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A decryption method based on CS is presented to retrieve the image from a double random phase digital holographic encrypted system. By applying CS to decrypt the incomplete hologram, signal can be ideally recovered.

Receptor activator of nuclear factor kappa B ligand (RANKL), a potent regulator of osteoclast formation and function, is expressed by osteoblasts and bone marrow stromal cells. However, the molecular mechanism underlying RANKL expression in osteoblast/stromal cells remains largely unclear. Here, we characterized the molecular mechanism controlling RANKL basal transcription in osteoblast/stromal cells. We cloned a 1,103-bp murine RANKL promoter (from -953 to +150, relative to the transcription start site). Using a series of deletion mutants of the 1,103-bp promoter, we identified a 100-bp region (-154 to -54) mediating RANKL basal transcription in both osteoblasts and bone marrow stromal cells. Electrophoretic mobility shift assay (EMSA) using five overlapping oligonucleotides (Probes 1-5) spanning the 100-bp region showed that Probes 1 and 2 specifically bound nuclear proteins with high affinity from both cell types. Computer analysis revealed that Probes 1 and 2 contain a putative Sp1-binding site. Supershift assays with Sp1 and Sp3 antibodies confirmed that the nuclear proteins binding to Probes 1 and 2 are Sp1 and Sp3. Functionally, the mutation of the Sp1/Sp3 site in Probe 1 profoundly reduced the basal promoter activity while the mutation of the one in Probe 2 resulted in moderate reduction in the basal promoter activity. Moreover, the mutation of both sites abrogated the RANKL basal promoter activity, indicating that Sp1 and Sp3 play a key role in the RANKL basal transcription in osteoblasts and bone marrow stromal cells.

Members of the tumour necrosis factor receptor family play a pivotal role in cell differentiation, function and apoptosis. However, signalling by many members of the family remains to be elucidated. In the present study, we developed a chimaeric receptor approach for studying signalling by receptors belonging to this family. The chimaeric receptor comprises the human Fas external domain linked to the transmembrane and cytoplasmic domains of a tumour necrosis factor receptor family member of interest. When the chimaera is expressed in mouse cells, the clustering of the chimaera induced by a human Fas-activating antibody activates the intracellular domain of the chimaera without affecting its endogenous counterpart. Since the antibody recognizes only human Fas, this approach can be used to dissect signalling by any tumour necrosis factor family member using any type of mouse cell including those endogenously expressing Fas. Moreover, we also showed that the chimaeric receptor approach can be used to study signalling at any stage of cell differentiation or function.

Rhythmic activation and repression of the frequency (frq) gene are essential for normal function of the Neurospora circadian clock. WHITE COLLAR (WC) complex, the positive element of the Neurospora circadian system, is responsible for stimulation of frq transcription. We report that a C2H2 finger domain-containing protein IEC-1 and its associated chromatin remodeling complex INO80 play important roles in normal Neurospora circadian clock function. In iec-1KO strains, circadian rhythms are abolished, and the frq transcript levels are increased compared to that of the wild-type strain. Similar results are observed in mutant strains of the INO80 subunits. Furthermore, ChIP data show that recruitment of the INO80 complex to the frq promoter is IEC-1-dependent. WC-mediated transcription of frq contributes to the rhythmic binding of the INO80 complex at the frq promoter. As demonstrated by ChIP analysis, the INO80 complex is required for the re-establishment of the dense chromatin environment at the frq promoter. In addition, WC-independent frq transcription is present in ino80 mutants. Altogether, our data indicate that the INO80 complex suppresses frq transcription by re-assembling the suppressive mechanisms at the frq promoter after transcription of frq., Author summary Circadian clocks organize inner physiology to anticipate changes in the external environment. These clocks are controlled by the oscillation of central clock proteins which form the central oscillator. Transcriptional regulation is a critical step in the regulation of the oscillation of these core proteins. In eukaryotes, chromatin remodeling is a common mechanism to regulate gene transcription by conquering or establishing nucleosomal barriers for the transcription machinery. Here, we showed that a C2H2 finger domain-containing protein IEC-1 and its associated chromatin remodeling complex INO80 are required for transcriptional repression of the core clock gene frq in the Neurospora circadian system. Moreover, the activator WHITE COLLAR (WC) complex is responsible for the transcriptional activation of frq; thus, considering the different timing of the transcriptional activation and suppression of frq, there must be a mechanism that coordinates the two opposite processes. We also demonstrated that the WC-mediated open state of the frq promoter facilitates the binding of INO80 to this region, which prepares for subsequent transcriptional suppression. Collectively, our data provide novel insights into the regulation of the frq gene and the circadian clock.

A computer generated holographic stereogram based on the wavefront recording plane (WRP) is presented. A WRP closed to the parallax image plane is introduced to record the complex amplitude in a small region for each point in the parallax image. By using three times of fast Fourier transform (FFT) to execute the Fresnel diffraction calculation between the WRP and the holographic stereogram plane, the object wave contributing to the hologram pattern can be achieved. The computation complexity of the proposed approach is dramatically reduced. The results show that the calculation time can be decreased by more than one order of magnitude.

The noise has a serious effect on the quality of the three-dimensional holographic images reconstructed. A new method is proposed to improve the Burch code and combine it with the three-step phase shifting method to remove the noise. The reconstructed images are with the high contrast and resolution. The three-dimensional reconstruction images are compared with the ones of median filtering and of 20-time intensity superposition method. The performance parameters of three methods are analyzed. The experimental results show that the zero-order light spot, conjugate image and speckle noise are suppressed effectively. The quality of the reconstructed image is noticeably improved.

Digital holography is a kind of coherent imaging method and inevitably affected by many factors in the process of recording. One of dominant problems is the speckle noise, which is essentially nonlinear multiplicative noise related to signals. So it is more difficult to remove than additive noise. Due to the noise pollution, the low resolution of image reconstructed is caused. A new solution for suppressing speckle noise in digital hologram is presented, which combines Canny filtering algorithm with wavelet threshold denoising algorithm. Canny filter is used to obtain the edge detail. Wavelet transformation performs denoising. In order to suppress speckle effectively and retain the image details as much as possible, Neyman-Pearson (N-P) criterion is introduced to estimate wavelet coefficient in every scale. An improved threshold function is proposed, whose curve is smoother. The reconstructed image is achieved by merging the denoised image with the edge details. Experimental results and performance parameters of the proposed algorithm are discussed and compared with other methods, which shows that the presented approach can not only effectively eliminate speckle noise, but also retain useful signals and edge information simultaneously.

Grain number, panicle seed setting rate, panicle number and grain weight are the most important components of rice grain yield. To date, several genes related to grain weight, grain number and panicle number have been described in rice. However, no genes regulating the panicle seed setting rate have been functionally characterized. Here we show that the domestication-related POLLEN TUBE BLOCKED 1 (PTB1), a RING-type E3 ubiquitin ligase, positively regulates the rice panicle seed setting rate by promoting pollen tube growth. The natural variation in expression of PTB1 which is affected by the promoter haplotype and the environmental temperature, correlates with the rice panicle seed setting rate. Our results support the hypothesis that PTB1 is an important maternal sporophytic factor of pollen tube growth and a key modulator of the rice panicle seed setting rate. This finding has implications for the improvement of rice yield.

A novel method for a straightforward computer-generated Fresnel hologram using multiple angular orthogonal projection images is presented. Without diffraction calculation and Fourier transforms calculation, the calculation complexity is simplified, and the accuracy is improved.

We report the construction of a poliovirus genome [pPVM-VPg(3F4A)] harboring a double mutation in VPg. This mutant, in which the tyrosine and the threonine at residues 3 and 4 of the VPg region were replaced by phenylalanine and alanine, respectively, is lethal, that is, all RNA synthesis was abolished and no revertants could be isolated. Using the properties of dicistronic polioviruses (with the general genotype PV 5′NTR-3AB-EMCV IRES-PV ORF-3′NTR), we have observed that the defect in RNA synthesis of the VPg(3F4A) mutant could be rescued by providing wild-type protein 3AB from the first open reading frame in trans. We conclude that the 3AB provided by the first cistron of the dicistronic construct was capable of "intragenomic complementation." Intragenomic complementation, however, was inefficient. Thus, the dicistronic RNAs were only quasi-infectious, and even first-passage viruses were found to have reverted to a functioning VPg in the polyprotein. This phenomenon underlines the role of polypeptide 3AB in multiple functions of viral proliferation. First-passage viruses, all of which expressed a small-plaque phenotype, had retained the original dicistronic genotype. At the fourth passage, however, all isolates were monocistronic, and they displayed complex genetic rearrangements revealing interesting information regarding IRES function.

The effect of deletion of each of the two authentic polyprotein translation initiation sites of foot-and-mouth disease virus on viral protein synthesis and replication was analyzed. Deletion of either the first or the second initiation site led to the expression of only one form of the leader protein, L or L', respectively, but in vitro processing of the viral polyprotein and cleavage of eIF-4 gamma were not affected by either deletion. Whereas RNA in which the first translation initiation site had been deleted led to the production of viruses in transfected BHK cells, deletion of the second translation initiation site abolished virus replication.

Background The ideal plant architecture (IPA) includes several important characteristics such as low tiller numbers, few or no unproductive tillers, more grains per panicle, and thick and sturdy stems. We have developed an indica restorer line 7302R that displays the IPA phenotype in terms of tiller number, grain number, and stem strength. However, its mechanism had to be clarified. Findings We performed re-sequencing and genome-wide variation analysis of 7302R using the Solexa sequencing technology. With the genomic sequence of the indica cultivar 9311 as reference, 307 627 SNPs, 57 372 InDels, and 3 096 SVs were identified in the 7302R genome. The 7302R-specific variations were investigated via the synteny analysis of all the SNPs of 7302R with those of the previous sequenced none-IPA-type lines IR24, MH63, and SH527. Moreover, we found 178 168 7302R-specific SNPs across the whole genome and 30 239 SNPs in the predicted mRNA regions, among which 8 517 were Non-syn CDS. In addition, 263 large-effect SNPs that were expected to affect the integrity of encoded proteins were identified from the 7302R-specific SNPs. SNPs of several important previously cloned rice genes were also identified by aligning the 7302R sequence with other sequence lines. Conclusions Our results provided several candidates account for the IPA phenotype of 7302R. These results therefore lay the groundwork for long-term efforts to uncover important genes and alleles for rice plant architecture construction, also offer useful data resources for future genetic and genomic studies in rice. Electronic supplementary material The online version of this article (doi:10.1186/1939-8433-5-18) contains supplementary material, which is available to authorized users.

Rice restorer lines play an important role in three-line hybrid rice production. Previous research based on molecular tagging has suggested that the restorer lines used widely today have narrow genetic backgrounds. However, patterns of genetic variation at a genome-wide scale in these restorer lines remain largely unknown. The present study performed re-sequencing and genome-wide variation analysis of three important representative restorer lines, namely, IR24, MH63, and SH527, using the Solexa sequencing technology. With the genomic sequence of the Indica cultivar 9311 as the reference, the following genetic features were identified: 267,383 single-nucleotide polymorphisms (SNPs), 52,847 insertion/deletion polymorphisms (InDels), and 3,286 structural variations (SVs) in the genome of IR24; 288,764 SNPs, 59,658 InDels, and 3,226 SVs in MH63; and 259,862 SNPs, 55,500 InDels, and 3,127 SVs in SH527. Variations between samples were also determined by comparative analysis of authentic collections of SNPs, InDels, and SVs, and were functionally annotated. Furthermore, variations in several important genes were also surveyed by alignment analysis in these lines. Our results suggest that genetic variations among these lines, although far lower than those reported in the landrace population, are greater than expected, indicating a complicated genetic basis for the phenotypic diversity of the restorer lines. Identification of genome-wide variation and pattern analysis among the restorer lines will facilitate future genetic studies and the molecular improvement of hybrid rice.

Morbidity and mortality of breast cancer patients are drastically increased when primary tumor cells are able to spread to distant sites and proliferate to become secondary lesions. Effective treatment of metastatic disease has been limited; therefore, an increased molecular understanding to identify biomarkers and therapeutic targets is needed. Breast cancer metastasis suppressor 1 (BRMS1) suppresses development of pulmonary metastases when expressed in a variety of cancer types, including metastatic mammary carcinoma. Little is known of Brms1 function throughout the initiation and progression of mammary carcinoma. The goal of this study was to investigate mechanisms of Brms1-mediated metastasis suppression in transgenic mice that express Brms1 using polyoma middle T oncogene-induced models. Brms1 expression did not significantly alter growth of the primary tumors. When expressed ubiquitously using a β-actin promoter, Brms1 suppressed pulmonary metastasis and promoted apoptosis of tumor cells located in the lungs but not in the mammary glands. Surprisingly, selective expression of Brms1 in the mammary gland using the MMTV promoter did not significantly block metastasis nor did it promote apoptosis in the mammary glands or lung, despite MMTV-induced expression within the lungs. These results strongly suggest that cell type-specific over-expression of Brms1 is important for Brms1-mediated metastasis suppression.

The bones are the most common sites of breast cancer metastasis. Upon arrival within the bone microenvironment, breast cancer cells coordinate the activities of stromal cells, resulting in an increase in osteoclast activity and bone matrix degradation. In late stages of bone metastasis, breast cancer cells induce apoptosis in osteoblasts, which further exacerbates bone loss. However, in early stages, breast cancer cells induce osteoblasts to secrete inflammatory cytokines purported to drive tumor progression. To more thoroughly evaluate the role of osteoblasts in early stages of breast cancer metastasis to the bones, we used green fluorescent protein-labeled human breast cancer cell lines MDA-MB-231 and MDA-MB-435, which both induce osteolysis after intra-femoral injection in athymic mice, and the murine pre-osteoblastic cell line MC3T3-E1 to modulate osteoblast populations at the sites of breast cancer metastasis. Breast cancer cells were injected directly into the femur with or without equal numbers of MC3T3-E1 cells. Tumors grew significantly larger when co-injected with breast cancer cells and MC3T3-E1 cells than injected with breast cancer cells alone. Osteolysis was induced in both groups, indicating that MC3T3-E1 cells did not block the ability of breast cancer cells to cause bone destruction. MC3T3-E1 cells promoted tumor growth out of the bone into the extraosseous stroma. These data suggest that breast cancer cells and osteoblasts communicate during early stages of bone metastasis and promote tumor growth.

Studies of the poliovirus genome-linked protein VPg have shown that this small viral protein is required for replication of virus-specific RNA (Q. Reuer, R. J. Kuhn, and E. Wimmer, J. Virol. 64:2967-2975, 1990). To understand the mechanism of RNA replication, we constructed a recombinant poliovirus genome encoding two tandemly arranged VPg coding sequences that were nearly identical in both nucleotide and amino acid sequence. Following transfection of this two-VPg-containing RNA into HeLa cells, we found a specific and selective deletion in the progeny virus genome. Sequence analysis of the recovered viral RNA indicated that the complete nucleotide sequence encoding the second (3C-proximal) VPg coding sequences was removed, restoring the authentic genome sequences in the poliovirus genome. Analysis of viral RNAs following transfection suggested that the deletion event occurred during genome replication. Deletion could have occurred via homologous recombination between two VPg sequences or via intramolecular deletion with loop-out of the template. In vitro translation of the two-VPg-containing transcript RNA indicated aberrant processing of the viral polyprotein. This result suggested that selection of the wild-type genotype in the transfected cells may occur at the level of viral protein synthesis.

The hypoxia-inducible factors have recently been identified as critical regulators of angiogenic-osteogenic coupling. Mice overexpressing HIFalpha subunits in osteoblasts produce abundant VEGF and develop extremely dense, highly vascularized long bones. In this study, we investigated the individual contributions of Hif-1alpha and Hif-2alpha in angiogenesis and osteogenesis by individually disrupting each Hifalpha gene in osteoblasts using the Cre-loxP method. Mice lacking Hif-1alpha demonstrated markedly decreased trabecular bone volume, reduced bone formation rate, and altered cortical bone architecture. By contrast, mice lacking Hif-2alpha had only a modest decrease in trabecular bone volume. Interestingly, long bone blood vessel development measured by angiography was decreased by a similar degree in both DeltaHif-1alpha and DeltaHif-2alpha mice suggesting a common role for these Hifalpha subunits in skeletal angiogenesis. In agreement with this idea, osteoblasts lacking either Hif-1alpha or Hif-2alpha had profound reductions in VEGF mRNA expression but only the loss of Hif-1alpha impaired osteoblast proliferation. These findings indicate that expression of both Hif-1alpha and Hif-2alpha by osteoblasts is required for long bone development. We propose that both Hif-1alpha and Hif-2alpha function through cell non-autonomous modes to promote vascularization of bone and that Hif-1alpha also promotes bone formation by exerting direct actions on the osteoblast.

The complete 5' and 3' non-coding regions of two attenuated South American foot-and-mouth disease virus (FMDV) vaccine strains, O1C-O/E and C3R-O/E, and their corresponding virulent parental strains, O1 Campos and C3 Resende, have been cloned from polymerase chain reaction-amplified primary cDNA. Differences observed in the derived nucleotide sequences between attenuated and virulent viruses seem not to affect regulatory signal structures, supporting the theory that genetic variations, primarily in the 3' halves of the viral genomes, contribute to the attenuation phenotype of the vaccine strains. In addition, this is the first report on the complete sequence of the 5' untranslated region of a C-type aphthovirus. Approximately 10% of the nucleotides differ from the corresponding known sequences of serotypes A or O.

Receptor activator of NF-kappa B ligand (RANKL) and its receptor activator of NF-kappa B (RANK) play pivotal roles in osteoclast differentiation and function. However, the structural determinants of the RANK that mediate osteoclast formation and function have not been definitively identified. To address this issue, we developed a chimeric receptor approach that permits a structure/function study of the RANK cytoplasmic domain in osteoclasts. Using this approach, we examined the role of six RANK putative tumor necrosis factor receptor-associated factor-binding motifs (PTM) (PTM1, ILLMT-REE(286-293); PTM2, PSQPS(349-353); PTM3, PFQEP(369-373); PTM4, VYVSQTSQE(537-545); PTM5, PVQEET(559-564); and PTM6, PVQEQG(604-609)) in osteoclast formation and function. Our data revealed that the RANK cytoplasmic domain possesses three functional motifs (PFQEP(369-373), PVQEET(559-564), and PVQEQG(604-609)) capable of mediating osteoclast formation and function. Moreover, we demonstrated that these motifs play distinct roles in activating intracellular signaling. PFQEP(369-373) initiates NF-kappa B, c-Jun N-terminal kinase, extracellular signal-regulated kinase, and p38 signaling pathways and PVQEET(559-564) activates NF-kappa B and p38 pathways in osteoclasts, whereas PVQEQG(604-609) is only capable of activating NF-kappa B pathway. Significantly, the revelation of these functional RANK cytoplasmic motifs has not only laid a foundation for further delineating RANK signaling pathways in osteoclasts, but, more importantly, these RANK motifs themselves represent potential therapeutic targets for bone disorders such as osteoporosis.

Objective—To examine the role of bovine viral diarrhea virus (BVDV) biotype on the establishment of fetal infection in cattle. Animals—30 mixed-breed pregnant cows. Procedure—Pregnant cows were inoculated oronasally with either i-VVNADL, originating from an infectious BVDV cDNA clone of the National Animal Disease Laboratory (NADL) isolate, or the parental virus stock, termed NADL-A. Results—All cows developed neutralizing antibodies to BVDV, and virus was commonly isolated from peripheral blood mononuclear cells or nasal swab specimens of NADL-A inoculated cows; however, virus was rarely isolated from specimens of i-VVNADL inoculated cows. i-VVNADL did not cause fetal infection, whereas all fetuses harvested from NADL-A inoculated cows at 6 weeks after inoculation had evidence of infection. Immunoblot analysis of fetal virus isolates revealed the absence of NS3, confirming a noncytopathic (NCP) biotype BVDV in the NADL-A stock. The sequence of the NCP contaminant (termed NADL-1102) and the i-VVNADL genome were virtually identical, with the exception of a 270 nucleotide-long insert in the i-VVNADL genome. Phylogenetic analyses revealed that NADL-1102 forms a monophyletic group with 6 other NADL genomes. Conclusions and Clinical Relevance—These data suggest that the contaminating NCP virus in the NADL-A stock was the ancestral NADL virus, which originally infected a bovine fetus and recombined to produce a cytopathic (CP) variant. Following oronasal infection of pregnant cows, viremia and transplacental transmission of CP BVDV to the fetus is rare, compared with the high occurrence of maternal viremia and fetal infection observed with NCP BVDV. (Am J Vet Res 2002;63:1455–1463)

The crosstalk severely affects the viewing experience for the auto-stereoscopic 3D displays based on frontal projection lenticular sheet. To suppress unclear stereo vision and ghosts are observed in marginal viewing zones(MVZs), aberration of the lenticular sheet combining with the frontal projector is analyzed and designed. Theoretical and experimental results show that increasing radius of curvature (ROC) or decreasing aperture of the lenticular sheet can suppress the aberration and reduce the crosstalk. A projector array with 20 micro-projectors is used to frontally project 20 parallax images one lenticular sheet with the ROC of 10 mm and the size of 1.9 m × 1.2 m. The 3D image with the high quality is experimentally demonstrated in both the mid-viewing zone and MVZs in the optimal viewing plane. The 3D clear depth of 1.2m can be perceived. To provide an excellent 3D image and enlarge the field of view at the same time, a novel structure of lenticular sheet is presented to reduce aberration, and the crosstalk is well suppressed.

The cellular receptor for poliovirus CD155 (or PVR) is the founding member of a new class of membrane-associated immunoglobulin-like proteins, which include the mouse tumor-associated antigen E4 (Tage4) and three proteins termed "nectins." Using a yeast two-hybrid screen we have discovered that the cytoplasmic domain of CD155 associates strongly and specifically with Tctex-1, a light chain of the dynein motor complex, the latter representing the major driving force for retrograde transport of endocytic vesicles and membranous organelles. We confirmed the interaction biochemically and by co-immunoprecipitation, and we mapped the Tctex-1 binding site to a SKCSR motif within the juxtamembrane region of CD155. Tctex-1 immunoreactivity was detected in mouse sciatic nerve and spinal cord (two tissues of central importance for poliovirus pathogenesis) in punctate, possibly vesicular, patterns. We propose that the cytoplasmic domain may target CD155-containing endocytic vesicles to the microtubular network. Neurotropic viruses like poliovirus, herpesvirus, rabies virus, and pseudorabies virus all utilize neuronal retrograde transport to invade the central nervous system. Association with Tctex-1 and, hence, with the dynein motor complex may offer an explanation for how poliovirus hijacks the cellular transport machinery to retrogradely ascend along the axon to the neuronal cell body.

The new technique of down-hole oil-water separation and injection in oilproduction is gaining wide popularity around the world, which can used to solvethe problems with high fluid production and thus large amount of separatingwork in development of high watercut field. It can also used to reduce theheavy investments and energy losses due to water injection. Therefor it is agood prospective technique. This technique was proposed and owned by ProductionTechnique Institute of Shengli Petroleum Administration Bureau. Since 1997, thestudying and manufacture of down-hole separation equipment have been finishedsuch as separate pumping pump and rod pumping production string of down-holeoil-water separation and injection etc. The factors influencing the effect ofthe oil-water separation and its efficiency have been found preliminarilythrough laboratory experiments. The field tests show that the technique willreduce the water production rate and expense, and increase oil productionapparently under appropriate conditions. Although there is obscure points ofthis technique, it is worth being researched and developed further.

An antiferromagnetic Fe24Mn4A15Cr covar alloy has been impulse-passivated by an alternating current(AC)voltage overlapping a direct current(DC)voltage,in order to improve its corrosion resistance due to the poor corrosion property of austenite matrix with a high Mn content,a low Cr and/or Al content.The impulse-passivated films were obtained in 1.0 mol/L Na_2SO_4 solution at an AC voltage with modulation amplitude of 180—480 mV during alternating period of 200- 400 ms for modification time of 5—15 min,and simultaneously at a DC voltage of 620 mV.The impulse-passivation parameters in 1.0 mol/L Na_2SO_4+0.5 mol/L H_2SO_4 solution were optimized as amplitude of 380 mV,period of 300 ms,and time of 10 min,by using the potential decline curves. The impulse-passivated films on the covar alloy under the optimum parameters were characterized by Auger electron spectroscopy and X-ray photoelectron spectroscopy(AES/XPS),and evaluated by anodic polarization and electrochemical impedance spectroscopy(EIS),respectively,and compared with those of the constant-passivated films at DC voltage of 620 mV for modification time of 15 min. In the impulse-passivated films on the covar alloy,an enrichment of elements Al and Cr,a lack of Mn in a thick barrier film composed of oxides Al_2O_3 and Cr_2O_3.The anodic polarization curves of the impulse-passivated films have a self-passivation with the higher corrosion potential of—100 mV and lower passive current density of 0.7μA/cm~2,than those of—360 mV and 2.6μA/cm~2 for the constant -passivated films.These electrochemical polarization behaviors were similar to those of the AISI 304 austenitic stainless steel.The EIS of the impulse-passivated films has larger diameter of capacitive arc,higher impedance modulus |Z|,and wider phase degree range,relative to the constant-passivated films.Correspondently,the impulse-passivated film resistant R_p increased to 54.0 kΩ·cm~2 from 14.8 kΩ·cm~2 and effective capacitance decreased to 10.2 /iF/cm2 from 14.0μF/cm~2,using an equivalent electric circuit of R_s-(R_P//CPE).The high insulation of the impulse-passivated films on the covar alloy led to an improved corrosion resistance of the cover alloy.The impulse-passivated antiferromagnetic Fe24Mn4Al5Cr covar alloy exhibits an application potential in wide industrial field.

The synthesis in vitro of poly(U) on a poly(A) template with oligo(dT)15 primer by poliovirus RNA polymerase 3Dpol (280 ng/ml) is strongly stimulated (50-100 fold) by the addition of purified poliovirus polypeptide 3AB. The synthesis of product continues linearly with time for up to 90 min. The reaction with 3Dpol alone can be reactivated and similarly enhanced by the addition of 3AB at 30 min of incubation. Optimal stimulation is achieved under conditions where the concentration of 3Dpol and of template is low, when the molar ratio of 3AB to 3Dpol is about 100:1 and that of 3AB to poly(A) is about 25:1. In the presence of 3AB, the yield of product made by 3Dpol is much increased but its size is unchanged. From a number of basic proteins and peptides tested, a few were found which also exhibited limited enhancement of polymerase activity. The stimulatory effect of 3AB is probably related to its ability to bind both the template-primer, poly(A).oligo(dT)15, and 3Dpol (Molla, A., Harris, K. S., Paul, A. V., Shin, S. H., Mugavero, J., and Wimmer, E. J. (1994) J. Biol. Chem. 269, 27015-27020). RNA synthesis on purified poliovirus RNA with oligo(dT)15 primer is enhanced by 3AB about 5-10 fold, and this reaction is highly sensitive to detergent.

INFECTIOUS eDNA AND IN VITRO SYNTHESIS OF INFECTIOUS VIRAL RNA 359 SELECTION AND GENERATION OF MUTANTS . . . . . . .... . . .. . . .. . . 360 Selection of Spontaneous Mutants . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 363 Random Mutagenesis .... _ . . . . . . . _ _ .... ... ... _ _ . . . . . . . . . 370 Mutagenesis Targeted to Existing or de novo Introduced Restriction Sites . . . . 370 Oligonucleotide-mediated, Site Directed Mutagenesis. . . . . . . . . . . . . . . . . . 371 Exchange of Genomic Segments . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 371 In Search for Revertants . . . . .. . .. . . .. . . . . . . .. . . . . . .. ..... . . 372

This work was also supported by National Institutes of Health Grant R01 DK46395 (to S. J. F.).

Global energy balance in mammals is controlled by the actions of circulating hormones that coordinate fuel production and utilization in metabolically active tissues. Bone-derived osteocalcin, in its undercarboxylated, hormonal form, regulates fat deposition and is a potent insulin secretagogue. Here, we show that insulin receptor (IR) signaling in osteoblasts controls osteoblast development and osteocalcin expression by suppressing the Runx2 inhibitor Twist2. Mice lacking IR in osteoblasts have low circulating undercarboxylated osteocalcin and reduced bone acquisition due to decreased bone formation and deficient numbers of osteoblasts. With age, these mice develop marked peripheral adiposity and hyperglycemia accompanied by severe glucose intolerance and insulin resistance. The metabolic abnormalities in these mice are improved by infusion of exogenous undercarboxylated osteocalcin. These results indicate the existence of a bone-pancreas endocrine loop through which insulin signaling in the osteoblast ensures osteoblast differentiation and stimulates osteocalcin production, which in turn regulates insulin sensitivity and pancreatic insulin secretion to control glucose homeostasis.