Unrestricted recognition of a nonpeptide antigen by CD8+ cytolytic T lymphocytes

CD8+ murine CTL that are specific for an unusual nonpeptide Ag, the heme moiety of hemoglobin, have been derived by in vitro stimulation of spleen cells with hemin. Such CTL demonstrate a requirement for the expression of class I Ag on target cells, yet appear to be unrestricted to the extent that both syngeneic and allogeneic targets precoated with hemin are sensitive to lysis. A series of CTL clones with specificity for hemin was derived from C57BL/6 mice. They exhibited the same type of promiscuous recognition that was observed in CTL populations from a number of different strains. The possibility that hemin acts as a nonspecific mediator of lysis by CTL was ruled out by the fact that a variety of CTL populations and clones specific for different Ag did not exhibit hemin-specific lysis. Some explanations offered to explain these results include 1) the possibility that hemin is recognized by binding to a site on the MHC other than the Ag-binding groove, and 2) the possibility that TCR recognition of a rigid molecule, such as hemin, may be less sensitive to polymorphic variation in the MHC than is recognition of a conventional peptide Ag whose conformation may differ significantly when bound to MHC molecules whose sequences differ within the Ag-binding groove.     

Visual evoked potentials in a sample of schizophrenic patients

Variations in evoked potentials utilizing a photic stimulus in a sample of psychiatric patients compared to a healthy sample were evaluated. A group of patients diagnosed as schizophrenic was tested against a sample of healthy volunteers in a trial combining visual evoked potentials and a simultaneous cognitive processing. The stimulus was a checkerboard pattern presented under three different conditions. The results indicate diminished P100 and lack the reactivity associated with cognitive processes in schizophrenic group. The P200 component also lacked, in the inpatient group the changes associated with the performance of the trial. Finally the multiple P300 component was shortened in latency and decreased in amplitude in the schizophrenia group. Besides, P300 interhemispheric shifts related to trials, were commonly inverted in schizophrenics. Results are interpreted as a lacked interhemispheric coordination in schizophrenics, rather than a fixed hemispheric alteration. Likewise, an attenuation in processing from specific cortical areas to association cortex is concluded.     

Distribution of neuropeptide Y-like immunoreactive fibers in the cat thalamus

Neuropeptide Y-like immunoreactivity was studied in the thalamus of the cat using an indirect immunoperoxidase method. The densest network of immunoreactive fibers was observed in the nucleus (n.) paraventricularis anterior. In the anterior, intralaminar and midline thalamic nuclei, as well as in the n. geniculatum medialis, n. geniculatum lateralis, n. habenularis lateralis, n. medialis dorsalis, n. lateralis posterior and n. pulvinar a low density of neuropeptide Y-like immunoreactive fibers was observed. Neuropeptide Y-like fibers were totally absent in the n. ventralis lateralis, n. ventralis medialis, n. ventralis postero-medialis and n. ventralis postero-lateralis. In addition, neuropeptide Y-like perikarya were found in the n. parafascicularis, n. suprageniculatus, n. geniculatum lateralis ventralis, n. medialis dorsalis and n. lateralis posterior.     

Genetically-based plant resistance traits affect arthropods,fungi, and birds

We examine how the distribution of a leafgalling aphid (Pemphigus betae) affects other species associated with natural stands of hybrid cottonwoods (Populus angustifolia x P. fremontii). Aphid transfers on common-garden clones and RFLP analysis show that resistance to aphids in cottonwoods is affected by plant genotype. Because susceptible trees typically support thousands of galls, while adjacent resistant trees have few or none, plant resistance traits that affect the distribution of this abundant herbivore may directly and/or indirectly affect other species. We found that the arthropod community of aphid-susceptible trees had 31% greater species richness and 26% greater relative abundance than aphid-resistant trees. To examine direct and indirect effects of plant resistance traits on other organisms, we experimentally excluded aphids and found that abundances and/or foraging behavior of arthropods, fungi, and birds were altered. First, exclusion of gall aphids on susceptible trees resulted in a 24% decrease in species richness and a 28% decrease in relative abundance of the arthropod community. Second, exclusion of aphids also caused a 2- to 3-fold decrease in foraging and/or presence of three taxa of aphid enemies: birds, fungi, and insects. Lastly, aphidexclussion resulted in a 2-fold increase in inquilines (animals who live in abodes properly belonging to another). We also found that fungi and birds responded to variation in gall density at the branch level. We conclude plant resistance traits affect diverse species from three trophic levels supporting a bottom-up influence of plants on community structure.     

Mutations in the consensus ATP-binding sites of XcpR and PilB eliminate extracellular protein secretion and pilus biogenesis in Pseudomonas aeruginosa.

The process of extracellular secretion in Pseudomonas aeruginosa requires specialized machinery which is widely distributed among bacteria that actively secrete proteins to the extracellular medium. One of the components of this machinery is the product of the xcpR gene, which is homologous to pilB, a gene encoding a protein essential for the biogenesis of type IV pili. Both XcpR and PilB are characterized by the presence of a conserved ATP-binding motif (Walker sequence). The codons of highly conserved glycine residues within the Walker sequences of xcpR and pilB were altered to encode a serine, and the effects of these substitutions were examined. Bacteria expressing mutant XcpR or PilB were unable to secrete exotoxin A or assemble pili, respectively. In addition, high-level expression of mutant XcpR in wild-type P. aeruginosa led to a pleiotropic extracellular secretion defect, resulting in the periplasmic accumulation of enzymes that are normally secreted from the cell. These studies show that the putative ATP-binding sites of XcpR and PilB are essential for their functions in protein secretion and assembly of pili, respectively. Moreover, the observed dominant negative phenotype of mutant XcpR suggests that this protein functions as a multimer or, alternatively, interacts with another essential component of the extracellular protein secretion machinery.     

Fluorescent in situ hybridization of the telomere repeat sequence in hamster sperm nuclear structures

The flat, hooked-shaped architecture of the hamster sperm nucleus makes this an excellent model for in situ hybridization studies of the three dimensional structure of the genome. We have examined the structure of the telomere repeat sequence (TTAGGG)_n with respect to the various nuclear structures present in hamster spermatozoa, using fluorescent in situ hybridization. In fully condensed, mature sperm nuclei, the telomere sequences appeared as discrete spots of various sizes interspersed throughout the volume of the nuclei. While the pattern of these signals was non-random, it varied significantly in different nuclei. These discrete telomere foci were seen to gradually lengthen into linear, beaded signals as sperm nuclei were decondensed, in vitro, and were not associated with the nuclear annulus. We also examined the relationship of telomeres to the sperm nuclear matrix, a residual nuclear structure that retains the original size and shape of the nucleus. In these structures the DNA extends beyond the perimeter of the nucleus to form a halo around it, representing the arrangement of the chromosomal DNA into loop domains attached at their bases to the nuclear matrix. Telomere signals in these structures were also linear and equal in length to those of the decondensed nuclei, and each signal represented part of a single DNA loop domain. The telomeres were attached at one end to the nuclear matrix and extended into the halo. Sperm nuclear matrices treated with Eco RI retained the telomere signals. These data support sperm DNA packaging models in which DNA is coiled into discrete foci, rather than spread out linearly along the length of the sperm nucleus.     

Real-time monitoring of biomass during Escherichia coli high-cell-density cultivations by in-line photon density wave spectroscopy

An efficient monitoring and control strategy is the basis for a reliable production process. Conventional optical density (OD) measurements involve superpositions of light absorption and scattering, and the results are only given in arbitrary units. In contrast, photon density wave (PDW) spectroscopy is a dilution-free method that allows independent quantification of both effects with defined units. For the first time, PDW spectroscopy was evaluated as a novel optical process analytical technology tool for real-time monitoring of biomass formation in Escherichia coli high-cell-density fed-batch cultivations. Inline PDW measurements were compared to a commercially available inline turbidity probe and with offline measurements of OD and cell dry weight (CDW). An accurate correlation of the reduced PDW scattering coefficient µ_s′ with CDW was observed in the range of 5–69 g L⁻¹ (R² = 0.98). The growth rates calculated based on µ_s′ were comparable to the rates determined with all reference methods. Furthermore, quantification of the reduced PDW scattering coefficient µ_s′ as a function of the absorption coefficient µ_a allowed direct detection of unintended process trends caused by overfeeding and subsequent acetate accumulation. Inline PDW spectroscopy can contribute to more robust bioprocess monitoring and consequently improved process performance.