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  • Life and Medical Sciences  (318)
  • Physics  (71)
  • Atomic, Molecular and Optical Physics  (54)
  • ASTROPHYSICS
  • Wiley-Blackwell  (443)
  • 1995-1999  (443)
  • 1
    ISSN: 0730-2312
    Keywords: osteocalcin ; transcriptional regulation ; homeodomain protein ; Msx ; bone-specific ; OC box ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Bone-specific expression of the osteocalcin gene is transcriptionally controlled. Deletion analysis of osteocalcin promoter sequences by transient transfection of osseous (ROS 17/2.8) and nonosseous (R2 fibroblast) cells revealed that the most proximal 108 nucleotides are sufficient to confer tissue-specific expression. By gel mobility shift assays with wild-type and mutated oligonucleotides and nuclear extracts from several different cell lines we identified a novel transcription factor complex which exhibits sequence-specific interactions with the primary transcriptional element, the OC box (nt -99 to -76). This OC box binding protein (OCBP) is present only in osteoblast-like cells. Methylation interference demonstrated association of the factor with OC box sequences overlapping the Msx homeodomain consensus binding site. By assaying several mutations of the OC box, both in gel shift and transient transfection studies using ROS 17/2.8, we show the following. First, binding of OCBP correlates with osteocalcin promoter activity in ROS 17/2.8 cells. Increased binding leads to a 2-3-fold increase in transcription, while decreased binding results in transcription 30-40% of control. Second, homeodomain protein binding suppresses transcription. However, Msx expression is critical for full development of the bone phenotype as determined by antisense studies. Last, we show that one of the mutations of the OC box permits expression of osteocalcin in non-osseous cell lines. In summary, we demonstrate association of at least two classes of tissue-restricted transcription factors with the OC box element, the OCBP and Msx proteins, supporting the concept that these sequences contribute to defining tissue specificity. © 1996 Wiley-Liss, Inc.
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  • 2
    ISSN: 0730-2312
    Keywords: epitope mapping ; monoclonal antibodies ; linear epitope ; immuno-dominant ; immuno-recessive ; ELISA ; competitive ELISA ; recombinant GST-PSP94 ; N-terminal and C-terminal peptides ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: PSP94 has shown potential to be a serum biomarker for evaluating prostate cancer. Studies of the epitope structure is crucial for this endeavour. In this article, we have used 15 different monoclonal antibodies (MAb) to analyse the epitope structure of PSP94 and to compare with the results obtained from our previous work using polyclonal antibody and recombinant PSP94. Firstly, we determined the relative activities of the 15 MAb population by direct and competitive ELISA. The two predominant MAbs (MAb PSP-6 and -19) in 15 MAbs were selected for further studies of the epitope structure. By comparing the binding activities of recombinant GST-PSP94 and natural PSP94 with MAbs, and by comparing their affinity with MAbs in an in vitro denaturing experiment, PSP94 was shown to have a similar, prevalently linear epitope structure as we demonstrated by polyclonal antibody. Using recombinant GST fusion protein with PSP94 and with each half of the N- and C-terminal 47 amino acids (GST-PSP-N47/C47) in E. coli cells, the different epitopes recognized by 15 monoclonal antibodies were delineated and the polar distribution of the epitope structure of PSP94 was characterized. Results of direct ELISA of recombinant N47 and C47 and their competitive binding against natural PSP94 (competitive ELISA) showed that the N- and C-termini represent the immuno-dominant and immuno-recessive area separately. A majority of the monoclonal antibodies (12/15) showed preferential binding of the N-terminal sequence of the PSP94 protein. Using GST-PSP-N47 as a standard protein, an epitope map of the 15 monoclonal antibodies was obtained. The results of this study will help to define the clinical utility of PSP94. J. Cell. Biochem. 65:186-197. © 1997 Wiley-Liss, Inc.
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  • 3
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Molecular Reproduction and Development 40 (1995), S. 69-83 
    ISSN: 1040-452X
    Keywords: SGP-1 ; Hypophysectomy ; Castration ; Efferent ducts ; Lysosomes ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The objective of this study was to define the factors regulating the endogenous production of sulfated glycoprotein-1 (SGP-1) in nonciliated cells of the efferent ducts. To this end we examined five different groups of animals undergoing the following experimental procedures: (1) hypophysectomized animals at 7, 14, and 28 days, (2) 7-day hypophysectomized rats receiving testosterone implants given at various time intervals thereafter, (3) castration at various time intervals up to 7 days, (4) 7-day castrated rats receiving testosterone implants at various time intervals thereafter, and (5) castrated rats given testosterone implants immediately after castration and sacrificed at different time intervals thereafter. Efferent ducts were fixed by perfusion with 4% paraformaldehyde and 0.5% glutaraldehyde in phosphate buffer for quantitative immunocytochemical analysis at the level of the electron microscope. For each experimental condition and their controls, the number of gold particles/μm2 within the endosomal and lysosomal compartments was calculated taking into account the changes in both the volume of the cell and organelles being quantified and expressed as labeling content. The results revealed that hypophysectomy (up to 4 weeks) caused a marked significant decrease in the SGP-1 labeling content of the endosomal and lysosomal compartments. The labeling content of the lysosomal compartment of efferent ducts from rats castrated for up to 1 week did not change significantly. However, there was a significant decrease in the labeling content of endosomes. This decrease is due to SGP-1, which is secreted by Sertoli cells, not being available for uptake in the efferent aucts. These results suggested that testosterone is not required for maintaining the high labeling content of SGP-1 within lysosomes of nonciliated cells, but that a pituitary factor appears to be needed. The administration of testosterone at different intervals to 7-day castrated animals resulted in a significant decrease of lysosomal SGP-1, suggesting that testosterone under these experimental conditions inhibits the production of a pituitary factor that maintains the high labeling content of SGP-1 within lysosomes of the nonciliated cells. Testosterone administered to 7-day hypophysectomized animals over a 24-hr period had no effect on the labeling content of SGP-1 within lysosomes. However, the administration of testosterone to animals immediately following castration showed no differences in the labeling content of SGP-1 within compared to controls. Together these results suggest that the labeling content of SGP-1 within lysosomes of nonciliated cells of the efferent ducts is not dependent on luminal or circulating androgens, nor is it dependent on a testicular factor entering the lumen of the ducts. It does appear, however, that SGP-1 synthesis and targeting to secondary lysosomes is dependent on a pituitary factor that may have a direct or an indirect effect on the nonciliated cells. © 1995 Wiley-Liss, Inc.
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  • 4
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Molecular Reproduction and Development 41 (1995), S. 287-299 
    ISSN: 1040-452X
    Keywords: SGP-1 ; SGP-2 ; Postnatal development ; Nonciliated cells ; Efferent ducts ; Rats ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Sulfated glycoprotein (SGP) -1 and -2, secretory products of Sertoli cells, are secreted into the lumen of seminiferous tubules where they bind to late spermatids. Once released, the spermatozoa traverse the efferent ducts where these proteins detach from their surface and are endocytosed by the nonciliated cells. In adult animals, SGP-1 and SGP-2 are also synthesized by nonciliated cells and targeted from the Golgi apparatus to lysosomes. The purpose of the present study was to determine the pattern of expression of SGP-1 and SGP-2 within nonciliated cells during postnatal development. The efferent ducts of animals at different postnatal ages were prepared for an electron microscopic immunocytochemical quantitative analysis as well as for Northern blot analysis. The data expressed as labeling content (no. gold particles/μm2 and taking into account the volume of the endocytic or-ganelles and the cell) revealed that anti-SGP-1 labeling in endosomes of nonciliated cells was minimal at 15, 21, and 29 days of age. On the other hand, the lysosomal labeling content showed a significant increase by day 29 compared to 15 and 21-day-old animals indicating that an endogenous form of SGP-1 was being synthesized by nonciliated cells and targeted to lysosomes. By day 39 a significant increase in endosomal labeling occurred; this was attributed to the endocytosis of Sertoli-derived SGP-1 which coincided with the entry of spermatozoa into the lumen of these ducts at this age. Lysosomal labeling showed further significant increases at days 39, 49, and then again at day 90. Northern blot analysis detected SGP-1 mRNA transcripts at all postnatal ages examined. While decreases or increases in transcripts could not be determined due to the greater amount of tissue present with increasing age, these data taken together support the idea of an endogenous form of SGP-1 being synthesized by nonciliated cells and targeted to lysosomes during postnatal development.In the case of SGP-2, endosomal labeling was minimal at 15, 21, and 29 days of age but was significantly increased by day 39, with similar values at all subsequent ages. The high value at day 39 was attributed to the endocytosis of SGP-2 which coincided with the entry of spermatozoa into the lumen at this age. Lysosomal labeling, on the other hand, was low at days 15 and 21 but peaked at day 29 at a time when endosomal labeling was minimal. These results suggested the synthesis of an endogenous form of SGP-2 which was being targeted to lysosomes. Similar values for SGP-2 lysosomal labeling comparable to that at day 29 were obtained at all other ages. Since SGP-2 endosomal labeling was significantly increased at day 39 and maintained thereafter, it is suggested that labeling in lysosomes at this and subsequent ages could also be due to the endocytosis of Sertoli-derived SGP-2. However, Northern blot analysis confirmed the presence of mRNA transcripts for SGP-2 at all postnatal ages examined, although increases or decreases in their amount were not determined. These results thus consolidate the hypothesis of an endogenous form of SGP-2 being synthesized by nonciliated cells and targeted to lysosomes. Finally, since the amounts of endogenous SGP-1 and SGP-2 peak at different ages, it is suggested that different factors are involved in regulation of these two proteins during postnatal development. © 1995 Wiley-Liss, Inc.
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  • 5
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Molecular Reproduction and Development 40 (1995), S. 286-291 
    ISSN: 1040-452X
    Keywords: Embryo development ; Growth factor ; Inhibin ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: It is evident that members of several growth factor families are actively involved in embryogenesis from its earliest phases. Several reports also indicate the oviduct as a possible source of growth factors, suggesting an active role of this organ in mammalian embryonic development. The aim of this study was to investigate the presence of activin/inhibin subunits in bovine oviduct since activin is a well-characterised morphogen in amphibian development. The presence of transcripts for α. βA, and βB subunits was investigated by analysing oviduct epithelial cells mRNA with reverse transcription-polymerase chain reaction (RT-PCR). Moreover, antisera specific for the three subunits were used for the Western blot analysis of the proteins secreted by oviduct epithelial cells in vitro and for their immunohistochemical localisation in different oviductal regions. Oviduct epithelial cells expressed only the βA-subunit gene. Immunoreactive material was present among in vitro secreted proteins, indicating that the transcript is translated into a polypeptide that has been localised in the epithelium of both the ampullary and isthmic tract of the organ. Consistent with these results, the antisera for the α and βB subunits did not recognise any specific antigen either among secreted proteins or in the sections. These results indicate that βA subunit gene is expressed in bovine oviduct epithelial cells, and the protein is secreted in vitro and can be found along the whole extension of the organ. In the absence of α or βB subunits, this suggests that activin A is present in bovine oviduct. Such a finding would be consistent with an embry-otrophic activity of this organ, but definitive conclusions on the target tissue and the specific functions of oviductal activin require further studies. © 1995 Wiley-Liss, Inc.
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  • 6
    ISSN: 1040-452X
    Keywords: Spermatogenesis ; Sperm-zona pellucida binding ; Transmembrane animal lectin proteins ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Galactosyl receptor, a cell surface Ca2+-dependent lectin with binding affinity for galactose, was evaluated by immunoblotting, immunoprecipitation, Northern blotting, and immunocytochemistry in human liver, testis, and sperm. Polyclonal antisera raised against the minor asialoglycoprotein receptor variant of rat hepatocytes (designated rat hepatic lectin-2/3, RHL-2/3), and its human liver-equivalent (designated H2), recognize native galactosyl receptor in the testis and sperm in immunoblotting, immunoprecipitation, and immunocytochemical experiments. An equivalent to the major hepatocyte asialoglycoprotein receptor variant (rat RHL-1 and human H1) was not detected. Human testis and sperm galactosyl receptor was resolved, after immunoprecipitation and immunoblotting, as a single protein component of molecular mass 50 kD. The single protein component in human testis and sperm contrasted with the doublet nature of rat testis and sperm galactosyl receptor, consisting of two components of molecular masses of 54 and 49 kD. Northern blotting experiments using radiolabeled H1 and H2 cDNA probes confirmed the presence of H2 mRNA and the lack of H1 mRNA in the human testis. Immunocytochemical studies detected specific antigenic sites on the entire surfaces of spermatogenic cells. However, immunoreactivity in epididymal and ejaculated sperm was confined to head surfaces overlying the acrosome. Results from these studies, and from previous studies in the rat, suggest that the testis/sperm galactosyl receptor is a C-type Ca2+-dependent lectin with possible roles in cell-cell interaction during spermatogenesis and sperm-zona pellucida binding at fertilization. © 1995 Wiley-Liss, Inc.
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  • 7
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    International Journal of Quantum Chemistry 60 (1996), S. 1525-1536 
    ISSN: 0020-7608
    Keywords: Computational Chemistry and Molecular Modeling ; Atomic, Molecular and Optical Physics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Chemistry and Pharmacology
    Notes: The local self-consistent field (LSCF) method which allows full SCF computations on a fragment of very long molecular systems represented by a classical force field has been adapted to the description of nonmetallic crystals. The periodicity of the network is achieved by modifying self-consistently the basic parameters of the classical subsystem (charges, geometric parameters) along the SCF iterative scheme. The method is tested on α-cristobalite. The parametrization of the quantum classical junction, achieved by localized bond orbitals has been performed with the help of a fragment located in the bulk. The stability of the method with respect to the size of the fragment and the size of the crystalline sample appears to be very good. The properties of fully hydroxylated (010) surface are corrected described. Modeling of water adsorption on the ideal surface as well as on two kinds of surface defects gives rise to very reasonable results with an absorption energies of ca 10 kcal/mol which are close to the upper limit of the experimental data. This preliminary study appears to be quite encouraging regarding the possibilities of using this method, which can be considered as an extension of the embedded cluster approach to covalent solids. Many applications to surface chemical reactivity studies can be imagined. © 1996 John Wiley & Sons, Inc.
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  • 8
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    International Journal of Quantum Chemistry 56 (1995), S. 371-384 
    ISSN: 0020-7608
    Keywords: Computational Chemistry and Molecular Modeling ; Atomic, Molecular and Optical Physics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Chemistry and Pharmacology
    Notes: Quantum crystallography is a developing technique for extracting quantum mechanically valid properties from X-ray diffraction experiments. Quantum mechanics and crystallography are joined through the fact that the electron distributions around atoms are the source of X-ray diffraction and electron density distributions are observables that lend themselves readily to quantum mechanical description. Direct contact with the X-ray diffraction data is made by equating the structure factor magnitudes, which are readily obtained from the measured X-ray diffraction intensities, with the magnitudes of certain Fourier transforms of the quantum mechanical description of the electron distribution. The quantum mechanical description of the electron density involves molecular orbitals and an associated matrix. By requiring that the associated matrix be a projector with a normalized trace, while optimizing the fit to the experimental structure factor magnitudes, strong constraints are imposed on the relationship between the X-ray data and the quantum mechanical description of the electron density distribution. The final result should be a wave function that is in good agreement with the X-ray diffraction information and from which a variety of properties, e.g., electron densities and electrostatic potentials, could be extracted. The method for making the fit to the X-ray data involves the use of least-squares calculations in which the defining equations are the structure factor equations and equations that arise from the conditions defining a projector matrix with a normalized trace. The variables are the elements of the projector. It is also possible to refine such parameters as atomic coordinates. The calculations are facilitated by use of good initial projector matrices. For this purpose, a method has been developed for generating projector matrices for large molecules from the sum of kernel matrices. Kernel matrices are obtained from the use of fragments of the known atomic coordinates of the substance of interest and the application of molecular orbital methods in quantum mechanics. The use of fragments is justified by the fact that overlap integrals rapidly approach zero as the distances between atoms increase. © 1995 John Wiley & Sons, Inc.
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  • 9
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    International Journal of Quantum Chemistry 60 (1996), S. 579-592 
    ISSN: 0020-7608
    Keywords: Computational Chemistry and Molecular Modeling ; Atomic, Molecular and Optical Physics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Chemistry and Pharmacology
    Notes: Position and momentum space plots are presented for localized molecular orbitals in hydrocarbons, boranes, a carborane, and two octahedral transition metal complexes. The p-space representation proves to be valuable for visualizing such orbitals since it highlights the differences in their character from one molecule to another. Factors influencing the form of the orbitals in p space, including the oscillatory behavior caused by contributions to an orbital from more than one center, are examined in detail. © 1996 John Wiley & Sons, Inc.
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  • 10
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    International Journal of Quantum Chemistry 60 (1996), S. 1691-1700 
    ISSN: 0020-7608
    Keywords: Computational Chemistry and Molecular Modeling ; Atomic, Molecular and Optical Physics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Chemistry and Pharmacology
    Notes: By performing ab initio calculations on fragments of leu1(SINGLEBOND)zervamicin, it is demonstrated that accurate wave functions can be obtained for this large structure by combining the results from the fragments. Input information consists of atomic coordinates as obtained, for example, from a crystal structure determination. The fragments are composed of a kernel of atoms surrounded by a chosen neighborhood of atoms. The entire molecule is divided into individual kernels and their neighborhoods are added for the individual fragment calculations on the basis of the distances of other atoms from the atoms in a kernel. The hydrated leu1(SINGLEBOND)zervamicin is composed of 295 atoms which could be handled in full by Gaussian 94. The results of the fragment calculations were combined to provide an electron density distribution for the molecule. This distribution was compared with one that represents the distribution obtained from a calculation on the entire molecule at once. The clear implication of this study is that it is feasible to perform ab initio calculations on structures by the use of fragments. The time involved increases essentially linearly with complexity. © 1996 John Wiley & Sons, Inc.
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