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M J Ward

Publications and source records attributed to M J Ward.

At least 37 records · Page 2Linked to original sources
PubMed1998

Regulation of motility behavior in Myxococcus xanthus may require an extracytoplasmic-function sigma factor.

Using interaction trap technology, we identified a putative extracytoplasmic-function (ECF) sigma factor (RpoE1) in Myxococcus xanthus, a bacterium which has a complex life cycle that includes fruiting body formation. The first domain of the response regulator protein FrzZ, a component of the Frz signal transduction system, was used as bait. Although the RpoE1 protein displayed no interactions with control proteins presented as bait, a weak interaction with a second M. xanthus response regulator (AsgA) was observed. While the specificity of the FrzZ-RpoE1 interaction therefore remains speculative, cloning and sequencing of the region surrounding rpoE1 localized it to a position downstream of the frzZ gene. A potential promoter site for binding of an ECF sigma factor was identified upstream of rpoE1, suggesting the gene may be autoregulated. However, primer extension studies suggested that transcription of rpoE1 occurs under both vegetative and developmental conditions from a sigma70-like promoter. Dot blot analysis of RNA preparations confirmed the low-level, constitutive expression of rpoE1 during both stages of the life cycle. Analysis of an insertion mutant also indicated a role for RpoE1 under both vegetative and developmental conditions, since swarming was reduced on nutrient-rich agar and developmental aggregation was effected under starvation conditions, especially at high cell densities. An insertion mutation introduced into the gene directly downstream of rpoE1 (orf5) did not result in either swarming or developmental aggregation defects, even though the gene is transcribed as part of the same operon. Therefore, we propose that this new ECF sigma factor could play a role in the transcriptional regulation of genes involved in motility behavior during both stages of the complex M. xanthus life cycle.

Amino Acid Sequence↗
PubMed1998

An ABC transporter plays a developmental aggregation role in Myxococcus xanthus.

Myxococcus xanthus is a gram-negative bacterium which has a complex life cycle. Autochemotaxis, a process whereby cells release a self-generated signaling molecule, may be the principal mechanism facilitating directed motility in both the vegetative swarming and developmental aggregation stages of this life cycle. The process requires the Frz signal transduction system, including FrzZ, a protein which is composed of two domains, both showing homology to the enteric chemotaxis response regulator CheY. The first domain of FrzZ (FrzZ1), when expressed as bait in the yeast two-hybrid system and screened against a library, was shown to potentially interact with the C-terminal portion of a protein encoding an ATP-binding cassette (AbcA). The activation domain-AbcA fusion protein did not interact with the second domain of FrzZ (FrzZ2) or with two other M. xanthus response regulator-containing proteins presented as bait, suggesting that the FrzZ1-AbcA interaction may be specific. Cloning and sequencing of the upstream region of the abcA gene showed the ATP-binding cassette to be linked to a large hydrophobic, potentially membrane-spanning domain. This domain organization is characteristic of a subgroup of ABC transporters which perform export functions. Cloning and sequencing downstream of abcA indicated that the ABC transporter is at the start of an operon containing three open reading frames. An insertion mutation in the abcA gene resulted in cells displaying the frizzy aggregation phenotype, providing additional evidence that FrzZ and AbcA may be part of the same signal transduction pathway. Cells with mutations in genes downstream of abcA showed no developmental defects. Analysis of the proposed exporter role of AbcA in cell mixing experiments showed that the ABC transporter mutant could be rescued by extracellular complementation. We speculate that the AbcA protein may be involved in the export of a molecule required for the autochemotactic process.

ATP-Binding Cassette Transporters↗
PubMed1998

Systemic activity of inhaled and swallowed beclomethasone dipropionate and the effect of different inhaler devices.

Inhaled glucocorticoids such as beclomethasone dipropionate, which are used in the treatment of asthma, may be associated with systemic adverse effects. To determine whether any systemic absorption following the inhalation of beclomethasone was a result of drug being absorbed from the lung (inhaled fraction) or the gastrointestinal tract (swallowed fraction), we studied normal subjects after the inhalation or swallowing of 2 mg beclomethasone dipropionate. Systemic activity was assessed using early morning cortisol suppression. Both inhaled and swallowed fractions produced significant systemic activity, the degree of which depended on the inhaler device used. Systemic activity was greater using a dry powder inhaler (52%) than using a metered dose inhaler with a large volume spacer (28%). These findings suggest that to limit potential adverse effects from high-dose beclomethasone dipropionate it is better to use a metered dose aerosol with large volume spacer than a dry powder.

Administration, Inhalation↗
PubMed1997

Regulation of directed motility in Myxococcus xanthus.

Myxococcus xanthus is a Gram-negative bacterium that exhibits a complex life cycle. During vegetative growth, cells move as large swarms. However, when starved, cells aggregate into fruiting bodies and sporulate. Both vegetative swarming and developmental aggregation require gliding motility, which involves the slow movement of cells on a solid surface in the absence of flagella. The frequency of cell reversals controls the direction of movement and is regulated by the frz genes, which encode the 'frizzy' signal-transduction proteins. These proteins contain domains which bear striking similarities to the major chemotaxis proteins of the enteric bacteria: CheA, CheY, CheW, CheR, CheB and Tar. However, significant differences exist between the Myxococcus Frz proteins and the enteric Che/MCP proteins. For example, the Frz system contains three CheY-like response-regulator domains: one is present on FrzE, which also contains a CheA-like domain, and two are present on FrzZ, which is a novel protein required for attractant, but not for repellent, responses. The identification of multiple CheY homologues in this system indicates a more complex regulatory pathway than that found in the enteric bacteria. While responses to repellent stimuli appear to follow the enteric paradigm, responses to attractants during vegetative swarming and development are more complex and may involve self-generated autoattractants. The Frz signal-transduction system regulates directed motility in M. xanthus and is essential for controlling both fruiting-body development and vegetative swarming.

Bacterial Proteins↗
PubMed1997

Bladder management in patients with pediatric onset neurogenic bladders.

Our objective was to determine which clean intermittent catheterization (CIC) methods and supplies were used by patients with pediatric onset neurogenic bladders and to relate methodology and materials to reported urinary tract infections. Data were collected via questionnaires distributed by mail and at clinic visits at our university tertiary care outpatient pediatric rehabilitation clinic. Questionnaires were given to 165 patients. Fifty-nine percent were returned (68 patients with myelomeningocele, 27 with pediatric onset spinal cord injury (SCI) and two with other diagnoses). Mean age was 12 years (range 1-27). Fifty-four percent of patients participated in their own CIC. Only two percent used sterile catheterization technique, whereas 98 percent used CIC. A sterile catheter was employed with clean technique by 22 percent. Catheters were reused by 76 percent. Subjects used a wide ranging number of catheters per month, with a median of 5.3. There was no correlation between the number of urinary tract infections (UTIs) per year and the type of catheter used or the use of prophylactic antibiotics. Compared with patients with myelomeningocele, subjects with SCI were significantly more likely to use sterile catheters (p = 0.04), > 10 catheters per month (p = 0.01) and gloves (p < 0.001). Subjects who used gloves or more catheters were more likely to experience UTI. These data suggest that clean reused supplies are not related to an increased likelihood of UTI and should be considered a way to lower costs in these populations.

Adolescent↗
PubMed1997

Localization of upstream silencer elements involved in the expression of cone transducin alpha-subunit (GNAT2).

PURPOSE: To localize cis-acting elements involved in the expression of the cone-specific G-protein, cone transducin alpha-subunit (GNAT2). METHODS: In this study, the authors used a genomic clone, HGLG3, to sequence 3139 base pairs of the upstream region of the GNAT2 gene and to localize cis-acting elements involved in the expression of GNAT2. Upstream elements were localized functionally by transfection of chloramphenicol acetyltransferase gene constructs containing nested deletions of this upstream region into WERI-Rb1 cells. Cell specificity of the localized elements was determined by transfection of the HeLa cells. Trans-acting factor-binding sites to functional cis-acting elements were determined by DNasel footprinting. Cell specificity of protein interaction with footprinted regions was tested by electrophoretic mobility shifts with nuclear extracts from WERI-Rb1 and HeLa cells. RESULTS: Transfection of WERI-Rb1 and HeLa cells revealed the presence of a strong, noncell-specific silencer region between -1130 and -23, a weak, cell-specific promoter between -151 and -10, and a stronger, noncell-specific element between +143 and +167. DNaseI footprinting showed three major footprints (S1, S2, and S3) between -807 and -176, indicating the binding sites for putative negative trans-acting factors. Individual footprinted sequences had similar electrophoretic mobility shifts when they were incubated with nuclear extracts from either WERI-Rb1 or HeLa cells, suggesting that these cells express the same negative factors. CONCLUSIONS: The expression of the GNAT2 gene is controlled by a strong silencer region, a weak upstream cell-specific promoter, and a strong downstream element. The silencer region interacts with similar proteins from retina- and nonretina-derived cell lines.

Base Sequence↗
PubMed1996

Spondyloepiphyseal dysplasia congenita with ventilator dependence: two case reports.

Home mechanical ventilation has recently become feasible in a number of conditions. Several small series and case reports mention respiratory insufficiency or failure in a subset of patients with spondyloepiphyseal dysplasia congenita. Two cases of successful home mechanical ventilation in children, now 7 years of age, who have spondyloepiphyseal dysplasia congenita and respiratory insufficiency, are reported. Novel rehabilitation planning has facilitated the growth and development of these children.

Child↗
PubMed1996

Identification and characterization of FrzZ, a novel response regulator necessary for swarming and fruiting-body formation in Myxococcus xanthus.

The frz genes of Myxococcus xanthus constitute a signal-transduction pathway that processes chemotactic information in a manner analogous to that found in enteric bacteria. Ultimately, these genes regulate the frequency of individual cell reversal. We report here the identification of a novel component of this signal-transduction pathway, designated frzZ, which was discovered as an open reading frame located 5' to the frz operon but transcribed in the opposite orientation. The translational start site of frzZ is 170 base pairs from that of frzA.frzZ utilizes a promoter similar to the sigma 70 promoters of Escherichia coli, and encodes a 290-amino-acid soluble protein, FrzZ (M(r) 30,500). FrzZ contains two domains, both of which show strong homology to CheY and other members of the response-regulator family. Linking these domains is a 39-amino-acid region that is very rich in alanine and proline (38% Ala and 33% Pro). A frzZ null mutant showed abnormally low reversal rates when compared to the wild-type control and was unable to form fruiting bodies on starvation medium, but it did form 'frizzy' aggregates. In addition, the frzZ mutant was defective in swarming, particularly on soft agar (0.3% w/v). However, unlike most frz mutants, the frzZ mutant was able to respond to attractants and repellents in the spatial chemotaxis assay. The discovery of FrzZ demonstrates that the M. xanthus frz signal-transduction pathway utilizes multiple response-regulator (CheY-like) proteins.

Amino Acid Sequence↗
PubMed1995

Tissue-engineered cartilage for implantation and grafting.

Facial plastic and reconstructive surgeons are continually faced with the dilemma of what material to use for implantation and grafting. Tissue-engineered cartilage is a relatively new and exciting concept which utilizes chondrocytes and cultures them on a three-dimensional biodegradable template. This template/cell complex is first briefly incubated in vitro, then implanted into a recipient host. In situ the template resorbs and is replaced with new cartilage that is viable, compatible, and mature. This paper discusses the biochemical composition of cartilage, the concept of tissue engineering, advances in template quality, and cartilage immunogenicity. Future clinical applications of this type of graft research include microtia repair, facial reconstruction, rhinoplasty, and other facial cosmetic procedures.

Animals↗
PubMed1995

Identification of a chemotaxis operon with two cheY genes in Rhodobacter sphaeroides.

A large chemotaxis operon was identified in Rhodobacter sphaeroides WS8-N using a probe based on the 3' terminal portion of the Rhizobium meliloti cheA gene. Two genes homologous to the enteric cheY were identified in an operon also containing cheA, cheW, and cheR homologues. The deduced protein sequences of che gene products were aligned with those from Escherichia coli and shown to be highly conserved. A mutant with an interrupted copy of cheA showed normal patterns of swimming, unlike the equivalent mutants in E. coli which are smooth swimming. Tethered cheA mutant cells showed normal responses to changes in organic acids, but increased, inverted responses to sugars. The unusual behaviour of the cheA mutant and the identification of two homologues of cheY suggests that R. sphaeroides has at least two pathways controlling motor activity. To identify functional similarity between the newly identified R. sphaeroides Che pathway and the methyl-accepting chemotaxis protein (MCP)-dependent pathway in enteric bacteria, the R. sphaeroides cheW gene was expressed in a cheW mutant strain of E. coli and found to complement, causing a partial return to a swarming phenotype. In addition, expression of the R. sphaeroides gene in wild-type E. coli resulted in the same increased tumbling and reduced swarming as seen when the native gene is overexpressed in E. coli. The identification of che homologues in R. sphaeroides and complementation by cheW suggests the presence of MCPs in an organism previously considered to use only MCP-independent sensing. The MCP-dependent pathway, appears conserved.(ABSTRACT TRUNCATED AT 250 WORDS)

Acetates↗
PubMed1995

Identification of a methyl-accepting chemotaxis protein in Rhodobacter sphaeroides.

Analysis of the DNA sequence directly upstream of the chemotaxis operon of Rhodobacter sphaeroides identified a single gene whose product has strong similarity to the methyl-accepting chemotaxis proteins (MCPs) found in enteric bacteria. The deduced protein had a highly conserved signalling sequence and only one very hydrophobic region at the N-terminus, in contrast to enteric MCPs. A possible cytoplasmic location of the majority of the protein was supported by Western blotting. The mcpA gene was insertionally inactivated and the resulting phenotype examined using swarm plate assays. The mutant lacking McpA lost chemotaxis to a wide range of attractant stimuli but only under aerobic conditions; it retained almost normal chemotaxis under anaerobic/photosynthetic conditions. The identification of a sensory protein which is active only under one set of growth conditions suggests that R. sphaeroides probably has several MCPs, which co-ordinately respond to changes in environmental conditions. Southern hybridization at relaxed stringency to the conserved sequence of the R. sphaeroides and Caulobacter crescentus mcp genes identified three possible additional mcp genes.

Amino Acid Sequence↗
PubMed1995

Associations among adult attachment representations, maternal sensitivity, and infant-mother attachment in a sample of adolescent mothers.

Associations among adolescent attachment organization, maternal sensitivity, and infant attachment organization were examined prospectively in 74 teenaged mother-infant dyads. Pregnant teenagers' attachment organizations predicted both sensitivity and infant-mother attachments. Mothers classified autonomous (F) in the prenatal period showed higher levels of sensitivity at both 3 and 9 months than mothers classified dismissing (Ds), preoccupied (E), or unresolved (U). Correspondence between maternal attachment (F vs. Ds/E/U) and infant attachment (secure [B] vs. avoidant [A]/resistant [C]/disorganized [D]) was observed in 58 of 74 (78%) dyads. Exact 4-group (Ds/E/F/U and A/B/C/D) agreement was observed in 50 of 74 (68%) families. In contrast, associations between maternal sensitivity and infant attachment were not significant, leading to questions about the processes that link attachment representations, maternal behavior, and infant attachment in adolescent mothers.

Adolescent↗
PubMed1994

Role of the mother's touch in failure to thrive: a preliminary investigation.

OBJECTIVE: This study investigated the hypothesis that specific types of maternal touch that might promote infant growth are reduced in cases of failure to thrive (FTT) and tested reports that maternal physical interaction and physical affection are diminished in FTT. METHODS: Frequencies of operationally defined maternal touch categories were scored by blinded raters from videotaped mother-child feeding and play interactions of 21 children, aged 9 to 19 months, with FTT and 18 normally growing comparison children. After scoring and statistical analyses were completed, investigators unblinded to group status and clinical data reviewed the videotapes of the dyads with the lowest touch scores. RESULTS: Mothers of children with FTT provided less matter-of-fact touch in feeding (p = .017) and unintentional touch in play (p = .048) than the comparison group, and there was a trend (p = .082) for them to provide less proprioceptive stimulation in play. Unblinded case reviews indicate that, among children with FTT, extremely infrequent touch signals a marked touch aversion by either the mother or child. CONCLUSIONS: Types of maternal touch that may promote growth or facilitate feeding are reduced in FTT, due, in extreme cases, to maternal or child touch aversion. Clinicians evaluating FTT should be alert to very infrequent touch in the mother-child interaction and consider whether it may represent a maternal intolerance of physical contact with her infant or a problem with the infant's feeding competence.

Child, Preschool↗