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YeiL, the third member of the CRP-FNR family in Escherichia coli.

The yeiL open reading frame located at 48.5 min (2254 kb) in the nfo-fruA region of the Escherichia coli chromosome was predicted to encode a CRP and FNR paralogue capable of forming inter- or intra-molecular disulphide bonds and incorporating one iron-sulphur centre per 25 kDa subunit. Purified MBP-YeiL (a maltose-binding-protein-YeiL fusion protein) was a high-molecular-mass oligomer or aggregate which released unstable monomers (68 kDa) under reducing conditions. The MBP-YeiL protein contained substoichiometric amounts of iron and acid-labile sulphide, and an average of one disulphide bond per monomer. The iron and sulphide contents increased consistent with the acquisition of one [4Fe-4S] cluster per monomer after anaerobic NifS-catalysed reconstitution. By analogy with FNR and FLP (the FNR-like protein of Lactobacillus casei) it was suggested that the transcription-regulatory activity of YeiL might be modulated by a sensory iron-sulphur cluster and/or by reversible disulphide bond formation. A yeiL-lacZ transcriptional fusion showed that aerobic yeiL expression increases at least sixfold during stationary phase, requires RpoS, and is positively autoregulated by YeiL, positively activated by Lrp (and IHF in the absence of FNR) and negatively regulated by FNR. A regulatory link between the synthesis of YeiK (a potential nucleoside hydrolase) and YeiL was inferred by showing that the yeiK and yeiL genes are divergently transcribed from overlapping promoters. A 10-15% deficiency in aerobic growth yield and an enhanced loss of viability under nitrogen starvation conditions were detected with a yeiL::kan(R) mutant, suggesting that YeiL might function as a post-exponential-phase nitrogen-starvation regulator.

ATP-Binding Cassette Transporters↗

Search for Bacillus anthracis potential vaccine candidates by a functional genomic-serologic screen.

Bacillus anthracis proteins that possess antigenic properties and are able to evoke an immune response were identified by a reductive genomic-serologic screen of a set of in silico-preselected open reading frames (ORFs). The screen included in vitro expression of the selected ORFs by coupled transcription and translation of linear PCR-generated DNA fragments, followed by immunoprecipitation with antisera from B. anthracis-infected animals. Of the 197 selected ORFs, 161 were chromosomal and 36 were on plasmids pXO1 and pXO2, and 138 of the 197 ORFs had putative functional annotations (known ORFs) and 59 had no assigned functions (unknown ORFs). A total of 129 of the known ORFs (93%) could be expressed, whereas only 38 (64%) of the unknown ORFs were successfully expressed. All 167 expressed polypeptides were subjected to immunoprecipitation with the anti-B. anthracis antisera, which revealed 52 seroreactive immunogens, only 1 of which was encoded by an unknown ORF. The high percentage of seroreactive ORFs among the functionally annotated ORFs (37%; 51/129) attests to the predictive value of the bioinformatic strategy used for vaccine candidate selection. Furthermore, the experimental findings suggest that surface-anchored proteins and adhesins or transporters, such as cell wall hydrolases, proteins involved in iron acquisition, and amino acid and oligopeptide transporters, have great potential to be immunogenic. Most of the seroreactive ORFs that were tested as DNA vaccines indeed appeared to induce a humoral response in mice. We list more than 30 novel B. anthracis immunoreactive virulence-related proteins which could be useful in diagnosis, pathogenesis studies, and future anthrax vaccine development.

Animals↗

Neurolinguistics: structural plasticity in the bilingual brain.

Humans have a unique ability to learn more than one language--a skill that is thought to be mediated by functional (rather than structural) plastic changes in the brain. Here we show that learning a second language increases the density of grey matter in the left inferior parietal cortex and that the degree of structural reorganization in this region is modulated by the proficiency attained and the age at acquisition. This relation between grey-matter density and performance may represent a general principle of brain organization.

Age Factors↗

Epigenetic regulation of translation reveals hidden genetic variation to produce complex traits.

Phenotypic plasticity and the exposure of hidden genetic variation both affect the survival and evolution of new traits, but their contributing molecular mechanisms are largely unknown. A single factor, the yeast prion [PSI(+)], may exert a profound effect on both. [PSI(+)] is a conserved, protein-based genetic element that is formed by a change in the conformation and function of the translation termination factor Sup35p, and is transmitted from mother to progeny. Curing cells of [PSI(+)] alters their survival in different growth conditions and produces a spectrum of phenotypes in different genetic backgrounds. Here we show, by examining three plausible explanations for this phenotypic diversity, that all traits tested involved [PSI(+)]-mediated read-through of nonsense codons. Notably, the phenotypes analysed were genetically complex, and genetic re-assortment frequently converted [PSI(+)]-dependent phenotypes to stable traits that persisted in the absence of [PSI(+)]. Thus, [PSI(+)] provides a temporary survival advantage under diverse conditions, increasing the likelihood that new traits will become fixed by subsequent genetic change. As an epigenetic mechanism that globally affects the relationship between genotype and phenotype, [PSI(+)] expands the conceptual framework for phenotypic plasticity, provides a one-step mechanism for the acquisition of complex traits and affords a route to the genetic assimilation of initially transient epigenetic traits.

Biological Evolution↗

Mechanisms of facilitation in primed perceptual identification.

We investigated the mechanism that produces priming in perceptual identification. In Experiment 1, subjects studied a series of compound words (e.g., OUTDOOR, SIDELINE); in Experiment 2, subjects studied a series of pictures (photographs) of objects. All subjects later received perceptual identification tests in which old (primed) and new (unprimed) words (Experiment 1) or pictures (Experiment 2) were presented for varying durations and masked. In both experiments, performance for primed and unprimed stimuli was predicted essentially perfectly by a model that assumes that prior exposure to a stimulus results in increased visual information-acquisition rate when it is subsequently encountered. An ancillary purpose of Experiment 1 was to test whether or not priming occurs for recombined words (e.g., OUTLINE); there was no evidence for such priming at any exposure duration.

Adult↗

Characterization of the Vibrio cholerae El Tor lipase operon lipAB and a protease gene downstream of the hly region.

We have cloned and sequenced a region encoding a lipase operon and a putative, previously uncharacterized metalloprotease of Vibrio cholerae O1. These lie downstream of hlyA and hlyB, which encode the El Tor hemolysin and methyl-accepting chemotactic factor, respectively. Previous reports identified the hlyC gene downstream of hlyAB, encoding an 18.3-kDa protein. However, we now show that this open reading frame (ORF) encodes a 33-kDa protein, and since the amino acid sequence is highly homologous to the triacylglyceride-specific lipase of Pseudomonas spp., hlyC has been renamed lipA. LipA contains the highly conserved pentapeptide and catalytic triad amino acid regions of the catalytic sites of other lipases. The region downstream of lipA has been sequenced and has revealed ORFs lipB and prtV. The amino acid sequence of lipB is homologous to those of the accessory lipase proteins (lipase-specific foldase) required by Pseudomonas and various other bacterial species for the production of mature active lipase, and in agreement with this, we show that both lipA and lipB are required to restore a lipase-deficient lipA null mutant of V. cholerae. The intergenic stop codon for lipA overlaps the ribosome-binding site for lipB, and a stem-loop resembling a rho-independent terminator is present immediately downstream from lipB, suggesting that lipA and lipB form a lipase operon in V. cholerae. prtV lies downstream of lipAB but is transcribed in the opposite direction and is predicted to share the same putative transcriptional terminator with lipAB. The zinc-binding and catalytic domains conserved among many metalloproteases are present in PrtV, which is highly homologous to the immune inhibitor A (InA) metalloprotease of Bacillus thuringiensis. PrtV was visualized as approximately 102 kDa, which is consistent with the coding capacity of the gene. The genetic organization of this region suggests that it is possibly part of a pathogenicity island, encoding products capable of damaging host cells and/or involved in nutrient acquisition by V. cholerae. However, neither lipA nor prtV null mutants were attenuated in the infant mouse model, nor did they exhibit reduced colonization potential compared with wild type in competition experiments.

Amino Acid Sequence↗

Auditory word discrimination in male children diagnosed as having minimal brain dysfunction.

Investigated auditory word discrimination skill in children diagnosed as having minimal brain dysfunction (MBD), a heretofore unexplored topic in the study of MBD. A lack of auditory discrimination skill seems relevant to learning deficiency in language acquisition, a typical sign of MBD. The present study tested the hypothesis that phonemic aural discriminations would be difficult to make for boys diagnosed as having MBD. Errors and nonresponses made while Ss performed an extended auditory word discrimination task were compared between a group of 26 boys diagnosed as having MBD and a matched normal group. As expected, the MBD group had a significantly higher error rate than the normal group even when Ss were discriminating easy word pairs composed of dissimilar items (e.g., reading, math). These and other results suggested that the MBD group was very susceptible to fatigue even while motivated to perform well and while able to improve with practice. The relatively inferior performance of the MBD group was interpreted as resulting directly from a lack of auditory discrimination skill and indirectly from fatigue and its accompanying attention loss.

Attention↗

Discovery and distribution of super-integrons among pseudomonads.

Until recently, integrons (systems for acquisition and expression of new genetic materials) have been associated generally with antibiotic resistance gene cassettes. The discovery of 'super-integrons' in Vibrionaceae suggests a greater impact of this gene acquisition mechanism on bacterial genome evolution than initially believed. Super-integrons may contain more than 100 gene cassettes and may encode other determinants, including biochemical functions or virulence factors. Here, we report the genetic organization of a super-integron from Pseudomonas alcaligenes ATCC 55044. This is the first evidence of a super-integron in a non-pathogenic bacterium, one which is widely distributed in a great number of ecological niches such as soil and aquatic habitats. Here, the sequence composition, open reading frame (ORF) content and organization of In55044 are described and found to have features intermediate between the multidrug-resistant integrons and the Vibrio cholerae super-integron. Similar structures are inferred to be present in several Pseudomonas species, based on polymerase chain reaction (PCR) experiments.

Amino Acid Sequence↗

Knowledge of Chlamydia trachomatis genital infection and its consequences in people attending a genitourinary medicine clinic.

OBJECTIVES: To assess knowledge of Chlamydia trachomatis infections, with a comparison of knowledge of Neisseria gonorrhoeae infections. METHODS: A cross sectional survey, by self completed questionnaire, of 200 subjects attending a genitourinary outpatient clinic. RESULTS: The response rate was 82% (90 male and 73 female). 51% of men (60% of females) had heard of chlamydia. 65 (82%) were unaware of the potential consequences of infection. 66% were unaware that the disease could be asymptomatic. Significantly more men (77%) than women (60%) had heard of gonorrhoea. Most participants (68%-82%) knew little of the possible consequences of this infection, and only 26% were aware that it could be asymptomatic. Knowledge was higher regarding fertility topics. There was no correlation between knowledge and either age or socioeconomic status. However, greater knowledge was displayed by those who read health information leaflets always or often. For both men and women, the preferred source of health information was the doctor. Other popular sources were health information leaflets, women's magazines, and television. CONCLUSIONS: Barely half the participants had heard of chlamydia and gonorrhoea. Further knowledge of either infection was very poor. There are serious implications for public health. The reasons for this are unclear and require exploration before targeted health promotion. Doctors and the popular media are acceptable, and potentially effective, sources of information. Acquisition of knowledge is important, both to reduce sexual risk taking behaviour and its consequences, and to allow for informed consent for chlamydia screening programmes.

Adolescent↗

Skill acquisition in the implementation of functional analysis methodology.

Functional analysis methodology is a powerful assessment tool for identifying contingencies that maintain a wide range of behavior disorders and for developing effective treatment programs. Nevertheless, concerns have been raised about the feasibility of conducting functional analyses in typical service settings. In this study, we examined the issue of skill acquisition in implementing functional analyses by evaluating an instructional program designed to establish a basic set of competencies. Eleven undergraduate students enrolled in a laboratory course in applied behavior analysis served as participants. Their performance was assessed during scripted simulations in which they played the roles of "therapists" who conducted functional analyses and trained graduate students played the roles of "clients" who emitted self-injurious and destructive behaviors. To approximate conditions under which an individual might conduct an assessment with limited prior training, participants read a brief set of materials prior to conducting baseline sessions. A multiple baseline design was used to assess the effects of training, which consisted of reading additional materials, watching a videotaped simulation demonstrating correct procedural implementation, passing a written quiz, and receiving feedback on performance during sessions. Results showed that participants scored a relatively high percentage of correct therapist responses during baseline, and that all achieved an accuracy level of 95% or higher following training that lasted about 2 hr. These results suggest that basic skills for conducting functional analyses can be acquired quickly by individuals who have relatively little clinical experience.

Feasibility Studies↗

The time course for language acquisition in biologically distinct populations: evidence from deaf individuals.

The present study provides evidence that individuals who have different patterns of cerebral lateralization and who develop along different maturational time courses can attain comparable levels of language proficiency. Right-handed individuals with left-handed family members (left-handed familials, LHFs) showed a shorter sensitive period for language acquisition than did right-handed individuals with only right-handed family members (right-handed familials, RHFs). The shorter sensitive period for LHFs may be due to a focus on non-linguistic, word-based conceptual information during language acquisition. RHFs may focus on grammatical relations during language acquisition, which matures later than lexical knowledge. This suggests that there may be different patterns of cerebral lateralization for language in all normal populations as a function of familial handedness.

Adolescent↗

The minimum domain of Pho81 is not sufficient to control the Pho85-Rim15 effector branch involved in phosphate starvation-induced stress responses.

The phosphate regulatory mechanism in yeast, known as the PHO pathway, is regulated by inorganic phosphate to control the expression of genes involved in the acquisition of phosphate from the medium. This pathway is also reported to contribute to other nutritional responses and as such it affects several phenotypic characteristics known also to be regulated by protein kinase A, including the transcription of genes involved in the general stress response and trehalose metabolism. We now demonstrate that transcription of post-diauxic shift (PDS)-controlled stress-responsive genes is solely regulated by the Pho85-Pho80 complex, whereas regulation of trehalose metabolism apparently involves several Pho85 cyclins. Interestingly, both read-outs depend on Pho81 but, while the previously described minimum domain of Pho81 is sufficient to sustain phosphate-regulated transcription of PHO genes, full-length Pho81 is required to control trehalose metabolism and the PDS targets. Consistently, neither the expression control of stress-regulated genes nor the trehalose metabolism relies directly on Pho4. Finally, we present data supporting that the PHO pathway functions in parallel to the fermentable growth medium- or Sch9-controlled pathway and that both pathways may share the protein kinase Rim15, which was previously reported to play a central role in the integration of glucose, nitrogen and amino acid availability.

Blotting, Northern↗

Analysis of neuro-behavioral Discovery data on the Macintosh computer.

This paper describes a suite of routines using IgorPro, a powerful analysis and graphing software package for the Macintosh computer, to enhance the ability to analyze, manipulate, and display data recorded with the Discovery acquisition software marketed by DataWave Technologies. The routines are able to time-align fast and slow data channels, and are especially useful for analyses that involve both neural and behavioral data. The software was designed for eyeblink conditioning and vocalization experiments, but it can easily be used for analyzing other types of neurobehavioral data. The data are first prepared on the PC with routines that inspect the header of the data file and translate the data file into a compact binary format that can be read by IgorPro. An option is also available to splice out data from unnecessary portions of an intertrial interval. The new file is then put on the Macintosh computer for display and analysis by IgorPro. These routines enable both neural and behavioral data to be quickly and easily reduced, manipulated, and statistically and graphically summarized.

Analysis of Variance↗

Transposon-aided capture (TRACA) of plasmids resident in the human gut mobile metagenome.

The bacterial community of the human gut is a complex ecosystem composed of >1,000 species, the majority of which are Gram positive and uncultured. To study plasmids resident within this community, we developed a culture-independent transposon aided capture method (TRACA), which does not rely on any plasmid-encoded traits. TRACA facilitated acquisition of plasmids from metagenomic DNA extracts, and subsequent maintenance and selection in an Escherichia coli host. We confirmed the presence of the transposon in captured plasmids and demonstrate that these plasmids are mainly of a Gram-positive origin. Sequencing of plasmids designated pTRACA10 (7 kb) and pTRACA17 (2.7 kb) revealed genes involved in plasmid mobilization and replication. From the homologies of these genes we conclude that pTRACA17 originates from a Gram-positive host belonging to the Firmicute division. pTRACA10 had two additional open reading frames with similarity to a conserved hypothetical protein and phosphoesterase or phosphohydrolase enzymes (Clusters of Orthologous Groups number 4186). Both plasmids lacked any conventional selectable markers.

DNA Transposable Elements↗

A 37-channel DC SQUID magnetometer system.

A 37-channel DC SQUID magnetometer system has been built for biomagnetic studies. The SQUID loop of each magnetometer serves as the active sensing element, thereby eliminating the need for flux coupling circuits. The magnetometers are located approximately 3 cm above the outer dewar bottom. The SQUIDs are directly coupled to highly simplified read-out electronics using only five wires per channel; no helium temperature impedance matching circuits are required. Each channel can be independently inserted into or removed from the dewar. Using a novel electronic noise reduction technique the system white and 1 Hz flux density noise values are typically 5 and 10 fT Hz-1/2, respectively, including the noise contribution of the in-house fabricated dewar and the magnetically shielded room. The two parts of the data-processing system allow independent handling of the acquisition and analysing task. Two example measurements demonstrate the advantage of the electronic noise reduction method.

Equipment Design↗

The acquisition of arithmetic in normal children: assessment by a cognitive model of dyscalculia.

Developmental dyscalculia (DC) is a learning disability affecting the acquisition of arithmetic skills. The authors studied the normal developmental of arithmetic function of 200 normal children in grades 3 to 6. Number comprehension and production were well established by grade 3. With age and experience, children became more adept at addition, subtraction, multiplication and division for both number facts and complex exercises. The scores did not display a Gaussian distribution, but were negatively skewed. Low scores on the battery correlated well with the teacher's rating of the child's arithmetic knowledge, but not with reading ability. An estimate of the fifth centile was calculated. On the basis of these results, the authors conclude that this battery assesses number knowledge. Use of the fifth centile as a cut-off point will aid in identification of children with DC.

Child↗

Lateralization in normal 6-year-olds as related to later reading disability.

The differentiation of lateral preference in hand, eye, ear, and foot was measured in 110 normal Grade 1 children and reading achievement was tested 3 and 5 years later. No support was found for the widely held belief that delays in early lateralization underlie subsequent reading impairment. Paradoxically, boys with serious reading disability showed a strong degree of early lateral concordance. Male, impaired readers displayed a tendency when they were in Grade 1 to be either uniformly right- or left-preferring across all 4 sensory-motor modalities. The results contradict the popular notion that lateral usage emerges from an undifferentiated state in the direction of greater degrees of specialization. Lateralization may be an expression of reflex constraints bound initially to the infant's tonic-neck posture, with later development less reflex-patterned during the acquisition of more sophisticated information-processing strategies. Such an interpretation implies a progressive discontinuity between lateral usage and neuro-maturational processes. Alternatively, the results can be accounted for by (1) a new maturational delay model in which maturation-dependent lateralization proceeds in graduated stages from a highly specialized beginning toward greater levels of neuromotor integration; or (2) a model that maintains the notion of emergent lateralization but in which delays in the lateralization process are actually beneficial to later performance.

Achievement↗

Nervous system reorganization following injury.

Contrary to the classical view of a pre-determined wiring pattern, there is considerable evidence that cortical representation of body parts is continuously modulated in response to activity, behavior and skill acquisition. Both animal and human studies showed that following injury of the peripheral nervous system such as nerve injury or amputation, the somatosensory cortex that responded to the deafferented body parts become responsive to neighboring body parts. Similarly, there is expansion of the motor representation of the stump area following amputation. Reorganization of the sensory and motor systems following peripheral injury occurs in multiple levels including the spinal cord, brainstem, thalamus and cortex. In early-blind subjects, the occipital cortex plays an important role in Braille reading, suggesting that there is cross-modal plasticity. Functional recovery frequently occurs following a CNS injury such as stroke. Motor recovery from stroke may be associated with the adjacent cortical areas taking over the function of the damaged areas or utilization of alternative motor pathways. The ipsilateral motor pathway may mediate motor recovery in patients who undergo hemispherectomy early in life and in children with hemiplegic cerebral palsy, but it remains to be determined if it plays a significant role in the recovery of adult stroke. One of the challenges in stroke recovery is to identify which of the many neuroimaging and neurophysiological changes demonstrated are important in mediating recovery. The mechanism of plasticity probably differs depending on the time frame. Rapid changes in motor representations within minutes are likely due to unmasking of latent synapses involving modulation of GABAergic inhibition. Changes over a longer time likely involve other additional mechanisms such as long-term potentiation, axonal regeneration and sprouting. While cross-modal plasticity appears to be useful in enhancing the perceptions of compensatory sensory modalities, the functional significance of motor reorganization following peripheral injury remains unclear and some forms of sensory reorganization may even be associated with deleterious consequences like phantom pain. An understanding of the mechanism of plasticity will help to develop treatment programs to improve functional outcome.

Aging↗