Biomedical subjects

J C Walker

Publications and source records attributed to J C Walker.

At least 19 recordsLinked to original sources

PubMed2001

BRS1, a serine carboxypeptidase, regulates BRI1 signaling in Arabidopsis thaliana.

Brassinosteroid-insensitive 1 (BRI1) of Arabidopsis thaliana encodes a cell surface receptor for brassinosteroids. Mutations in BRI1 severely affect plant growth and development. Activation tagging of a weak bri1 allele (bri1-5) resulted in the identification of a new locus, brs1-1D. BRS1 is predicted to encode a secreted carboxypeptidase. Whereas a brs1 loss-of-function allele has no obvious mutant phenotype, overexpression of BRS1 can suppress bri1 extracellular domain mutants. Genetic analyses showed that brassinosteroids and a functional BRI1 protein kinase domain are required for suppression. In addition, overexpressed BRS1 missense mutants, predicted to abolish BRS1 protease activity, failed to suppress bri1-5. Finally, the effects of BRS1 are selective: overexpression in either wild-type or two other receptor kinase mutants resulted in no phenotypic alterations. These results strongly suggest that BRS1 processes a protein involved in an early event in the BRI1 signaling.

Amino Acid Sequence↗

PubMed2001

Investigating sources of response variability and neural mediation in human nasal irritation.

A major component of indoor air complaints is nasal irritation (NI), yet there is an extreme paucity of quantitative concentration-response data from normosmics (individuals who report normal odor sensation). Due to an assumption that NI is mediated solely by the activation of the trigeminal (fifth cranial) nerve, much of the small amount of available information has been obtained from anosmic individuals, who lack olfactory (first cranial) nerve input to the brain and, thus, only have nasal trigeminal input remaining. In a repeated measurements design, the NI responses of 31 normosmic and four anosmic individuals were quantified in response to a range of concentrations of propionic acid generated by an automated air-dilution olfactometer. A variance analysis approach was used to apportion different nested sources of variation (within-session, within-individual, inter-individual) in NI responses. In contrast to anosmic NI and normosmic odor performance, NI response by normosmics exhibited considerable variation at all three levels. However, this variation did not obscure the observation that, in agreement with electrocortical measurements by Hummel et al. (1996), NI sensitivity in normosmics clearly exceeded that of anosmics. These observations provide support for enhanced research efforts to better understand the neural basis of NI so that its occurrence in actual environments may be effectively minimized.

Adult↗

PubMed2001

Human responses to propionic acid. II. Quantification of breathing responses and their relationship to perception.

In 20 normal and four anosmic participants, instantaneous inhalation and exhalation flow rates were recorded in response to 15 s stimulations with clean air or propionic acid concentrations (0.16, 1.14, 8.22 and 59.15 p.p.m., v/v) that ranged from peri-threshold for normals to clearly supra-threshold for anosmics. Each odorant/irritant delivery to the face-mask began with an exhalation. This allowed concentration to reach full value before stimulus onset, defined as the point where the participant began to bring the stimulus into the nose by inhalation. Two seconds after this stimulus onset, normals exhibited cumulative inhaled volume (CIV) declines of 39 and 14%, and latencies of 500 and 710 ms, with presentations of 59.15 and 8.22 p.p.m., respectively. With anosmics, 59.15 p.p.m. caused a 19% decline in CIV that began at 730 ms. Examination of the first inhalation after stimulus onset shows that the CIV declines in normals were achieved by a progressive decline in volume (InVol), beginning with a slight drop at 1.14 p.p.m., and a marked decline in duration (InDur) with only the highest concentration. Anosmics exhibited declines in InDur and InVol with only the 59.15 p.p.m. stimulus, and these declines were much more modest than the changes seen in normals. Comparison of these breathing results with perceptual responses from this same experiment demonstrates that: (i) in normals, odor perception rises slightly, but breathing does not change, with the lowest concentration; (ii) the higher breathing sensitivity (declines in InVol) of normals is paralleled by both the higher nasal irritation of these individuals and the presence of odor sensation; (iii) InDur declines in normals only with a stimulus concentration sufficient to cause marked nasal irritation in anosmics; and iv) in anosmics, modest but reliable declines in both InDur and InVol mirror the marked elevation in nasal irritation magnitude seen with only the highest concentration. In view of the failure of prior work to provide evidence that olfactory activation alone can cause any of the breathing changes we observed, we conclude that some breathing parameters are quite useful as rapid and sensitive measures of nasal irritation that arises from activation of nasal trigeminal afferents alone or in combination with the olfactory nerve.

Adult↗

PubMed2001

A mutant Arabidopsis heterotrimeric G-protein beta subunit affects leaf, flower, and fruit development.

A genetic screen was performed to find new mutants with an erecta (er) phenotype and to identify genes that may function with ER, a receptor-like kinase. These mutants were named elk (for erecta-like) and were placed into five complementation groups. We positionally cloned ELK4 and determined that it encodes AGB1, a putative heterotrimeric G-protein beta subunit. Therefore, elk4 was renamed agb1. agb1-1 plants express similar fruit phenotypes, as seen in er plants, but differ from er in that the stem is only slightly shorter than that in the wild type, the pedicel is slightly longer than that in the wild type, and the leaves are rounder than those in er mutants. Molecular analysis of agb1-1 indicates that it is likely a null allele. AGB1 mRNA is expressed in all tissues tested but is highest in the silique. Analysis of agb1-1 er double mutants suggests that AGB1 may function in an ER developmental pathway regulating silique width but that it functions in parallel pathways affecting silique length as well as leaf and stem development. The finding that AGB1 is involved in the control of organ shape suggests that heterotrimeric G-protein signaling is a developmental regulator in Arabidopsis.

Arabidopsis↗

PubMed2001

Chlamydia trachomatis infection does not enhance local cellular immunity against concurrent Candida vaginal infection.

Although Th1-type cell-mediated immunity (CMI) is the predominant host defense mechanism against mucosal Candida albicans infection, CMI against a vaginal C. albicans infection in mice is limited at the vaginal mucosa despite a strong Candida-specific Th1-type response in the draining lymph nodes. In contrast, Th1-type CMI is highly effective against an experimental Chlamydia trachomatis genital tract infection. This study demonstrated through two independent designs that a concurrent Candida and Chlamydia infection could not accelerate or modulate the anti-Candida CMI response. Together, these results suggest that host responses to these genital tract infections are independent and not influenced by the presence of the other.

Animals↗

PubMed2001

Human breathing and eye blink rate responses to airborne chemicals.

Increased levels of air pollution have been linked with morbidity and mortality, but mechanisms linking physiologic responses to quality of life and productivity issues remain largely unknown. Individuals often report irritation of the nose and/or eyes upon exposures to environmental contaminants. Evaluation of these self-reports would be greatly aided by the development of valid physiological markers. Chamber studies (unencumbered exposures) of nonsmoker responses to environmental tobacco smoke offer two candidate end points: (a) Tidal volume increases and breathing frequency declines with stimuli that elicit only moderate irritation. (b) Eye blink rate increases only with a concentration sufficiently high to cause progressive worsening of eye irritation with prolonged exposure. Experiments with very brief nasal-only presentations also suggest the value of breathing changes as sensitive markers of irritation: (a) Tidal volume is inversely related to perceived nasal irritation (NI) intensity in both normal and anosmic (lacking olfactory input) individuals, although normals exhibit greater NI sensitivity. (b) Inhalation duration, in both groups, declines only with trigeminal activation sufficient to cause readily perceptible NI in anosmics. Changes in eye blink rate and breathing may be useful in the investigation of irritation and other effects of air pollution, and could be quite useful in investigations of mixtures of volatile organic compounds.

Air Pollutants↗

PubMed2001

The performance of the human nose in odour measurement.

Over the last 20 years or so, there has been steadily increasing activity in the area of applied human odour measurement. This has been especially true outside of the United States. Yet, for about 40 years, there has also been decreasing interest and activity, on the part of academic smell researchers, in rigorous quantitative measurement of the functional properties of the human olfactory system. There are some optimistic signs, however, that this situation may be improving. Applied meetings such as this one are reaching out to learn more about basic research in human olfaction and some research groups are venturing out to indoor air quality, environmental health, water quality and other applied areas. In this paper I hope to support and accelerate the increasingly fruitful interactions that are beginning. The paper aims to make four main points. First, some of the most important ways in which the laboratory differs from everyday life will be noted. Keeping these differences in mind lessens the risk that laboratory data will be used uncritically to make predictions of real-world responses to chemical stimuli. Next, the specific benefits that would accrue from more fruitful interactions between basic and applied researchers will be highlighted; this is perhaps best seen by noting problem areas resulting from too little cross-fertilisation. Third, the CEN standard for the measurement of odour thresholds will be discussed in light of what is known concerning both the functional aspects of the human olfactory system and the current state of knowledge concerning best methods for investigating this system. Finally, some recent work we have done that was designed to help characterise human odour responses and demonstrate improved methodology, will be briefly mentioned. The paper concludes with suggestions as to how the scientific basis of applied odour measurement may best be enhanced.

Air Pollutants↗

PubMed2000

Individual variation in the expression profiles of nicotinic receptors in the olfactory bulb and trigeminal ganglion and identification of alpha2, alpha6, alpha9, and beta3 transcripts.

Nicotine evokes dose-dependent and often variable chemosensory responses in animals and humans. Earlier observations that nicotine binds to some nicotinic acetylcholine receptor (nAChR) subtypes in the olfactory bulb (OB) and trigeminal ganglion (TG) led us to investigate the complete nAChR expression profile in each tissue and to determine whether inter-individual differences exist in male and female rats. Total RNA was extracted from individual samples of dissected OB and TG and analyzed by a sensitive reverse transcription-polymerase chain reaction (RT-PCR) assay to determine the messenger RNA profiles of ten transcripts encoded by the alpha2, alpha3, alpha4, alpha5, alpha6, alpha7, alpha9, beta2, beta3, and beta4 nAChR genes. We found that (a) in the OB, all animals expressed alpha2, alpha3, alpha4, alpha5, alpha7, beta2, and beta4 subunit mRNAs, whereas alpha6, beta3, and alpha9 transcripts were expressed in only 17, 28, and 33% of the animals, respectively, and (b) in the TG, all animals expressed alpha2, alpha3, alpha6, alpha7, beta2, and beta4 subunit mRNAs, whereas alpha9, beta3, alpha4, and alpha5 transcripts were expressed in 4, 38, 88, and 92% of the animals, respectively. These results also identified new subunits that are expressed in each tissue (alpha2, alpha6, alpha9, and beta3) and demonstrated that individual rats may have different tissue-specific expression profiles for alpha4, alpha5, alpha6, alpha9, and beta3 transcripts. Such variations are likely to be reflected in the composition of functional receptor subtypes in the rat OB and TG that have different activation and desensitization characteristics to acetylcholine and nicotine.

Acetylcholine↗

PubMed2000

HAESA, an Arabidopsis leucine-rich repeat receptor kinase, controls floral organ abscission.

Abcission, the natural shedding of leaves, flowers and fruits, is a fundamental component of plant development. Abscission is a highly regulated process that occurs at distinct zones of cells that undergo enlargement and subsequent separation. Although some components of abscission, including accumulation of the hormone ethylene and cell wall-degrading enzymes, have been described, the regulatory pathways remain largely unknown. In this paper we describe a critical component required for floral organ abscission in Arabidopsis thaliana, the receptor-like protein kinase HAESA. Histochemical analysis of transgenic plants harboring a HAESA promoter:: beta-glucuronidase reporter gene and in situ RNA hybridization experiments show HAESA expression in the abscission zones where the sepals, petals, and stamens attach to the receptacle, at the base of pedicels, and at the base of petioles where leaves attach to the stem. Immunodetection, immunoprecipitation, and protein kinase activity assays reveal HAESA is a plasma membrane serine/threonine protein kinase. The reduction of function of HAESA in transgenic plants harboring an antisense construct results in delayed abscission of floral organs, and the severity of the phenotype is directly correlated with the level of HAESA protein. These results demonstrate that HAESA functions in developmentally regulated floral organ abscission.

Arabidopsis↗

PubMed2000

Method for calibrating olfactometer output. Part 2. Amyl acetate.

A method for measuring amyl acetate in air was developed and validated. Known volumes of air samples from the output of an olfactometer, a device used to generate odor stimuli, were passed through charcoal sorbent tubes. Following extraction of the sorbent with carbon disulfide, the amount of amyl acetate collected on each tube was determined by gas chromatography with flame ionization detection. The method was used to determine the actual concentrations of amyl acetate presented to experimental participants in odor sensitivity testing.

Air Pollutants↗

PubMed2000

Differential regulation of CD4 lymphocyte recruitment between the upper and lower regions of the genital tract during Chlamydia trachomatis infection.

Genital infection with Chlamydia trachomatis results in both the local recruitment of protective immune responses and an inflammatory infiltrate that may also participate in tubal pathology. As a beginning to understanding the etiology of immune system-mediated tubal pathology, we evaluated the regional recruitment of lymphocyte subsets to different areas of the female genital tract (GT) over the course of a murine infection with the mouse pneumonitis agent of Chlamydia trachomatis (MoPn). Using flow cytometric techniques we found that the CD4 lymphocyte subset was preferentially recruited to the upper GT (oviduct and uterine horn) over the lower GT (cervical-vaginal region) throughout the course of MoPn infection. The influx of CD4 cells also correlated with the expression of endothelial cell adhesion molecules (ECAMs) and in vitro lymphocyte adherence in the upper GT. Interestingly, the expression of ECAMs in the lower GT was not maintained longer than 7 days after infection, even in the presence of viable chlamydiae. Taken together, these data suggest that regulatory mechanisms of lymphocyte recruitment differ between the upper and lower regions of the GT and may influence the clearance of chlamydiae and the development of tubal pathology.

Animals↗

PubMed2000

Sedation for vitreoretinal surgery: a comparison of anaesthetist-administered midazolam and patient-controlled sedation with propofol.

Local anaesthesia is increasingly being used for vitreoretinal surgery, but the optimal technique for sedation remains unclear. Anaesthetist-administered midazolam, which is often used, was compared in this study to patient-controlled sedation with propofol in 43 patients undergoing 50 vitreoretinal procedures. A variety of patient, anaesthetist and surgical endpoints were measured. There were no significant outcome differences between the two agents except that midazolam produced more amnesia for the local anaesthetic eye block. However, several outcomes and the observations in patients who experienced both agents showed a trend in favour of propofol for intraoperative sedation. We conclude that both approaches are safe and that patient-controlled sedation with propofol is at least as satisfactory as anaesthetist-administered midazolam.

Procedural Sedation↗

PubMed2000

The FHA domain mediates phosphoprotein interactions.

The forkhead-associated (FHA) domain is a phosphopeptide-binding domain first identified in a group of forkhead transcription factors but is present in a wide variety of proteins from both prokaryotes and eukaryotes. In yeast and human, many proteins containing an FHA domain are found in the nucleus and involved in DNA repair, cell cycle arrest, or pre-mRNA processing. In plants, the FHA domain is part of a protein that is localized to the plasma membrane and participates in the regulation of receptor-like protein kinase signaling pathways. Recent studies show that a functional FHA domain consists of 120-140 amino acid residues, which is significantly larger than the sequence motif first described. Although FHA domains do not exhibit extensive sequence similarity, they share similar secondary and tertiary structures, featuring a sandwich of two anti-parallel (beta)-sheets. One intriguing finding is that FHA domains may bind phosphothreonine, phosphoserine and sometimes phosphotyrosine, distinguishing them from other well-studied phosphoprotein-binding domains. The diversity of proteins containing FHA domains and potential differences in binding specificities suggest the FHA domain is involved in coordinating diverse cellular processes.

Arabidopsis↗

PubMed1999

Kinase interaction domain of kinase-associated protein phosphatase, a phosphoprotein-binding domain.

Kinase-associated protein phosphatase interacts specifically with plant receptor-like protein kinases. This interaction is thought to be a key step in signal perception and transduction. The minimal kinase interaction (KI) domain of kinase-associated protein phosphatase was mapped to a 119-aa segment spanning residues 180 to 298. A forkhead-associated (FHA) homology region resides in this minimal KI domain. Site-directed mutagenesis of four highly conserved sites in this FHA homology region abolishes the KI domain's interaction with receptor-like protein kinases, indicating that the FHA region is essential for binding. Serial deletion analysis indicates that 30 aa on each side of the FHA region are also needed for binding; this minimal functional unit is designated as the KI domain. Kinetic studies using surface plasmon resonance indicate that the binding between the KI domain and receptor-like protein kinases has a dissociation constant (KD) of about 25-100 nM, which is similar to the binding affinity of two other well characterized phosphorylation-dependent protein-binding domains (14-3-3 and Src homology 2) and their high-affinity phosphopeptide ligands.

Amino Acid Sequence↗

PubMed1999

Molecular evolution of type 1 serine/threonine protein phosphatases.

Type 1 serine/threonine protein phosphatases (PP1s) play key roles in many cellular processes. To understand the evolutionary relationships among PP1s from various kingdoms and to provide a valid basis to evaluate the structure-function relationships of these phosphatases, 44 PP1 sequences were aligned, revealing a high sequence similarity among PP1 homologs. About one-third of the total amino acids are conserved in all the sequences studied. Most of these conserved amino acids are located within a 270-amino-acid core region. They include most sites critical to the activity and regulation of PP1s based on three-dimensional structural studies of mammalian PP1s. Positional variation analysis using a sliding window approach revealed two variable blocks in the 270-amino-acid core region. The major variable block corresponds to a subdomain composed of three alpha-helices (alphaG, alphaH, and alphaI) and three beta-sheets (beta7, beta8, and beta9). Phylogenetic analyses suggested that plant and animal PP1s form distinct monophyletic groups. The plant PP1 family contains several subgroups that may be older than the monocot-dicot divergence. In the animal PP1 family, different vertebrate isoforms appear to form distinct subgroups. Relative substitution rate studies indicated that plant PP1s are more diverse than animal PP1s, with an average substitution rate 1.5 times as large as that of animal PP1s. The possible involvement of PP1s in the establishment of multicellularity is discussed.

Amino Acid Sequence↗

PubMed1998

Challenges in understanding RLK function.

Plants use receptor-like kinases (RLKs) to transduce extracellular signals into the cell. Recent advancements in RLK research include the cloning of the BRASSINOSTEROID INSENSITIVE 1 and CLAVATA 1 genes, revealing RLK roles in development. Our understanding of RLK function has also been broadened by transgenic approaches in the study of the RLKs pollen receptor kinase 1, and wall associated kinase 1. These results extend the observations that RLKs function in developmental processes and plant defense responses. Additionally, expression based studies suggest roles for other newly reported RLKs in development and light responses. Taken together, the studies confirm the importance of RLKs in diverse plant processes, yet major challenges remain. These include identifying ligands that activate RLKs and characterizing downstream pathways. These challenges can be conquered by coordinated efforts from investigators using molecular, genetic, and biochemical approaches.

Amino Acid Sequence↗

PubMed1998

Molecular cloning and chromosomal mapping of type one serine/threonine protein phosphatases in Arabidopsis thaliana.

Type one serine/threonine protein phosphatases (PP1s) have been implicated in various processes of plant growth and development. In all plant species studied, PP1s are encoded by multigene families. Previous studies in our laboratory identified five Arabidopsis thaliana PP1 genes (TOPP1, TOPP2, TOPP3, TOPP4 and TOPP5). In the present study, we report the isolation of three additional PP1 genes (TOPP6, TOPP7 and TOPP8). Southern blot analyses indicate that these three newly isolated genes are single-copy genes in A. thaliana genome. All the three genes are expressed in roots, rosettes and flowers, although their expression levels appear to be lower than those of the five previously identified TOPP genes. Six of the eight TOPP genes were mapped to different positions on four of five A. thaliana chromosomes. Sequence comparison revealed that TOPP genes belong to different subgroups of plant PP1 genes, suggesting that they may encode proteins with distinct functions.

Amino Acid Sequence↗

PubMed1998

Seroconversion of type B to O erythrocytes using recombinant Glycine max alpha-D-galactosidase.

Recombinant alpha-D-galactosidase (rGal) from soybean (Glycine max) hydrolyzed the immunodominant alpha-D-galactose residue from the B epitope of red blood cells. This converted type B erythrocytes to type O which are "universally" transfusable. Type B red blood cells were obtained from four different donors and enzymatically converted. Cell function parameters, including red cell indices, pH, methemoglobin, carboxyhemoglobin, osmotic fragility, hemolysis, 2,3-diphosphoglycerate, cholinesterase, ATP, and antigen typing of treated cells were compared to controls. These pilot studies indicate that rGal could have potential biotechnical application in the production of universally transfusable red blood cells.

2,3-Diphosphoglycerate↗