Thermoregulatory behavior of the bluespotted sunfish,Enneacanthus Gloriousus

Twenty bluespotted sunfish, Enneacanthus gloriosus, were individually tested for thermoregulatory behavior in an electronic shuttlebox. The first fishes of this genus to be tested for thermoregulatory behavior, these individuals were captured near the northern limit of their zoogeographic range, in northeastern Pennsylvania. The mean final preferendum from pooled data of all 20 fish was 28.5°C; there were no significant differences between day and night.     

In vitro adhesion of tufted oral streptococci toBacterionema matruchotii

The adhesive interaction between the ectosymbionts of the corn cob configuration (CCC), a naturally occurring bacterial consortium in human dental plaque, was studied. In vitro association was produced in mixed cultures consisting ofBacterionema matruchotii and streptococci resemblingStreptococcus sanguis previously isolated from human CCC. Phase-contrast, selective immunofluorescence, and scanning electron microscopy showed that in this aggregative system, a tuftlike polar structure typical of the coccal epibiont mediated binding to the core filament. This aggregative model provides evidence that a specialized structure on the cell surface of epiphytic organisms may participate in interbacterial adherence.     

Behavioral thermoregulation by Ammocoete larvae of the sea lamprey (Petromyzon marinus) in an electronic shuttlebox

Ammocoete larvae of the sea lamprey Petromyzon marinus, a member of the primitive vertebrate class Agnatha, were tested for thermoregulatory behavior in an electronic shuttlebox (ichthyotron). The final preferendum derived from pooled data for 24 individually tested ammocoetes was characterized by a mean of 13.6 ± 0.17 (s.e.m.)°C, a mode and median of 140°C, and a range of voluntarily occupied temperatures from 10–19°C over a 3-day period.     

The molar ratio of the two major polypeptide components of human high density lipoprotein.

Human high density lipoprotein (HDL) and its subfractions (HDL2 and HDL3) were separated by ultracentrifugation and the molar ratio of the two major polypeptide chains apo-Gln-I and apo-Gln-II was determined by fluorescence tagging of sodium dodecyl sulfate-denatured proteins combined with polyacrylamide disc gel electrophoresis. Using purified apo-Gln-I and apo-Gln-II standards, it was found that holo HDL, holo HDL2, and holo HDL3 from all plasma samples contained a molar ratio of apo-Gln-I to the disulfide-bound dimer of apo-Gln-II of 2:1, that is a 1:1 ratio in terms of each species of polypeptide chain. The method described is useful for making repeated and rapid measurements on microgram quantities of intact lipoproteins.     

Functional and molecular characterization of an anion exchanger in airway serous epithelial cells

Serous cells secreteCl and HCO₃ and play an importantrole in airway function. Recent studies suggest that aCl/HCO₃ anion exchanger (AE) maycontribute to Cl secretion by airway epithelial cells.However, the molecular identity, the cellular location, and thecontribution of AEs to Cl secretion in serous epithelialcells in tracheal submucosal glands are unknown. The goal of thepresent study was to determine the molecular identity, the cellularlocation, and the role of AEs in the function of serous epithelialcells. To this end, Calu-3 cells, a human airway cell line with aserous-cell phenotype, were studied by RT-PCR, immunoblot, andelectrophysiological analysis to examine the role of AEs inCl secretion. In addition, the subcellular location of AEproteins was examined by immunofluorescence microscopy. Calu-3 cellsexpressed mRNA and protein for AE2 as determined by RT-PCR and Westernblot analysis, respectively. Immunofluorescence microscopy identified AE2 in the basolateral membrane of Calu-3 cells in culture and rattracheal serous cells in situ. InCl/HCO₃/Na⁺-containingmedia, the 8-(4-chlorophenylthio)adenosine 3',5'-cyclic monophosphate(CPT-cAMP)-stimulated short-circuit anion current (I_sc) was reduced by basolateral but not byapical application of 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid(50 µM) and 4,4'-dinitrostilbene-2,2'-disulfonic acid [DNDS (500 µM)], inhibitors of AEs. In the absence of Na⁺ in thebath solutions, to eliminate the contributions of the Na⁺/HCO₃ andNa⁺/K⁺/2Cl cotransporters toI_sc, CPT-cAMP stimulated a small DNDS-sensitive I_sc. Taken together with previous studies, theseobservations suggest that a small component of cAMP-stimulatedI_sc across serous cells may be referable toCl secretion and that uptake of Cl acrossthe basolateral membrane may be mediated by AE2.

     

Automation of protein assays with a microcomputer interfaced to a spectrophotometer

We have developed a system for semi-automatic protein quantitation assays based on an inexpensive, general-purpose microcomputer. A commercial curve fitting program was modified for direct data input from the spectrophotometer via an analog to digital converter. The software provides specific prompts for protein assays resulting in a rapid and user friendly, semi-automated system. A listing of the protein assay results are printed, or stored to a magnetic disk, along with a graphic representation of the standard curve.     

Locus Heterogeneity for Waardenburg Syndrome is Predictive of Clinical Subtypes

Waardenburg syndrome (WS) is a dominantly inherited and clinically variable syndrome of deafness, pigmentary changes, and distinctive facial features. Clinically, WS type I (WS1) is differentiated from WS type II (WS2) by the high frequency of dystopia canthorum in the family. In some families, WS is caused by mutations in the PAX3 gene on chromosome 2q. We have typed microsatellite markers within and flanking PAX3 in 41 WS1 kindreds and 26 WS2 kindreds in order to estimate the proportion of families with probable mutations in PAX3 and to study the relationship between phenotypic and genotypic heterogeneity. Evaluation of heterogeneity in location scores obtained by multilocus analysis indicated that WS is linked to PAX3 in 60% of all WS families and in 100% of WS1 families. None of the WS2 families were linked. In those families in which equivocal lod scores (between −2 and +1) were found, PAX3 mutations have been identified in 5 of the 15 WS1 families but in none of the 4 WS2 families. Although preliminary studies do not suggest any association between the phenotype and the molecular pathology in 20 families with known PAX3 mutations and in four patients with chromosomal abnormalities in the vicinity of PAX3, the presence of dystopia in multiple family members is a reliable indicator for identifying families likely to have a defect in PAX3.     

High titer production of tetracenomycins by heterologous expression of the pathway in a Streptomyces cinnamonensis industrial monensin producer strain

Streptomyces cinnamonensis C730.1 and C730.7, are industrially mutagenized strains that produce moderate and high levels of the polyketide polyether antibiotic monensin A, respectively, in an oil-based fermentation medium. The possibility that these strains could be used for high titer production of a heterologous polyketide product was investigated by expression of the entire tetracenomycin (TCM) biosynthetic pathway using an integrative plasmid, pSET154. Expression in C730.1 led to stable production of ~0.44 g/l TCM C (the final biosynthetic product) and ~2.69 g/l TCM A2 (the penultimate biosynthetic product), and resulted in a 40% decrease in monensin production. Expression in the C730.7 led to higher levels of TCMs, ~0.6 g/l TCM C and ~4.35 g/l TCM A2, without any detectable decrease in the higher titer monensin production. Abrogation of monensin production in this strain through deletion of the corresponding biosynthetic genes did not lead to higher levels of TCM products. In the case of the C730.7 host, 85% of the TCM C and virtually all of the TCM A2 were intracellular, suggesting feedback inhibition leads to the accumulation of the final pathway intermediate. These observations contrast those made for the native producer Streptomyces glaucescens where the predominant product is TCM C and TCM titers are significantly lower levels (~0.3 g/l), and demonstrate the potential utility of S. cinnamonensis strains as heterologous hosts for high level expression of a variety of polyketide synthase derived products.