Human erythrocyte calmodulin. Further chemical characterization and the site of its interaction with the membrane.

Human erythrocyte and bovine brain calmodulins were indistinguishable by tryptic peptide mapping, indicating that the primary sequence of the two proteins is either very similar or identical. Calcium binding determinations of human erythrocyte calmodulin, by equilibrium dialysis and fluorescence titration, were in close agreement with previous studies on other calmodulins. The calcium-activated adenosine triphosphatase which is stimulated by calmodulin was shown to be firmly associated with smooth erythrocyte plasma membranes devoid of spectrin and actin. Kinetic titration demonstrated that there are 4500 calmodulin binding sites per erythrocyte and that the turnover number of this calcium-activated adenosine triphosphatase is 3000 mumol of Pi . (mumol of site)-1 . min-1 which is similar to the turnover numbers of other transport adenosine triphosphatases. Furthermore, calmodulin stimulates calcium-activated adenosine triphosphatase by a simple enzyme-ligand association.     

Mechanisms of acetylcholine-mediated vasodilation in systemic arteries from mourning doves (Zenaida macroura)

For mammals, acetylcholine (ACh) promotes endothelium-dependent vasodilation primarily through nitric oxide (NO) and prostaglandin-mediated pathways, with varying reliance on endothelial-derived hyperpolarizing factors. Currently, no studies have been conducted on small systemic arteries from wild birds. We hypothesized that ACh-mediated vasodilation of isolated small arteries from mourning doves (Zenaida macroura) would likewise depend on endothelial-derived factors. Small resistance mesenteric and cranial tibial (c. tibial) arteries (80–150 μm, inner diameter) were cannulated and pre-constricted to 50 % of resting inner diameter with phenylephrine then exposed to increasing concentrations of ACh (10^?9–10^?5 M) or the NO donor, sodium nitroprusside (SNP; 10^?12–10^?3 M). For mesenteric arteries, ACh-mediated vasodilation was significantly blunted with the potassium channel antagonist tetraethylammonium chloride (TEA, 10 mM); whereas responses were only moderately impaired with endothelial disruption or inhibition of prostaglandins (indomethacin, 10 μM). In contrast, endothelial disruption as well as exposure to TEA largely abolished vasodilatory responses to ACh in c. tibial arteries while no effect of prostaglandin inhibition was observed. For both vascular beds, responses to ACh were moderately dependent on the NO signaling pathway. Inhibition of NO synthase had no impact, despite complete reversal of phenylephrine-mediated tone with SNP, whereas inhibition of soluble guanylate cyclase (sGC) caused minor impairments. Endothelium-independent vasodilation also relied on potassium channels. In summary, ACh-mediated vasodilation of mesenteric and c. tibial arteries occurs through the activation of potassium channels to induce hyperpolarization with moderate reliance on sGC. Prostaglandins likewise play a small role in the vasodilatory response to ACh in mesenteric arteries.     

Feline viruses in wildcats from Scotland

Few data are available on the prevalence of feline viruses in European wildcats (Felis silvestris). Previous surveys have indicated that wildcats may be infected with the common viruses of domestic cats, apart from feline immunodeficiency virus (FIV). In the present study, 50 wildcats trapped throughout Scotland (UK) between August 1992 and January 1997 were tested for evidence of viral infection. All were negative for FIV by several serological or virological methods. By contrast, 10% of the cats were positive for feline leukemia virus (FeLV) antigen and infectious virus was isolated from 13% of a smaller subset. Of the wildcats tested for respiratory viruses, 25% yielded feline calicivirus (FCV) and although no feline herpesvirus was isolated, 16% of the samples had neutralizing antibodies to this virus. Antibodies to feline coronavirus (FCoV) were found in 6% of samples. Feline foamy virus (FFV) was an incidental finding in 33% of samples tested. This study confirms that wildcats in Scotland are commonly infected with the major viruses of the domestic cat, except for FIV.     

Defining core habitat of local populations of the gray treefrog (Hyla versicolor) based on choice of oviposition site

Concern over amphibian population declines and loss of terrestrial and aquatic habitat have emphasized the need to define habitat requirements for each stage in a species' life history. The realization that pond-breeding amphibians spend most of their lives in the terrestrial environment suggests the need to protect terrestrial as well as aquatic habitat. Many studies on amphibian populations have focused on emigration from breeding sites to define habitat use; however these studies do not typically elucidate terrestrial activities of adults within the breeding season. We measured colonization rates of artificial pools by gray treefrogs (Hyla versicolor) at multiple distances from natural breeding ponds. We found a non-random distribution of egg deposition among distances, with 95% of eggs deposited within 15 m of the breeding pond. Additionally, we found that the time to first colonization of artificial pools increased with respect to distance. Our results indicate that adult gray treefrogs may travel up to 200 m within a breeding season, and that multiple breeding ponds may be considered part of a single population. We suggest that a minimum core terrestrial habitat of 60 m surrounding breeding sites is appropriate for protection of local populations of gray treefrogs.     

Sequence analysis of insecticidal genes from Xenorhabdus nematophilus PMFI296

Three strains of Xenorhabdus nematophilus showed insecticidal activity when fed to Pieris brassicae (cabbage white butterfly) larvae. From one of these strains (X. nematophilus PMFI296) a cosmid genome library was prepared in Escherichia coli and screened for oral insecticidal activity. Two overlapping cosmid clones were shown to encode insecticidal proteins, which had activity when expressed in E. coli (50% lethal concentration [LC(50)] of 2 to 6 microg of total protein/g of diet). The complete sequence of one cosmid (cHRIM1) was obtained. On cHRIM1, five genes (xptA1, -A2, -B1, -C1, and -D1) showed homology with up to 49% identity to insecticidal toxins identified in Photorhabdus luminescens, and also a smaller gene (chi) showed homology to a putative chitinase gene (38% identity). Transposon mutagenesis of the cosmid insert indicated that the genes xptA2, xptD1, and chi were not important for the expression of insecticidal activity toward P. brassicae. One gene (xptA1) was found to be central for the expression of activity, and the genes xptB1 and xptC1 were needed for full activity. The location of these genes together on the chromosome and therefore present on a single cosmid insert probably accounted for the detection of insecticidal activity in this E. coli clone. Although multiple genes may be needed for full activity, E. coli cells expressing the xptA1 gene from the bacteriophage lambda P(L) promoter were shown to have insecticidal activity (LC(50) of 112 microg of total protein/g of diet). This is contrary to the toxin genes identified in P. luminescens, which were not insecticidal when expressed individually in E. coli. High-level gene expression and the use of a sensitive insect may have aided in the detection of insecticidal activity in the E. coli clone expressing xptA1. The location of these toxin genes and the chitinase gene and the presence of mobile elements (insertion sequence) and tRNA genes on cHRIM1 indicates that this region of DNA represents a pathogenicity island on the genome of X. nematophilus PMFI296.