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J D Taylor

Publications and source records attributed to J D Taylor.

At least 19 recordsLinked to original sources
PubMed2000

Cultivar-specific avirulence and virulence functions assigned to avrPphF in Pseudomonas syringae pv. phaseolicola, the cause of bean halo-blight disease.

The avrPphF gene was cloned from Pseudomonas syringae pathovar phaseolicola (PPH:) races 5 and 7, based on its ability to confer avirulence towards bean cultivars carrying the R1 gene for halo-blight resistance, such as Red Mexican. avrPphF comprised two open reading frames, which were both required for function, and was located on a 154 kb plasmid (pAV511) in PPH: Strain RW60 of PPH:, lacking pAV511, displayed a loss in virulence to a range of previously susceptible cultivars such as Tendergreen and Canadian Wonder. In Tendergreen virulence was restored to RW60 by avrPphF alone, whereas subcloned avrPphF in the absence of pAV511 greatly accelerated the hypersensitive resistance reaction caused by RW60 in Canadian Wonder. A second gene from pAV511, avrPphC, which controls avirulence to soybean, was found to block the activity of avrPphF in Canadian Wonder, but not in Red Mexican. avrPphF also conferred virulence in soybean. The multiple functions of avrPphF illustrate how effector proteins from plant pathogens have evolved to be recognized by R gene products and, therefore, be classified as encoded by avirulence genes.

Base Sequence↗
PubMed2000

Exclusion limits on the WIMP-nucleon cross section from the cryogenic dark matter search.

The Cryogenic Dark Matter Search (CDMS) employs Ge and Si detectors to search for weakly interacting massive particles (WIMPs) via their elastic-scattering interactions with nuclei while discriminating against interactions of background particles. CDMS data, accounting for the neutron background, give limits on the spin-independent WIMP-nucleon elastic-scattering cross section that exclude unexplored parameter space above 10 GeV/c2 WIMP mass and, at >75% C.L., the entire 3sigma allowed region for the WIMP signal reported by the DAMA experiment.

Journal Article↗
PubMed2000

Multiplexed single nucleotide polymorphism genotyping by oligonucleotide ligation and flow cytometry.

BACKGROUND: We have developed a rapid, high throughput method for single nucleotide polymorphism (SNP) genotyping that employs an oligonucleotide ligation assay (OLA) and flow cytometric analysis of fluorescent microspheres. METHODS: A fluoresceinated oligonucleotide reporter sequence is added to a "capture" probe by OLA. Capture probes are designed to hybridize both to genomic "targets" amplified by polymerase chain reaction and to a separate complementary DNA sequence that has been coupled to a microsphere. These sequences on the capture probes are called "ZipCodes". The OLA-modified capture probes are hybridized to ZipCode complement-coupled microspheres. The use of microspheres with different ratios of red and orange fluorescence makes a multiplexed format possible where many SNPs may be analyzed in a single tube. Flow cytometric analysis of the microspheres simultaneously identifies both the microsphere type and the fluorescent green signal associated with the SNP genotype. RESULTS: Application of this methodology is demonstrated by the multiplexed genotyping of seven CEPH DNA samples for nine SNP markers located near the ApoE locus on chromosome 19. The microsphere-based SNP analysis agreed with genotyping by sequencing in all cases. CONCLUSIONS: Multiplexed SNP genotyping by OLA with flow cytometric analysis of fluorescent microspheres is an accurate and rapid method for the analysis of SNPs.

Chromosomes, Human, Pair 19↗
PubMed2000

Excision from tRNA genes of a large chromosomal region, carrying avrPphB, associated with race change in the bean pathogen, Pseudomonas syringae pv. phaseolicola.

Pseudomonas syringae pv. phaseolicola (Pph) race 4 strain 1302A carries avirulence gene avrPphB. Strain RJ3, a sectoral variant from a 1302A culture, exhibited an extended host range in cultivars of bean and soybean resulting from the absence of avrPphB from the RJ3 chromosome. Complementation of RJ3 with avrPphB restored the race 4 phenotype. Both strains showed similar in planta growth in susceptible bean cultivars. Analysis of RJ3 indicated loss of > 40 kb of DNA surrounding avrPphB. Collinearity of the two genomes was determined for the left and right junctions of the deleted avrPphB region; the left junction is approximately 19 kb and the right junction > 20 kb from avrPphB in 1302A. Sequencing revealed that the region containing avrPphB was inserted into a tRNALYS gene, which was re-formed at the right junction in strain 1302A. A putative lysine tRNA pseudogene (PsitRNALYS) was found at the left junction of the insertion. All tRNA genes were in identical orientation in the chromosome. Genes near the left junction exhibited predicted protein homologies with gene products associated with a virulence locus of the periodontal pathogen Actinobacillus actinomycetemcomitans. Specific oligonucleotide primers that differentiate 1302A from RJ3 were designed and used to demonstrate that avrPphB was located in different regions of the chromosome in other strains of Pph. Deletion of a large region of the chromosome containing an avirulence gene represents a new route to race change in Pph.

Base Sequence↗
PubMed2000

A microsphere-based assay for multiplexed single nucleotide polymorphism analysis using single base chain extension.

A rapid, high throughput readout for single-nucleotide polymorphism (SNP) analysis was developed employing single base chain extension and cytometric analysis of an array of fluorescent microspheres. An array of fluorescent microspheres was coupled with uniquely identifying sequences, termed complementary ZipCodes (cZipCodes), which allowed for multiplexing possibilities. For a given assay, querying a polymorphic base involved extending an oligonucleotide containing both a ZipCode and a SNP-specific sequence with a DNA polymerase and a pair of fluoresceinated dideoxynucleotides. To capture the reaction products for analysis, the ZipCode portion of the oligonucleotide was hybridized with its cZipCodes on the microsphere. Flow cytometry was used for microsphere decoding and SNP typing by detecting the fluorescein label captured on the microspheres. In addition to multiplexing capability, the ZipCode system allows multiple sets of SNPs to be analyzed by a limited set of cZipCode-attached microspheres. A standard set of non-cross reactive ZipCodes was established experimentally and the accuracy of the system was validated by comparison with genotypes determined by other technologies. From a total of 58 SNPs, 55 SNPs were successfully analyzed in the first pass using this assay format and all 181 genotypes across the 55 SNPs were correct. These data demonstrate that the microsphere-based single base chain extension (SBCE) method is a sensitive and reliable assay. It can be readily adapted to an automated, high-throughput genotyping system. [Primer sequences used in this study are available as online supplementary materials at www.genome.org.]

DNA, Complementary↗
PubMed1999

Identification of a pathogenicity island, which contains genes for virulence and avirulence, on a large native plasmid in the bean pathogen Pseudomonas syringae pathovar phaseolicola.

The 154-kb plasmid was cured from race 7 strain 1449B of the phytopathogen Pseudomonas syringae pv. phaseolicola (Pph). Cured strains lost virulence toward bean, causing the hypersensitive reaction in previously susceptible cultivars. Restoration of virulence was achieved by complementation with cosmid clones spanning a 30-kb region of the plasmid that contained previously identified avirulence (avr) genes avrD, avrPphC, and avrPphF. Single transposon insertions at multiple sites (including one located in avrPphF) abolished restoration of virulence by genomic clones. Sequencing 11 kb of the complementing region identified three potential virulence (vir) genes that were predicted to encode hydrophilic proteins and shared the hrp-box promoter motif indicating regulation by HrpL. One gene achieved partial restoration of virulence when cloned on its own and therefore was designated virPphA as the first (A) gene from Pph to be identified for virulence function. In soybean, virPphA acted as an avr gene controlling expression of a rapid cultivar-specific hypersensitive reaction. Sequencing also revealed the presence of homologs of the insertion sequence IS100 from Yersinia and transposase Tn501 from P. aeruginosa. The proximity of several avr and vir genes together with mobile elements, as well as G+C content significantly lower than that expected for P. syringae, indicates that we have located a plasmid-borne pathogenicity island equivalent to those found in mammalian pathogens.

Bacterial Proteins↗
PubMed1999

Apoptosis induction by a novel anti-prostate cancer compound, BMD188 (a fatty acid-containing hydroxamic acid), requires the mitochondrial respiratory chain.

We recently developed a class of novel anti-prostate cancer compounds, cyclic hydroxamates that elicit a potent apoptotic response in many tumor cells cultured in vitro (D.G. Tang et al., Biochem. Biophys. Res. Commun., 242: 380-384, 1998). The lead compound, termed BMD188, induces programmed cell death in a variety of prostate cancer cells in vitro as well as in vivo (L. Li et al., Anticancer Res., 19: 51-70, 1999). BMD188 kills androgen-independent prostate cancer cells as well as prostate cancer cells with a multidrug-resistance phenotype. The apoptotic effect of BMD188 in prostate cancer cells does not depend on cell cycle, p53 status, or its purported target, arachidonate 12-lipoxygenase, but does require caspase activation and seems to involve mitochondria. To synthesize more specific and effective anti-prostate cancer hydroxamic acid compounds, it is important to understand their mechanism(s) of action. In the present study, we studied the role of mitochondrial respiratory chain (MRC) in BMD188-induced apoptosis in androgen-independent prostate cancer PC3 cells and compared its effect with that of staurosporine (STS), a widely used apoptosis inducer. Several lines of evidence indicate that BMD188-induced cell death depends on MRC: (a) the death could be significantly inhibited by several complex-specific respiration inhibitors; (b) respiration-deficient rho0 cells were more resistant than wild-type parent cells to apoptosis induction by BMD188; and (c) BMD188 induced a rapid increase in reactive oxygen species in mitochondria, an up-regulation of cytochrome c oxidase subunits, a biphasic alteration (i.e., an early hyperpolarization, followed by later hypopolarization) in the mitochondrial membrane potential (delta psi(m)), dramatic changes in mitochondrial morphology and distribution prior to caspase activation, and an abnormal proliferation of mitochondria at the ultrastructural level. By contrast, STS-induced PC3 apoptosis seemed not to depend on MRC. Taken together, the data suggest that the MRC represents a functional target for anti-prostate cancer hydroxamates.

Antineoplastic Agents↗
PubMed1999

Prostate secretory protein (PSP94) suppresses the growth of androgen-independent prostate cancer cell line (PC3) and xenografts by inducing apoptosis.

BACKGROUND: PSP94 (prostate secretory protein of 94 aa; also called PIP), one of the predominant proteins secreted into the seminal fluid, was proposed as an independent diagnostic/prognostic marker for prostate cancers. It was also shown to inhibit rat prostate cancer growth. In this study, we investigated the effect of purified PSP94 on the growth of androgen-independent human prostate cancer cells (PC3) and its potential mechanism of action. METHODS AND RESULTS: PSP94, in a dose- and time-dependent manner, inhibited the growth of PC3 cells. The protein demonstrated a stronger inhibitory effect on the colony-forming ability of PC3 cells in soft agar. A daily injection of PSP94 at 5 microg/kg/body weight resulted in a 50-60% inhibition in the growth of PC3 xenografts in athymic mice. PC3 cell growth inhibition by PSP94 resulted from cell death characteristic of morphological apoptosis, which was confirmed by dual fluorescence microscopy, electron microscopy, and DNA fragmentation assays. Mechanistic studies indicated that PSP94 enhanced the expression of proapoptotic protein Bax without affecting Bcl-2 levels. CONCLUSIONS: This study suggests that PSP94 may represent a novel, apoptosis-based, antitumor agent applicable to the treatment of hormone-refractory human prostate cancers.

Androgens↗
PubMed1998

12(S)-hydroxyeicosatetraenoic acid increases the actin microfilament content in B16a melanoma cells: a protein kinase-dependent process.

12(S)-hydroxyeicosatetraenoic acid [12(S)-HETE], a lipoxygenase metabolite of arachidonic acid, has been shown to be involved in a wide variety of cellular activities (i.e., adhesion, spreading, motility, invasion) which promote metastasis to occur in tumor cells. In this study, several techniques (Western blotting, flow cytometry and DNase I assay) were performed to examine the alterations in the distribution of G- and F-actin expressed in B16a melanoma cells. Each of these methods independently revealed that 12(S)-HETE treatment (0.1 mM, 15 min) resulted in an increase in the F-actin content in the cytoskeletal preparations. Since the integrity of cytoskeletal networks (i.e., actin filaments) can be dynamically regulated through protein phosphorylation, we investigated the potential role of several protein kinases in the 12(S)-HETE-induced actin polymerization. By flow cytometric analysis, 12(S)-HETE was found to increase the actin filament contents. This effect could be inhibited by protein kinase C (PKC) inhibitors (calphostin C and staurosporine) as well as by protein tyrosine kinase (PTK) inhibitor (genistein) but not by protein kinase A inhibitor (H8), suggesting that the 12(S)-HETE effect involves PKC and PTK. This conclusion is consistent with the observations that phorbol 12-myristate-13-acetate (PMA) mimics the biological effect of 12(S)-HETE in promoting the F-actin formation in B16a cells. As a final analysis, direct protein phosphorylation studies indicate that 12(S)-HETE treatment led to enhanced phosphorylation of myosin light chain, which may contribute to the increased stress fiber formation following 12(S)-HETE stimulation.

12-Hydroxy-5,8,10,14-eicosatetraenoic Acid↗
PubMed1998

Actin cleavage in various tumor cells is not a critical requirement for executing apoptosis.

Actin is a major cytoskeletal protein which is involved in many physiological cellular functions such as motility, cell shape, and adhesion. Recently, actin has also been reported to be cleaved by apoptotic proteases (i.e., caspases) and this cleavage is thought to contribute to the apoptotic process. However, conflicting data also exists as to whether actin represents a true caspase substrate during apoptosis induction in vivo (i.e., inside the cells). In this study, we critically examined the actin cleavage patterns during apoptosis of several tumor cell lines derived from three different species (i.e., mouse, rat, and human). Our findings demonstrate that: 1) actin cleavage in vivo is not a common phenomenon since apoptosis caused by multiple inducers in most cell types examined occurs without evidence of actin degradation; and 2) in certain cell types (e.g., U937), spontaneous, actin cleavage is observed which is not prevented by various specific chemical/peptide inhibitors of proteases such as caspases or serine proteases although apoptosis per se is retarded by some of these inhibitors. Our results conclude that actin is not a critical substrate for apoptotic proteases in vivo during apoptosis.

Actins↗
PubMed1998

A novel in vitro system to study extravasated tumor cell-induced angiogenesis.

Angiogenesis, the formation of new blood vessels from preexisting ones, is a fundamental stage in the metastatic pathway. For the primary tumor, this neovascularization provides nutrients and oxygen as well as a route by which metastatic tumor cells gain access to the circulatory system. Among these metastatic tumor cells, there are subgroups of cells that express an angiogenesis-inducing cells phenotype (AICs) as well as others that do not. Tumor cells not expressing the angiogenesis-inducing cells phenotype (non-AICs) invade new tissues and remain as dormant micrometastases unless they accompany AICs. Thus, either alone or with non-AICs, angiogenesis-inducing cells form rapidly growing, clinically detectable metastases. Much of the current research in this area is concentrated on the vascularization of primary tumors, but the regulation of angiogenesis by extravasating or invading tumor cells has not being extensively studied. We have developed a working model, which demonstrates that human metastatic prostate cancer cells (PC-3) appear to induce human vascular endothelial cells (HUVECs) to translocate across a Matrigel-coated 8 mm membrane. The parameters of this model (i.e. pore size, seeding-cell density, seeding times) were established using highly invasive murine melanoma cells (B16F10) seeded on murine microvascular endothelial cells (CD3). We have further modified our model in order to include a host compartment made of collagen gel, in order to mimic the in vivo site of metastases-induced angiogenesis.

Adenocarcinoma↗
PubMed1998

Present of a toxin in the salivary glands of the marine snail Cymatium intermedius that targets nicotinic acetylcholine receptors.

Presence of a toxin in the salivary glands of the marine snail Cymatium intermedius that targets nicotinic acetylcholine receptors. Toxicon 36, 25-29, 1998.-We present evidence of a neurotoxin from the salivary glands of Cymatium intermedius that displays acetylcholine-like effects on vertebrate (mouse ileum) and invertebrate (molluscan smooth muscle; molluscan heart; leech body wall) tissues. These effects were completely blocked by (+)-tubocurarine (10-100 muM) but not by atropine (up to 200 muM) suggesting that the toxin targets nicotinic-like acetylcholine receptors. This affirms the proposal that this genus may overcome their prey with a paralytic secretion.

Animals↗
PubMed1998

Sequence variations in alleles of the avirulence gene avrPphE.R2 from Pseudomonas syringae pv. phaseolicola lead to loss of recognition of the AvrPphE protein within bean cells and a gain in cultivar-specific virulence.

The bean halo blight pathogen, Pseudomonas syringae pv. phaseolicola (Psph), is differentiated into nine races based on the presence or absence of five avirulence (avr) genes in the bacterium, which interact with corresponding resistance genes. R1-R5, in Phaseolus vulgaris. The resistance gene R2 is matched by avrPphE, which is located adjacent to the cluster of hrp genes that are required for pathogenicity of Psph. Although only races 2, 4, 5 and 7 are avirulent on cultivars with R2 (inducing the hypersensitive response; HR), homologues of avrPphE are present in all races of Psph. DNA sequencing of avrPphE alleles from races of Psph has demonstrated two routes to virulence: via single basepair changes conferring amino acid substitutions in races 1, 3, 6 and 9 and an insertion of 104bp in the allele in race 8. We have demonstrated that these base changes are responsible for the difference between virulence and avirulence by generating transconjugants of a virulent race harbouring plasmids expressing the various alleles of avrPphE. Agrobacterium tumefaciens-directed expression of avrPphE from race 4 in bean leaves induced the HR in a resistance gene-specific manner, suggesting that the AvrPphE protein is alone required for HR induction and is recognized within the plant cell. The allele from race 6, which is inactive if expressed in Psph, elicited a weak HR if expressed in planta, whereas the allele from race 1 did not. Our results suggest that the affinity of interaction between AvrPphE homologues and an unknown plant receptor mediates the severity of the plant's response. Mutation of avrPphE alleles did not affect the ability to colonize bean from a low level of inoculum. The avirulence gene avrPphB, which matches the R3 resistance gene, also caused a gene-specific HR following expression in the plant after delivery by A. tumefaciens.

Alleles↗
PubMed1997

Suppression of W256 carcinosarcoma cell apoptosis by arachidonic acid and other polyunsaturated fatty acids.

Serum-cultured rat W256 carcinosarcoma cells of the monocytoid origin undergo rapid apoptosis in response to the lipoxygenase inhibitor NDGA (nordihydroguaiaretic acid). Exogenous arachidonic acid (AA), in a time- and dose-dependent fashion, suppressed NDGA-induced W256 cell apoptosis as well as DNA fragmentation, with the maximal effect observed at approximately 25 microM. Mobilization of endogenous AA by calcium ionophore A23187 provided an even stronger and longer-lasting protection against NDGA-caused cell death. The A23187 effect on AA release as well as W256 cell death can be blocked by bromophenacyl bromide, thus suggesting involvement of phospholipase A2 activation. Serum withdrawal similarly caused W256 cells to undergo typical apoptosis, which was not rescued by several growth factors commonly found in serum. However, exogenous AA suppressed serum starvation-induced W256 cell apoptosis and significantly extended cell survival in a dose-dependent manner. Lipoxygenase products, 12(S)- and 15(S)-, but not 5(S)-hydroxyeicosatetraenoic acid (HETE), in a dose-dependent fashion, also prevented both NDGA- and serum-starvation-induced W256 cell apoptosis. AA appears to suppress W256 cell apoptosis via distinct signaling pathway(s) since it does not prevent cell death triggered by several other inducers. Examination of a panel of polyunsaturated fatty acids revealed that alpha-linolenic and linoleic acid can also suppress NDGA-induced W256 cell apoptosis. Our data suggest that AA and other polyunsaturated fatty acids and/or their metabolites may enhance tumor growth not only by promoting cell proliferation but also by suppressing apoptosis.

12-Hydroxy-5,8,10,14-eicosatetraenoic Acid↗
PubMed1997

Nonlethal gamma radiation displays similar cytoplasmic rearrangements as 12(S)-HETE.

Timar et al. (Int. J. Canc., 52:594-603, 1992) and Onoda et al. (Rad. Res., 130:281-288, 1992) implicated 12(S)-HETE and nonlethal gamma radiation respectively, in enhanced expression of the alpha IIb beta 3 integrin receptor in B16a tumor cells. 12(S)-HETE has been reported to induce rearrangement of cytoplasmic elements and movement of cytoplasm (Haddad and Taylor, Proc. Amer. Cancer Res., 34:51, 1993); Timar et al., Cell Motil. Cytoskel., 26:49-65, 1993). Here, we report similar results in B16a tumor cells by post 90 cGy gamma radiation. Control cells were circular without cytoplasmic processes with the majority of their organelles and elements in perinuclear regions both surrounded by a ring of actin microfilaments. The cell's extensive leading edge was organelle free with the exception of occasional endoplasmic reticular cisternae parallel to one or two microtubules, and short bundles of actin microfilaments perpendicular to the plasma membrane. As early as 5 sec, but better defined by 60-90 sec, one end of elongated microfilaments bundles became associated with the plasma membrane while the other end of the bundles became associated with perinuclear mass protrusions. By 5 min, prominent by 15 min, protrusions, now separated from the leading edge by mixture of bundle and ring microfilaments, reached the plasma membrane to form cellular processes. By 30 min, some cytoplasmic masses showed signs of retraction along with the disappearance of their associated mixed microfilaments. As well as numerous vesicles of unknown origin, primarily associated with microtubules perpendicular to the plasma membrane, mitochondria and endoplasmic reticular cisternae were found in the cytoplasmic masses.

12-Hydroxy-5,8,10,14-eicosatetraenoic Acid↗