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Przysługujący widmom wywrotowy, niemalże rewolucyjny potencjał został odsunięty oraz zagubiony poprzez pozbycie się tej kontrowersyjnej kategorii z terytorium metafizyki i zbyt pośpieszne odesłanie jej do przestrzeni badań nad literaturą, sprowadzając ją jedynie do roli użytecznego, bezpiecznego narzędzia interpretacji.

Książka Andrzeja Marca odzyskuje niepokojący charakter widmontologii i ponownie wprowadza widma do myśli metafizycznej. Czym nawiedzona i przeniknięta jest tradycyjna metafizyka, co właściwie ją dręczy, nie daje spokoju oraz nieustannie podmywa jej stabilność?

Kategoria widma Derridy posłużyła Marcowi przede wszystkim do rozważań nad charakterem współczesności i przeobrażeń, jakie dokonały się w obrębie myśli ponowoczesnej. Współczesna kultura, zdaniem Marca, stała się miejscem powstawania widm, nieustannie powracających z przeszłości. Uznając, że nie jesteśmy już w stanie tworzyć utopijnych wizji przyszłości, autor stara się zdać sprawę z widmowego charakteru współczesnej kultury, analizując m. in. strategie vintage, retro, nurt no future w kulturze lat 70. czy nurt fantastyki naukowej w literaturze.

This report describes the generation of novel encapsidated RNA particles and their evaluation as in-tube internal controls in diagnostic real-time reverse-transcription PCR (rRT-PCR) assays for the detection of RNA viruses. A cassette containing sequences of 2 diagnostic primer sets for foot-and-mouth disease virus (FMDV) and a set for swine vesicular disease virus (SVDV) was engineered into a full-length cDNA clone containing the RNA-2 segment of Cowpea Mosaic Virus (CPMV). After co-inoculation with a plasmid that expressed CPMV RNA-1, recombinant virus particles were rescued from cowpea plants (Vigna unguiculata). RNA contained in these particles was amplified in diagnostic rRT-PCR assays used for detection of FMDV and SVDV. Amplification of these internal controls was used to confirm that rRT-PCR inhibitors were absent from clinical samples, thereby verifying negative assay results. The recombinant CPMVs did not reduce the analytical sensitivity of the rRT-PCRs when amplification of the insert was performed in the same tube as the diagnostic target. This system provides an attractive solution to the production of internal controls for rRT-PCR assays since CPMV grows to high yields in plants, the particles are thermostable, RNase resistant and simple purification of RNA-2 containing capsids yields a preparation which is non-infectious.

The cell-cell adhesion molecule 1 (C-CAM1) plays an important role as a tumor suppressor for prostate cancer. Decreased expression of C-CAM1 was detected in prostate, breast, and colon carcinoma. Reexpression of C-CAM1 in prostate and breast cancer cell lines was able to suppress tumorigenicity in vivo. These observations suggest that C-CAM1 may be used as a marker for cancer detection or diagnosis. To generate monoclonal antibodies specific to C-CAM1, we have overexpressed full-length human C-CAM1 in Sf9 cells using a baculovirus expression system. The protein was purified 104-fold using nickel affinity chromatography. About 0.4 mg purified C-CAM1 was obtained from 200 mg of infected cells. When the purified protein was digested with peptidyl-N-glycosidase, the apparent mobility of the protein on SDS-PAGE changed from 90 to 58 kDa, which is close to the molecular weight predicted from the cloned cDNA sequence. This observation suggests that C-CAM1 was glycosylated on asparagine residues when expressed in Sf9 cells. Western blotting and internal protein sequencing analysis confirmed that the purified protein is human C-CAM1. Biochemical and functional assays indicate that this protein expressed in Sf9 cells displays characteristics similar to those of native protein, including adhesion function and glycosylation modification. Using this protocol, sufficient quantity of this protein can be produced with purity suitable for monoclonal antibody generation and biochemical study.

Affective states, anxiety in particular, have been shown to negatively influence human postural control efficiency as measured by posturographic means, while exposure to a full-length mirror image of one's body exerts a stabilizing effect. We tested the hypothesis that body image concerns and preoccupations would relate negatively to this stabilising effect. Sixty-six healthy students, who screened negative for psychiatric disorders, completed rating scales for anxiety, depression and body image concerns. Posturography recordings of body sway were taken under three conditions: with eyes closed, looking at a vertical bar and looking at a full-length mirror. The Eyes Open/Mirror Stabilometric Quotient [EOMQ=(sway path with eyes closed/sway path looking at the mirror)x100], an index of how much postural control is stabilized by mirror feedback in comparison to the visual vertical bar condition, was significantly inversely related to body image concerns and preoccupations, and to trait anxiety. This finding confirms the impact of emotional factors on human postural control, which warrant further studies. If confirmed in clinical populations characterized by high levels of body image disturbances, e.g. eating disorders, it could lead to developments in the assessment and monitoring of these patients.

A recent species status investigation of the pathogenic fungus Paracoccidioides brasiliensis suggested the existence of three cryptic species. In the present study the sequences of the PRP8 intein from P. brasiliensis isolates belonging to the three described genetic groups and two unidentified isolates were determined and analyzed in order to check their functionality and usefulness for species identification. All the isolates presented a full-length intein, although the Endonuclease domain seems to be inactive due to substitutions in the second essential aspartic acid residue. Phylogenetic analysis by Maximum-Parsimony, Maximum Likelihood, and Bayesian Analysis clearly separated the isolates from the three species and revealed a significant difference between the Pb01 isolate and the remaining ones. The Pb01 isolate does not belong to any of the groups previously described since it presented a high divergence level compared to the three different genetic groups, corroborating some previous studies that suggested this isolate represents a new species of Paracoccidioides.

The Arabidopsis Nucleolar Protein Database (http://bioinf.scri.sari.ac.uk/cgi-bin/atnopdb/home) provides information on 217 proteins identified in a proteomic analysis of nucleoli isolated from Arabidopsis cell culture. The database is organized on the basis of the Arabidopsis gene identifier number. The information provided includes protein description, protein class, whether or not the plant protein has a homologue in the most recent human nucleolar proteome and the results of reciprocal BLAST analysis of the human proteome. In addition, for one-third of the 217 Arabidopsis nucleolar proteins, localization images are available from analysis of full-length cDNA–green fluorescent protein (GFP) fusions and the strength of signal in different parts of the cell—nucleolus, nucleolus-associated structures, nucleoplasm, nuclear bodies and extra-nuclear—is provided. For each protein, the most likely human and yeast orthologues, where identifiable through BLASTX analysis, are given with links to relevant information sources.

Pyrethroid resistance in human head louse populations is widespread in the United States and worldwide. We previously documented that the knockdown resistance of permethrin-resistant head louse populations is associated with the T929I and L932F (T917I and L920F in the numbering of the louse amino acid sequence) mutations in the voltage-sensitive sodium channel a-subunit gene. In order to identify additional sodium channel mutations potentially associated with knockdown resistance, we cloned and sequenced full-length cDNA fragments from insecticide-susceptible (Ecuador) and permethrin-resistant (Florida) head louse populations and from an insecticide-susceptible body louse population (Israel). Sequence comparisons of the complete open reading frames of the sodium channel genes identified one additional novel mutation (M815I), which was located in the IIS1-2 extracellular loop of the a-subunit, from the permethrin-resistant head louse population. Absolute conservation of the Met815 residue at the corresponding positions within sodium channels from all known susceptible populations of insect species implied that the M815I mutation likely has a functional significance in resistance. Sequence analyses of cloned cDNA fragments and genomic DNA fragments from individual louse samples, both containing the three mutation sites, confirmed that all the mutations exist en bloc as a haplotype. Northern blot analysis identified a single 7.2 kb transcript. The comparison of complete open reading frame sequences (6156 bp) of sodium channel gene between head and body lice revealed 26 polymorphic nucleotides, of which only one resulted in a conservative amino acid substitution (glutamic versus aspartic acid at 11th amino acid position). The virtual identity in nucleotide sequences indicated that both body and head lice are conspecific, and lends justification of the use of the body louse as a surrogate organism for the head louse in bio-Pesticide Biochemistry and Physiology 75 79-91 www.elsevier.com/locate/ypest * Corresponding

A hybrid approach involving synthetic DNA, fusion PCR, and ectopic expression has been used to genetically manipulate the expression of the D1 protein of photosystem II (PSII) in the model cyanobacterium Synechocystis sp. PCC6803. Due to the toxicity of the full-length psbA gene in E. coli, a chimeric psbA2 gene locus was commercially synthesised and cloned in two halves. High-fidelity fusion PCR utilizing sequence overlap between the two synthetic gene halves allowed the production of a DNA fragment that was able to recombine the full-length psbA2 gene into the Synechocystis chromosome at an ectopic (non-native) location. This was accomplished by designing the synthetic DNA/fusion PCR product to have the psbA2 gene, with control sequences, interposed between chimeric sequences corresponding to an ectopic target chromosomal location. Additionally, a recipient strain of Synechocystis lacking all three psbA genes was produced by a combination of traditional marker replacement and markerless replacement techniques. Transformation of this multiple deletion strain by the synthetic DNA/fusion PCR product faithfully restored D1 expression in terms of its expression and PSII repair capacity. The advantages and potential issues for using this approach to rapidly introduce chimeric sequence characteristics as a general tool to produce novel genetic constructs are discussed.

Growing evidence suggests that oxidative stress is involved in the neuronal degeneration and can promote the aggregation of α-synuclein. However, the role of α-synuclein under physiological and pathological conditions remains poorly understood. In the present study, we examined the possible interaction between the α-synuclein and oxidative stress. In a dopaminergic cell line MES23.5, we have found that the 200μM H2O2

The infectious and diagnostic stage of Giardia lamblia (also known as G. intestinalis or G. duodenalis) is the cyst. The Giardia cyst wall contains fibrils of a unique b-1,3-linked N-acetylgalactosamine (GalNAc) homopolymer and at least three cyst wall proteins (CWPs) composed of Leu-rich repeats (CWP LRR) and a C-terminal conserved Cys-rich region (CWP CRR). Our goals were to dissect the structure of the cyst wall and determine how it is disrupted during excystation. The intact Giardia cyst wall is thin (,400 nm), easily fractured by sonication, and impermeable to small molecules. Curled fibrils of the GalNAc homopolymer are restricted to a narrow plane and are coated with linear arrays of oval-shaped protein complex. In contrast, cyst walls of Giardia treated with hot alkali to deproteinate fibrils of the GalNAc homopolymer are thick (,1.2 mm), resistant to sonication, and permeable. The deproteinated GalNAc homopolymer, which forms a loose lattice of curled fibrils, is bound by native CWP1 and CWP2, as well as by maltose-binding protein (MBP)-fusions containing the full-length CWP1 or CWP1 LRR. In contrast, neither MBP alone nor MBP fused to CWP1 CRR bind to the GalNAc homopolymer. Recombinant CWP1 binds to the GalNAc homopolymer within secretory vesicles of Giardia encysting in vitro. Fibrils of the GalNAc homopolymer are exposed during excystation or by treatment of heat-killed cysts with chymotrypsin, while deproteinated fibrils of the GalNAc homopolymer are degraded by extracts of Giardia cysts but not trophozoites. These results show the Leu-rich repeat domain of CWP1 is a lectin that binds to curled fibrils of the GalNAc homopolymer. During excystation, host and Giardia proteases appear to degrade bound CWPs, exposing fibrils of the GalNAc homopolymer that are digested by a stage-specific glycohydrolase.

Ferritin, a cytosolic iron storage protein composed of 24 subunits of heavy chain and light chain, is an intracellular protein primarily involved in iron metabolism. It can sequester up to 4500 ferric ions in its inner core to protect cells against toxicity of iron. Ferritin is known to play important roles in detoxification and is also involved in immunity processes. In this study, a full-length ferritin cDNA was cloned from the haemocyte of the Pacific white shrimp, Litopenaeus vannamei: it comprises 1249 bp, including 132 bp in the 5 0 -untranslated region, 510 bp in the open reading frame which encodes 170 amino acid residues, and 607 bp in the 3 0untranslated region. Alignments of the deduced amino acid sequence showed that the Pacific white shrimp ferritin shares 74%, 69%, 62%, 67%, 50% and 48% identity with crayfish, tick, brine shrimp, oyster, human and rat, respectively. The tissue-specific expression pattern was examined by reverse transcription polymerase chain reaction and real-time quantitative PCR. The ferritin mRNA is expressed in various tissues of the shrimp in the order of haemocyte, midgut gland, brain ganglion, gill, hepatopancreas, abdominal ganglion, eyestalk, muscle, thoracic ganglion, and heart.

Humor is a uniquely human quality whose neural substrates remain enigmatic. The present report combined dynamic, real-life content and event-related functional magnetic resonance imaging (fMRI) to dissociate humor detection (''getting the joke'') from humor appreciation (the affective experience of mirth). During scanning, subjects viewed full-length episodes of the television sitcoms Seinfeld or The Simpsons. Brain activity time-locked to humor detection moments revealed increases in left inferior frontal and posterior temporal cortices, whereas brain activity time-locked to moments of humor appreciation revealed increases in bilateral regions of insular cortex and the amygdala. These findings provide evidence that humor depends critically upon extant neural systems important for resolving incongruities (humor detection) and for the expression of affect (humor appreciation).

Amelogenin, the predominant matrix protein in developing dental enamel, has been considered essential for proper enamel formation . Amelogenin self-assembles into nanospheres that serve as a scaffold for the oriented elongation of enamel crystals. As enamel matures, amelogenin proteins are progressively processed by proteinases, including matrix metallopro-

The laccase gene lac48424-1 and its corresponding full-length cDNA were cloned and characterized from a novel white-rot fungi Trametes sp. 48424 which had the high yield of laccase and strong ability for decolorizing different dyes. The 1563 bp full-length cDNA of lac48424-1 encoded a mature laccase protein containing 499 amino acids preceded by a signal peptide of 21 amino acids. The deduced protein sequence of LAC48424-1 showed high similarity with other known fungal laccases and contained four copper-binding conserved domains of typical laccase protein. The functionality of lac48424-1 gene encoding active laccase was verified by expressing the gene in the yeast Pichia pastoris successfully. It was found that the recombinant laccase produced by the yeast transformant could decolorize different dyes. The 5 0 -flanking sequence upstream of start codon was obtained by Self-Formed Adaptor PCR. Many putative cis-acting responsive elements involved in the transcriptional regulation were identified in the promoter region of lac48424-1.

Pichia pastoris strain GS115, using the Saccharomyces cerevisiae Mat ␣ signal peptide. In both cases, a high level of the secreted protein could be detected by SDS-PAGE. The truncated gene was also expressed using the human gelatinase B native signal peptide. Secretion using the Mat ␣ signal peptide was significantly greater than that from the native signal peptide. The recombinant products were purified and characterized biochemically. The recombinant proteins, FLGelB and dGelB, were found to have similar biochemical properties and activity to that of the human gelatinase B native protein.

Although it is known that deletions or mutations of the SMN1 gene on chromosome 5 cause decreased levels of the SMN protein in subjects with proximal autosomal recessive spinal muscular atrophy (SMA), the exact sequence of pathological events leading to selective motoneuron cell death is not fully understood yet. In this review, new Wndings regarding the dual cellular role of the SMN protein (translocation of -actin to axonal growth cones and snRNP biogenesis/pre-mRNA splicing) were integrated with recent data obtained by detailed neuropathological examination of SMA and control subjects. A presumptive series of 10 pathogenetic events for SMA is proposed as follows: (1) deletions or mutations of the SMN1 gene, (2) increased SMN mRNA decay and reduction in full-length functional SMN protein, (3) impaired motoneuron axonoand dendrogenesis, (4) failure of motoneurons to form synapses with corticospinal Wbers from upper motoneurons, (5) abnormal motoneuron migration towards ventral spinal roots, (6) inappropriate persistence of motoneuron apoptosis due to impaired diVerentiation and motoneuron displacement, (7) substantial numbers of motoneurons continuing to migrate abnormally ("heterotopic motoneurons") and entering into the ventral roots, (8) attracted glial cells following these heterotopic motoneurons, which form the glial bundles of ventral roots, (9) impaired axonal transport of actin, causing remaining motoneurons to become chromatolytic, and (10) eventual death of all apoptotic, heterotopic and chromatolytic neurons, with apoptosis being more rapid and predominating in the earlier stages, with death of heterotopic and chromatolytic neurons occurring more slowly by necrosis during the later stages of SMA. According to this model, the motoneuron axonopathy is more important for pathogenesis than the ubiquitous nuclear splicing deWcit. It is also supposed that individually variable levels of SMN protein, together with inXuences of other phenotype modiWer genes and their products, cause the clinical SMA spectrum through diVerential degree of motoneuron functional loss.

Background: Occult or latent hepatitis B virus (HBV) infection is defined as infection with detectable HBV DNA and undetectable surface antigen (HBsAg) in patients' blood. The cause of an overt HBV infection becoming an occult one is unknown. To gain insight into the mechanism of the development of occult infection, we compared the full-length HBV genome from a blood donor carrying an occult infection (d4) with global genotype D genomes. The phylogenetic analysis of polymerase, core and X protein sequences did not distinguish d4 from other genotype D strains. Yet, d4 surface protein formed the evolutionary outgroup relative to all other genotype D strains. Its evolutionary branch was the only one where accumulation of substitutions suggests positive selection (dN/dS = 1.3787). Many of these substitutiions accumulated specifically in regions encoding the core/surface protein interface, as revealed in a 3D-modeled protein complex. We identified a novel RNA splicing event (deleting nucleotides 2986-202) that abolishes surface protein gene expression without affecting polymerase, core and X-protein related functions. Genotype D strains differ in their ability to perform this 2986-202 splicing. Strains prone to 2986-202 splicing constitute a separate clade in a phylogenetic tree of genotype D HBVs. A single substitution (G173T) that is associated with clade membership alters the local RNA secondary structure and is proposed to affect splicing efficiency at the 202 acceptor site. We propose an evolutionary scenario for occult HBV infection, in which 2986-202 splicing generates intracellular virus particles devoid of surface protein, which subsequently accumulates mutations due to relaxation of coding constraints. Such viruses are deficient of autonomous propagation and cannot leave the host cell until it is lysed. Occult HBV infections are defined as the presence of HBV DNA and the absence of HBV surface antigen (HBsAg encoded by the S gene) in plasma or serum of HBVinfected patients . This infection may persist in individuals for years without emerging symptoms of overt HBV

Caenorhabditis elegans should soon be the first multicellular organism whose complete genomic sequence has been determined. This achievement provides a unique opportunity for a comprehensive assessment of the signal transduction molecules required for the existence of a multicellular animal. Although the worm C. elegans may not much resemble humans, the molecules that regulate signal transduction in these two organisms prove to be quite similar. We focus here on the content and diversity of protein kinases present in worms, together with an assessment of other classes of proteins that regulate protein phosphorylation. By systematic analysis of the 19,099 predicted C. elegans proteins, and thorough analysis of the finished and unfinished genomic sequences, we have identified 411 full length protein kinases and 21 partial kinase fragments. We also describe 82 additional proteins that are predicted to be structurally similar to conventional protein kinases even though they share minimal primary sequence identity. Finally, the richness of phosphorylation-dependent signaling pathways in worms is further supported with the identification of 185 protein phosphatases and 128 phosphoprotein-binding domains (SH2, PTB, STYX, SBF, 14-3-3, FHA, and WW) in the worm genome.

A-Synuclein, a protein implicated in neurodegenerative diseases and of elusive physiological function owes its name to an observed presence in presynaptic and nuclear compartments. However, its nuclear localisation has remained controversial. We expressed synuclein-eGFP fusion proteins in organotypic rat hippocampal slice cultures and murine hippocampal primary neurons using a Sindbis virus expression system. Recombinant full-length A-synuclein accumulated in presynaptic locations, mimicking its native distribution. Expression of deletion mutant A-synuclein revealed that presynaptic targeting depended on the presence of its N-terminal and core region. This domain also causes nuclear exclusion of the Asynuclein fusion protein. In contrast, the C-terminal domain of Asynuclein directs fusion proteins into the nuclear compartment. The related protein ;-synuclein contains a similar N-terminal and core domain as A-synuclein. However, ;-synuclein lacks a C-terminal domain that causes nuclear localisation of the fusion protein, suggesting that the two synucleins might have different roles relating to the cell nucleus.

CWR22 has been a valuable xenograft model for the study of prostate cancer progression from an androgen-dependent tumor to one that grows in castrated animals. Herein, we report the identification and characterization of a novel androgen receptor (AR) mutation occurring in a relapsed tumor (CWR22R-2152) and in the CWR22Rv1 cell line established from it. The mutation was not detected in the original, hormone-dependent CWR22 xenograft, indicating that this change occurred during the progression to androgen independence. It is characterized by an in-frame tandem duplication of exon 3 that encodes the second zinc finger of the AR DNA-binding domain. Accordingly, immunoblot analyses demonstrated the expression of an AR species having an approximately 5-kDa increase in size relative to the LNCaP AR. This was accompanied by a COOH-terminally truncated AR species migrating with a relative mass of 75-80 kDa, referred to as ARDeltaLBD because it lacks the ligand-binding domain. By recreating ...

Since the early 1980s, it has been known that high Prostate-specific antigen (PSA) is a widely used serum prostate-specific antigen (PSA) 2 levels are well marker for screening and monitoring prostate cancer. correlated with the presence of prostate cancer (1, 2). Because PSA levels are normally quite low, an anti-By using simple immunometric assays it is now possible body-based assay must be used to detect PSA. However, to quantitate serum PSA levels in less than an hour (3, not all PSA-specific antibodies bind equally well to PSA 4). However, current problems with many commercial or to its different isoforms. Therefore, a better undertests point to the lack of understanding regarding how standing of how PSA interacts with PSA-specific anti-

Environmental microbiology studies commonly use terminal restriction fragment length polymorphism (T-RFLP) of 16S rRNA genes, for example, to analyze changes in community structure in relation to changing physicochemical and biological conditions over space and time. Although T-RFLP is most useful for comparing samples from different environments, a large number of samples makes effective analysis difficult using the Web-based tools that are currently available. To resolve this dilemma, we used a new approach for calculating data from multiple T-RFLP samples by estimating terminal fragment combinations, then applying a correlation analysis using two different fluorescent dyes generated simultaneously from all samples. This calculation was based on the expectation that the proportions of two terminal fragments from one full-length polymerase chain reaction fragment would be nearly the same in each analysis. Using this program, the oral microflora in 73 human saliva samples were analyzed, and 24 bacterial groups, with peak areas of at least 0.5% and correlation coefficients of 0.55 or greater, were identified from the T-RFs within 40 s.

AbstractÐThis paper deals with the computation of reciprocals, square roots, inverse square roots, and some elementary functions using small tables, small multipliers, and, for some functions, a final ªlargeº (almost full-length) multiplication. We propose a method, based on argument reduction and series expansion, that allows fast evaluation of these functions in high precision. The strength of this method is that the same scheme allows the computation of all these functions. We estimate the delay, the size/number of tables, and the size/number of multipliers and compare with other related methods.

Plasmodium falciparum is the most prevalent and lethal of the malaria parasites infecting humans, yet the origin and evolutionary history of this important pathogen remain controversial. Here we develop a single-genome amplification strategy to identify and characterize Plasmodium spp. DNA sequences in faecal samples from wild-living apes. Among nearly 3,000 specimens collected from field sites throughout central Africa, we found Plasmodium infection in chimpanzees (Pan troglodytes) and western gorillas (Gorilla gorilla), but not in eastern gorillas (Gorilla beringei) or bonobos (Pan paniscus). Ape plasmodial infections were highly prevalent, widely distributed and almost always made up of mixed parasite species. Analysis of more than 1,100 mitochondrial, apicoplast and nuclear gene sequences from chimpanzees and gorillas revealed that 99% grouped within one of six host-specific lineages representing distinct Plasmodium species within the subgenus Laverania. One of these from wester...

Transgenic tomato resistant to tomato leaf curl disease (ToLCD) using replicase (rep) gene sequences of Tomato leaf curl virus in antisense orientation were developed via Agrobacterium-mediated transformation. A binary vector carrying the antisense rep gene (untranslatable full length sequence, 1086 bp) along with the npt II gene was used for transformation. High level of resistance and inheritability of the transgene was observed up to T 2 stage following challenge inoculation with the virus. The mechanism of resistance appears RNA-mediated, since the plants carried the untranslatable antisense rep gene. Progeny analysis of these plants showed classical Mendelian pattern of inheritance in two of the six transgenic lines having single transgene insertion.

Pathogenesis-related (PR) proteins are plant proteins that are induced in response to pathogen attack. PR proteins are grouped into independent families based on their sequences and properties. The PR-4 family comprises class I and class II chitinases. We have isolated a full-length cDNA encoding a chitinase from maize which shares a high degree of nucleotide and amino acid sequence homology with the class II chitinases of the PR-4 family of PR proteins. Our results indicate that fungal infection, and treatment either with fungal elicitors or with moniliformin, a mycotoxin produced by the fungus Fusarium moniliforme, increase the level of ZmPR4 mRNA. In situ mRNA hybridization analysis in sections obtained from fungus-infected germinating embryos revealed that ZmPR4 mRNA accumulation occurs in those cell types that first establish contact with the pathogen. ZmPR4 mRNA accumulation is also stimulated by treatment with silver nitrate whereas the application of the hormones gibberellic acid or acetylsalicylic acid has no effect. Wounding, or treatment with abscisic acid or methyl jasmonate, results in accumulation of ZmPR4 mRNA in maize leaves. Furthermore, the ZmPR4 protein was expressed in Escherichia coli, purified and used to obtain polyclonal antibodies that specifically recognized ZmPR4 in protein extracts from fungus-infected embryos. Accumulation of ZmPR4 mRNA in fungus-infected maize tissues was accompanied by a significant accumulation of the corresponding protein. The possible implications of these findings as part of the general defence response of maize plants against pathogens are discussed.

Theileria parva is the causative agent of Corridor disease in cattle in South Africa. The African buffalo (Syncerus caffer) is the reservoir host, and, as these animals are important for eco-tourism in South Africa, it is compulsory to test and certify them disease free prior to translocation. A T. parva-specific real-time polymerase chain reaction (PCR) test based on the small subunit ribosomal RNA (18S rRNA) gene is one of the tests used for the diagnosis of the parasite in buffalo and cattle in South Africa. However, because of the high similarity between the 18S rRNA gene sequences of T. parva and Theileria sp. (buffalo), the latter is also amplified by the real-time PCR primers, although it is not detected by the T. parva-specific hybridization probes. Preliminary sequencing studies have revealed a small number of sequence differences within the 18S rRNA gene in both species but the extent of this sequence variation is unknown. The aim of the current study was to sequence the 18S rRNA genes of T. parva and Theileria sp. (buffalo), and to determine whether all identified genotypes can be correctly detected by the real-time PCR assay. The reverse line blot (RLB) hybridization assay was used to identify T. parva and Theileria sp. (buffalo) positive samples from buffalo blood samples originating from the Kruger National Park, Hluhluwe-iMfolozi Park, the Greater Limpopo Transfrontier Park, and a private game ranch in the Hoedspruit area. Theileria parva and Theileria sp. (buffalo) were identified in 42 % and 28 %, respectively, of 252 samples, mainly as mixed infections. The full-length 18S rRNA gene of selected samples was amplified, cloned and sequenced. From a total of 20 sequences obtained, 10 grouped with previously published T. parva sequences from GenBank while 10 sequences grouped with a previously published Theileria sp. (buffalo) sequence. All these formed a monophyletic group with known pathogenic Theileria species. Our phylogenetic analyses confirm the distinction between Theileria sp. (buffalo) and T. parva and indicate the existence of a single group of T. parva and two Theileria sp. (buffalo) 18S rRNA gene variants in the African buffalo. Despite the observed variation in the full-length parasite 18S rRNA gene sequences, the area in the V4 hypervariable 3 region where the RLB and real-time PCR hybridization probes were developed was relatively conserved. The T. parva specific real-time PCR assay was able to successfully detect all T. parva variants and, although amplicons were obtained from Theileria sp. (buffalo) DNA, none of the Theileria sp. (buffalo) 18S rRNA sequence variants were detected by the T. parva-specific hybridization probes.

PCR-RFLP with nine restriction enzymes was applied to the 165 and 235 rRNA genes of 42 rhizobial and agrobacterial strains to determine the phylogenetic position of Rhizobium galegae and increase the understanding of the evolution of ribosomal operons. The strains were selected based on previous phylogenetic studies. PCR primers were designed so that they amplified a 2.3 kb fragment of the 23s rRNA gene (excluding the B8 loop). Universal primers rD1 and fD1 were used to amplify the full-length 165 rRNA. The RFLP analysis resulted in 27 and 32 different restriction patterns for 165 and 235, respectively. The RFLP patterns were transformed to genetic distances and dendrograms were constructed from the data using the unweighted pair group method with averages. The shapes of the dendrograms derived from the analysis of the 165 and 235 rRNA genes correlated well, with only a f e w strains having different positions. The 235 tree generally had deeper branching than the 165 tree, allowing better discrimination between species and strains. The combined data from the two analyses described 36 genotypes. The eight R. galegae strains formed a homogeneous cluster in all dendrograms. The RFLP analysis was confirmed by partial sequence analysis of the 165 rRNA gene (the first 800 bp), which correlated well with full-length 165 rRNA sequence analysis. The 165 data placed R. galegae near the genus Agrobacterium with Agrobacterium witis as its nearest neighbour, whereas in the 235 and the combined dendrograms it showed closer affinity to the genus Rhizobium. I Keywords: rhizobia, Rhizobium galegae, PCR-RFLP, 16S/23S rDNA, phylogeny I 94 92 90 Rhizobrum sp.(Acacia senegal) HAMBII 932 A. tumefaciens C58 Rhizobium sp. (Asfragalus sinicus) DUS551 Rhizobium sp.(Brownea ariza) MP13W4 A rubi ATCCI 3335 Rhizobium sp. (Glycynhiza pallidifbra) SH19352 A radiobacfer ATCC4718 A. furnefaciens B6 Rhizobium sp. (Leucaena sp.) INPA45R A. radiobacfer ATCCl9358 R. lropici CIAT899 A. rhizogenes ATCCll325 A. vitis LMG8750 A. vilis AG61 Rhizobium sp. (Camgana microphylla) SH215214 S. ferangae ORS1009 Rhizobium sp. (Acacia senegal) HAMBll705 R. leguminosarum bv. viceae ATCC1004 S. meliloti NZP4027 R. galegae (Galega orientalis) HAMB1540 (3) R. ga/egae (Galega officinalis) HAMBI1141 (4) R. galegae (Galega orientalis) HAMB11428 Rhizobium sp. (Hedysarum coronarium) MAP835 Rhizobium sp. (Caragana pruinosa) SH21321 Rhizobium sp. (Asfragalus cornplanatus) JL84 Rhizobium sp. (Leucaena leucocephala) MP13031 Rhizobium sp. (Sophora viicifolia) SH1124 M. ciceri UPM-Ca7 M. huakuii CCBAU2609 M. loti NZP2037 M. loti NZP2213 Rhizobium sp. ClusterU ORSlOOl 100 98 96 8 9 Rhizobium sp. (Brownea ariza) MPI 3004 Rhizobium sp. (Centrosema plumier!) MPI 3008 A. rubi ATCC 13335 Rhizobium sp. (Asfragalus siniCUs) DUS 551 Rhizobium sp. (Acacia senegal) HAMBI 1625 (3) Rhizobium sp. (Glycyrrhiza pallidiflora) SH19352 A. radiobacfer ATCC 471 8 Rhizobium sp. (Glycyrrhiza pallidiflora) SH19312 A. tumefaciens B6 A radiobacter ATCC19358 Rhuobium sp. (Acacia senegal) HAMBI 1932 A. fumefaciens C58 A. vifis AG 61 A. vitis LMG 0750 R. tropici ClAT 899 A. rhimgenes ATCCll325 R. leguminosarum ATCC 1004 Rhizobium sp. (Acacia senegal) HAMBH 705 R. galegae (Galega officinalislorient) (5) R. gdegae (Gakga orientalis) HAMB11461 R. galegae (Galega orientalis) HAMB11428 R. galegae (Galega oflicinalis) HAMBI1141 Rhizobium sp. (Sophora viicifolia) SHll24 Rhizobium sp. (Leucaena lecocephala) MP13031 S. meliloti NZP4027 Rhizobiurn sp. (Hedysarum coronarium) IMAP835 Rhhobium sp. (Astragalus complanatus) JL84 Rhizobium sp. (Camgana pruinosa) SH21321 S. terangae ORS1009 Rhizobium sp. (Leucaena sp.) INPA45R Rhizobium sp. (Camgana microphylla) SH215214 M. huakuii CCBAU2609 M. loti NZP2037 M. cicen UPM-Ca7 M. loti NZP2213

Chronic sleep disorders affect more than 50 million people in the US, and there are numerous clinical populations, such as spinal cord injury patients, cognitively-impaired elderly persons living at home, and infants at risk for Sudden Infant Death Syndrome, who could ...

During mammalian brain development, neurons are generated along the ventricle, migrate radially, and become aligned in defined patterns. These precise patterns of neuronal alignment are regulated by an extracellular matrix protein Reelin, and binding of Reelin to its receptors induces tyrosine phosphorylation of the intracellular adaptor protein disabled 1 (Dab1). We recently reported that Reelin molecules assemble to form a homomeric protein complex. Although the number of molecules in the full-length complex is unknown, recombinant N-terminal fragments, which contain the epitope for the function-blocking CR-50 antibody, assembled to form a complex of more than 40 monomers. When the N-terminus was deleted from Reelin, the truncated protein did not form a stable complex. To further characterize the Reelin assembly, we performed biochemical analysis of the full-length Reelin assembly in this study. Here, we report that a full-length Reelin forms a disulfide-linked homodimer. A chemic...

Classical heme-containing plant peroxidases have been ascribed a wide variety of functional roles related to development, defense, lignification, and hormonal signaling. More than 40 peroxidase genes are now known in Arabidopsis thaliana for which functional association is complicated by a general lack of peroxidase substrate specificity. Computational analysis was performed on 30 near full-length Arabidopsis peroxidase cDNAs for annotation of start codons and signal peptide cleavage sites. A compositional analysis revealed that 23 of the 30 peroxidase cDNAs have 5P untranslated regions containing 40^71% adenine, a rare feature observed also in cDNAs which predominantly encode stress-induced proteins, and which may indicate translational regulation.

Although acute bouts of exercise have consistently been found to enhance self-efficacy, recent evidence suggests that the exercise environment may moderate this relation. The purpose of this study was to investigate the influence of an acute bout of exercise performed in environments differing in self-evaluative potential on self-efficacy. Forty-three female undergraduates performed 20 min of aerobic exercise in front of and in the absence of a full-length mirror. Self-efficacy was assessed at 4 time points: before entry into the experimental condition, pre-exercise, postexercise, and 20 min postexercise. Results revealed that self-efficacy did not significantly change following exercise or recovery in the no mirror condition. However, the mirror condition resulted in moderate increases in self-efficacy postexercise and large increases following recovery. Results indicate that self-evaluation following exercise may increase one's perceptions of capabilities and appear to be in contrast to past research demonstrating a negative effect of mirrors on self-efficacy.

During a viewing of Disney's animated film Cinderella (1950), one author (AI) noticed a dog having nightmares with dreamenactment that strongly resembled RBD. This prompted a study in which all Disney classic full-length animated films and shorts were analyzed for other examples of RBD. Three additional dogs were found with presumed RBD in the classics films Lady and the Tramp (1955) and The Fox and the Hound (1981), and in the short Pluto's Judgment Day (1935). These dogs were elderly males who would pant, whine, snuffle, howl, laugh, paddle, kick, and propel themselves while dreaming that they were chasing someone or running away. In Lady and the Tramp the dog was also losing both his sense of smell and his memory, two associated features of human RBD. These four films were released before RBD was first formally described in humans and dogs. In addition, systematic viewing of the Disney films identified a broad range of sleep disorders, including nightmares, sleepwalking, sleep related seizures, disruptive snoring, excessive daytime sleepiness, insomnia and circadian rhythm sleep disorder. These sleep disorders were inserted as comic elements. The inclusion of a broad range of accurately depicted sleep disorders in these films indicates that the Disney screenwriters were astute observers of sleep and its disorders.

We report the cloning of a gene and the characterization of the encoded protein, which is released by the teratocytes of the parasitoid Aphidius ervi in the haemocoel of the host aphid Acyrthosiphon pisum. The studied protein was identified by LC-MS/MS, and the gathered information used for isolating the full length cDNA. The corresponding gene was made of 3 exons and 2 introns, and was highly expressed in the adult wasps and in parasitized hosts. The translation product, which was named Ae-ENO, showed a very high level of sequence identity with insect enolases. In vivo immunodetection experiments evidenced Ae-ENO localization in round spots, present in the teratocytes and released in the host haemocoel. Moreover, strong immunoreactivity was detected on the surface of A. ervi larvae and of host embryos. Ae-ENO expressed in insect cells was not secreted in the medium, indicating the occurrence in the teratocytes of an unknown pathway for Ae-ENO release. The recombinant protein produced in bacteria under native conditions was a dimer, with evident enolase activity (K m ¼ 0.086 AE 0.017 mM). Enolase is a well known enzyme in cell metabolism, which, however, is associated with a multifunctional role in disease, when present in the extracellular environment, on the surface of prokaryotic and eukaryotic cells. In these cases, the enolase mediates the activation of enzymes involved in the invasion of tissues by pathogens and tumour cells, and in the evasion of host immune response. The possible role played by Ae-ENO in the host regulation process is discussed in the light of this information.