridges a system reliability-based approach has to be used. This is accomplished by a probabilistic finite element geometrically nonlinear analysis approach. This study forms part of an investigation into the system reliability evaluation of geometrically nonlinear large span bridges recently undertaken at the University of Colorado. A brief review of reliability analysis of geometrically nonlinear elastic structures allows for the determination of its relevance to the assessment of suspension bridges. A probabilistic finite element geometrically nonlinear elastic code is used for system reliability evaluation of suspension structures. The allowable stress design procedures used by the Honshu Shikoku Bridge Authority for the design of suspension bridges are presented along with their application to the design of an existing bridge. This bridge is studied from a system reliability viewpoint to evaluate its reliability under different loading and damage scenarios. Such information calls attention to the fact that the reliability of cables, hanger ropes and girders are very different. Therefore, optimal maintenance decisions for suspension bridges designed according to allowable stress method are not consistent with those based on equal component reliability values.     

A case with dicentric translocation between chromosome 9 and 18: confirmation by fluorescent in situ hybridization on metaphase spread

A female child with dicentric translocation between chromosome 9 and chromosome 18 presented non-specific minor anomalies with laryngomalacia. Chromosomal analyses were performed by the G-banding method and a fluorescence in situ hybridization (FISH) technique with a specific probe for the centromeric region of chromosome 18 and the painting probe for the chromosomes 9 and 18. Her full karyotype was confirmed as 45,XX,tdic(9;18)(p24;p11). This is the first case of dicentric translocation between chromosomes 9 and 18. The FISH technique is an important tool in chromosome diagnostics.     

HDR codec with concatenated multilevel codes and multiple-symboldifferential detection for satellite application

The performance of the multilevel code based high data rate (HDR) codec is evaluated with multiple-symbol differential detection over satellite channels. It is shown that multiple-symbol observation effectively fills the gap between conventional differentially coherent detection and ideal coherent detection with differential encoding. Being especially suitable for HDR application where carrier acquisition and tracking is extremely difficult, the codec reliably supports broadband ISDN transmission over a 72 MHz satellite transponder     

Polarization diversity detection performance of 2.5-Gb/s CPFSKregenerators intended for field use

2.5-Gb/s continuous-phase frequency-shift keying (CPFSK) regenerators are studied in laboratory and field experiments. A polarization diversity receiver is constructed with newly designed automatic gain and automatic frequency controllers. It achieves a sensitivity fluctuation of about 0.5 dB against all input signal polarization states. The automatic gain control is used in the intermediate-frequency circuits of the receiver and stably operates over 15 dB of input signal power variation. The automatic frequency control (AFC) is sufficiently stable to withstand a 30-kHz input signal polarization variation between two orthogonal linear polarization modes. AFC stability is maintained even if the environmental temperature of the laser diode modules fluctuates from 15 to 35 C. The polarization fluctuation in both cases results in only 0.1-dB degradation in receiver sensitivity     

Establishment and characterization of a new erythropoietin-dependent acute myeloid leukemia cell line, AS-E2

We have established an erythropoietin-dependent human leukemia cell line, AS-E2, from a patient with acute myeloid leukemia. These cells have many characteristics of late erythroid progenitor cells, they are positive for CD36, Glycophorin A, and CD71 but negative for CD41, and positive for benzidine and PAS staining. These cells express GATA-1 and have low affinity erythropoietin (EPO) receptor on their surface. Interestingly, AS-E2 cells are strictly dependent on EPO for their growth and survival; other cytokines including GM-CSF, stem cell factor, or IL-3 cannot support the growth of this cell line. These features are similar to late erythroid lineage cells, like normal BFU-E or CFU-E, and we have demonstrated that EPO stimulation induces the tyrosine phosphorylation of several proteins in AS-E2 cells including the EPO receptor and JAK2 kinase. This new cell line is a useful reagent to study biological and molecular events during the late stages of erythropoiesis, and to understand transforming events in human erythroid cells.     

Monoclonal antibodies against pig ovarian follicular granulosa cells induce apoptotic cell death in cultured granulosa cells

Two monoclonal antibodies capable of inducing granulosa cell apoptosis were produced against granulosa cells prepared from antral follicles of pig ovaries. The healthy follicles, 4-5 mm in diameter, were dissected from the ovaries of gilts, and then granulosa cells were isolated. BALB/c female mice were immunized with the isolated granulosa cells. Antibodies against the granulosa cells were detected by immunofluorescent staining using frozen ovarian sections. The isolated spleen cells prepared from immunized mice producing antibodies against the granulosa cells were fused with Sp2/O-Ag 14 mouse myeloma cells by standard hybridization techniques. Two hybridoma clones, PFG-1 and PFG-2, which produced specific IgM antibodies against granulosa cells were selected. Western blotting analysis revealed that PFG-1 and PFG-2 antibodies specifically recognized cell-membrane proteins with molecular weights of 55 and 70 kD and isoelectric points of 5.9 and 5.4, respectively. The monoclonal antibodies immunohistochemically reacted with granulosa cells of healthy follicles. When the isolated granulosa cells prepared from healthy follicles were cultured in medium containing 0.1 or 10 micrograms/m/PFG-1 or PFG-2 antibodies, respectively, the cells underwent apoptosis as determined by nuclear morphology, DNA electrophoresis and flow cytometric analysis. In conclusion, these two monoclonal antibodies against granulosa cells have cell-killing activity in cultured granulosa cells.     

Increased sensitivity of gastric cancer cells to natural killer and lymphokine-activated killer cells by antisense suppression of N-acetylgalactosaminyltransferase

UDP-N-acetylgalactosamine:polypeptide N-acetylgalactosaminyltransferases (GalNAc transferases) catalyze the initial reaction in O-linked (mucin type) oligosaccharide biosynthesis. To attempt to inhibit the synthesis of O-glycan, we transfected antisense cDNA of GalNAc transferase type 1 (GalNAc-T1) into a human gastric cancer cell line, JRST. A decreased expression of GalNAc-T1 at the level of both mRNA and protein was observed in the resultant transfectants. They demonstrated a significantly increased sensitivity to NK and lymphokine-activated killer cells in vitro compared with parental cells and mock transfectants. Although there was no significant difference in in vitro cell proliferation among parental cells, mock transfectants, or antisense transfectants, the in vivo growth rate of antisense transfectants using SCID mice was clearly lower than that of parental cells and mock transfectants. Furthermore, the treatment of mice with anti-asialo-G(M1) Ab abolished this growth-inhibitory effect of antisense transfection. From these results, we conclude that antisense suppression of GalNAc-T1 could increase the sensitivity of tumor cells to NK and lymphokine-activated killer cells, suggesting that this strategy may be of use as a new immunogene therapy.