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61.
Identification of the sea urchin egg receptor for sperm using an antiserum raised against a fragment of its extracellular domain 总被引:7,自引:0,他引:7 下载免费PDF全文
Sea urchin egg fertilization requires the species-specific interaction of molecules on the sperm and egg surfaces. Previously, we isolated an extracellular, 70-kD glycosylated fragment of the S. purpuratus egg receptor for sperm by treating the eggs with lysylendoproteinase C (Foltz, K. R., and W. J. Lennarz. 1990. J. Cell Biol. 111:2951-2959). To characterize the receptor further, we have generated a polyclonal antiserum (anti-70KL) against the purified 70-kD fragment. Anti-70KL was found to react with a single polypeptide of approximately 350 kD on Western blots, presumed to be the intact receptor, in an egg cell surface preparation. This polypeptide appeared to be tightly associated with the plasma membrane/vitelline layer complex, as it was released from these preparations only by detergent treatment. Immunofluorescence microscopy revealed that the receptor was distributed evenly over the egg surface. The anti-70KL was species specific both in its ability to recognize the egg surface protein and to inhibit sperm binding. Fab fragments generated from affinity-purified anti-70KL also bound to the egg surface and inhibited sperm binding in a concentration-dependent manner. Interestingly, treatment with Fabs caused a small percentage of eggs to undergo cortical granule exocytosis, even in the absence of external Ca2+. These results confirm earlier findings indicating that the receptor is a cell surface glycoprotein of high molecular weight that species specifically binds sperm. This antiserum provides a powerful tool for further investigation of gamete interactions and the structure of the sperm receptor. 相似文献
62.
63.
Black BE Jansen LE Maddox PS Foltz DR Desai AB Shah JV Cleveland DW 《Molecular cell》2007,25(2):309-322
Active centromeres are marked by nucleosomes assembled with CENP-A, a centromere-specific histone H3 variant. The CENP-A centromere targeting domain (CATD), comprised of loop 1 and the alpha2 helix within the histone fold, is sufficient to target histone H3 to centromeres and to generate the same conformational rigidity to the initial subnucleosomal heterotetramer with histone H4 as does CENP-A. We now show in human cells and in yeast that depletion of CENP-A is lethal, but recruitment of normal levels of kinetochore proteins, centromere-generated mitotic checkpoint signaling, chromosome segregation, and viability can be rescued by histone H3 carrying the CATD. These data offer direct support for centromere identity maintained by a unique nucleosome that serves to distinguish the centromere from the rest of the chromosome. 相似文献
64.
Ewelina Zasadzińska Meghan C Barnhart‐Dailey P Henning J L Kuich Daniel R Foltz 《The EMBO journal》2013,32(15):2113-2124
The epigenetic mark of the centromere is thought to be a unique centromeric nucleosome that contains the histone H3 variant, centromere protein‐A (CENP‐A). The deposition of new centromeric nucleosomes requires the CENP‐A‐specific chromatin assembly factor HJURP (Holliday junction recognition protein). Crystallographic and biochemical data demonstrate that the Scm3‐like domain of HJURP binds a single CENP‐A–histone H4 heterodimer. However, several lines of evidence suggest that HJURP forms an octameric CENP‐A nucleosome. How an octameric CENP‐A nucleosome forms from individual CENP‐A/histone H4 heterodimers is unknown. Here, we show that HJURP forms a homodimer through its C‐terminal domain that includes the second HJURP_C domain. HJURP exists as a dimer in the soluble preassembly complex and at chromatin when new CENP‐A is deposited. Dimerization of HJURP is essential for the deposition of new CENP‐A nucleosomes. The recruitment of HJURP to centromeres occurs independent of dimerization and CENP‐A binding. These data provide a mechanism whereby the CENP‐A pre‐nucleosomal complex achieves assembly of the octameric CENP‐A nucleosome through the dimerization of the CENP‐A chaperone HJURP. 相似文献
65.
Rubio-Aliaga I Boll M Vogt Weisenhorn DM Foltz M Kottra G Daniel H 《The Journal of biological chemistry》2004,279(4):2754-2760
The new member of the mammalian amino acid/auxin permease family, PAT2, has been cloned recently and represents an electrogenic proton/amino acid symporter. PAT2 and its paralog, PAT1/LYAAT-1, are transporters for small amino acids such as glycine, alanine, and proline. Our immunodetection studies revealed that the PAT2 protein is expressed in spinal cord and brain. It is found in neuronal cell bodies in the anterior horn in spinal cord and in brain stem, cerebellum, hippocampus, hypothalamus, rhinencephalon, cerebral cortex, and olfactory bulb in the brain. PAT2 is expressed in neurons positive for the N-methyl-d-aspartate subtype glutamate receptor subunit NR1. PAT2 is not found in lysosomes, unlike its paralog PAT1, but is present in the endoplasmic reticulum and recycling endosomes in neurons. PAT2 has a high external proton affinity causing half-maximal transport activation already at a pH of 8.3, suggesting that its activity is most likely not altered by physiological pH changes. Transport of amino acids by PAT2 activity is dependent on membrane potential and can occur bidirectionally; membrane depolarization causes net glycine outward currents. Our data suggest that PAT2 contributes to neuronal transport and sequestration of amino acids such as glycine, alanine, and/or proline, whereby the transport direction is dependent on the sum of the driving forces such as substrate concentration, pH gradient, and membrane potential. 相似文献
66.
Foltz DJ Castro-Perez J Riley P Entwisle JR Baker TR 《Journal of chromatography. B, Analytical technologies in the biomedical and life sciences》2005,825(2):144-151
The identification of in vitro and in vivo metabolites is vital to the discovery and development of new pharmaceutical therapies. Analytical strategies to identify metabolites at different stages of this process vary, but all involve the use of liquid chromatography separations combined with detection via mass spectrometry (HPLC/MS). Reported here is the use of narrow-bore column (0.5-1.0 mm i.d.) trapping of metabolites, followed by back-flushing onto a matching analytical column. Separated metabolites were then identified using quadrupole time-of-flight mass spectrometry (MS) and tandem MS. Metabolites in human plasma and from low-level in vitro incubations, that were not identified using standard HPLC/MS approaches, were characterized using the instrumental configuration described here. 相似文献
67.
Barley aleurone cells contain two types of vacuoles. Characterization Of lytic organelles by use of fluorescent probes 总被引:10,自引:1,他引:9 下载免费PDF全文
Light microscopy was used to study the structure and function of vacuoles in living protoplasts of barley (Hordeum vulgare cv Himalaya) aleurone. Light microscopy showed that aleurone protoplasts contain two distinct types of vacuole: the protein storage vacuole and a lysosome-like organelle, which we have called the secondary vacuole. Fluorescence microscopy using pH-sensitive fluorescent probes and a fluorogenic substrate for cysteine proteases showed that both protein storage vacuoles and secondary vacuoles are acidic, lytic organelles. Ratio imaging showed that the pH of secondary vacuoles was lower in aleurone protoplasts incubated in gibberellic acid than in those incubated in abscisic acid. Uptake of fluorescent probes into intact, isolated protein storage vacuoles and secondary vacuoles required ATP and occurred via at least two types of vanadate-sensitive, ATP-dependent tonoplast transporters. One transporter catalyzed the accumulation of glutathione-conjugated probes, and another transported probes not conjugated to glutathione. 相似文献
68.
Segregation and linkage studies of allozyme loci in pair crosses of the oysterCrassostrea virginica 总被引:4,自引:0,他引:4
David W. Foltz 《Biochemical genetics》1986,24(11-12):941-956
The genetic control of 11 electrophoretically detected allozyme polymorphisms in the oyster Crassostrea virginica was investigated in 10 pair crosses. For nine allozyme loci, each offspring shared at least one band (electromorph) with each parent. For the remaining two loci (mannosephosphate isomerase and leucine aminopeptidase-2), some offspring failed to share a band with one or both parents. Several lines of evidence indicated that these anomalous results were due to transmission of null alleles. There was evidence of distorted segregation at 8 of the 11 loci. The departures from the Mendelian expectations within the pair crosses might be due either to viability selection in the offspring or to gametic selection in one or both parents, although the possibility that the distortion is due to a locus linked to the allozyme locus cannot be ruled out. However, there was no evidence that heterozygosity per se had an effect on viability of offspring within a cross. Linkage analysis revealed two linkage groups, one consisting of four allozyme loci and the other consisting of three loci. 相似文献
69.
Charmaine Foltz ;Larry Carbone ;David DeLong ;Bernard E. Rollin ;Pascalle Van Loo ;Julia Whitaker ;Axel Wolff 《中国实验动物学杂志》2008,(1):26-26
At the 2006 National Meeting of the American Association of Laboratory Animal Science, a panel discussed the question of what constitutes optimal or acceptable housing density for mice. Though there is a consensus that present guidelines are somewhat arbitrarily defined, scientific research has not yet been able to provide clear recommendations for amending them. Speakers explored the many factors that influence decisions on mouse housing, including regulatory requirements, scientific data and their interpretation, financial considerations and ethical concems. The panel largely agreed that animal well-being should be the measure of interest in evaluating housing density and that well-being includes not only physical health, but also animals' behavior, productivity and preference. Lab Animal 2007,36(10) 相似文献
70.
Nine of 100 plant food samples investigated yielded salmonellae. A brilliant green tetrathionate-brilliant green agar combination gave the most salmonellae recoveries. 相似文献