Cytogenetics for the model system Arabidopsis thaliana

A detailed karyotype of Arabidopsis thaliana is presented using meiotic pachytene cells in combination with fluorescence in situ hybridization. The lengths of the five pachytene bivalents varied between 50 and 80 microns, which is 20-25 times longer than mitotic metaphase chromosomes. The analysis confirms that the two longest chromosomes (1 and 5) are metacentric and the two shortest chromosomes (2 and 4) are acrocentric and carry NORs subterminally in their short arms, while chromosome 3 is submetacentric and medium sized. Detailed mapping of the centromere position further revealed that the length variation between the pachytene bivalents comes from the short arms. Individual chromosomes were unambiguously identified by their combinations of relative lengths, arm-ratios, presence of NOR knobs and FISH signals with a 5S rDNA probe and chromosome specific DNA probes. Polymorphisms were found among six ecotypes with respect to the number and map positions of 5S rDNA loci. All ecotypes contain 5S rDNA in the short arms of chromosomes 4 and 5. Three different patterns were observed regarding the presence and position of a 5S rDNA locus on chromosome 3. Repetitive DNA clones enabled us to subdivide the pericentromeric heterochromatin into a central domain, characterized by pAL1 and 106B repeats, which accommodate the functional centromere and two flanking domains, characterized by the 17 A20 repeat sequences. The upper flanking domains of chromosomes 4 and 5, and in some ecotypes also chromosome 3, contain a 5S rDNA locus. The detection of unique cosmids and YAC sequences demonstrates that detailed physical mapping of Arabidopsis chromosomes by cytogenetic techniques is feasible. Together with the presented karyotype this makes Arabidopsis a model system for detailed cytogenetic mapping.     

Inhibition of purinergic transmission by prostaglandin E1 and E2 in the guinea-pig vas deferens: an electrophysiological study

1. The effects of prostaglandin E1 (PGE1) and E2 (PGE2) on postjunctional electrical activity in the guinea-pig vas deferens evoked by sympathetic nerve stimulation were investigated using both intracellular and focal extracellular recording techniques in vitro. 2. Bath application of PGE1 (1-100 nM) or PGE2 (0.1-100 nM) concentration-dependently inhibited the amplitudes of all excitatory junction potentials (e.j.ps) evoked during short trains of stimuli (10 stimuli at 1 Hz). Increasing the duration of nerve stimulation (100 stimuli at 1 Hz) did not overcome this inhibitory effect. At these concentrations PGE1 and PGE2 were without any apparent inhibitory effect on the amplitudes of spontaneous e.j.ps. 3. Local application of PGE1 (10-100 nM) or PGE2 (10-30 nM) markedly reduced the frequency of occurrence of excitatory junction currents (e.j.cs) evoked by trains of 20-100 stimuli at 1 to 4 Hz without changing the amplitudes of spontaneous e.j.cs or the configuration of the nerve terminal impulse. 4. In the presence of PGE1 or PGE2, raising the frequency of stimulation (from 1 to 4 Hz), increased the likelihood of e.j.c. occurrence. 5. The postjunctional electrical activity recorded in the guinea-pig vas deferens is believed to be due to ATP released from the sympathetic nerve endings. Thus the present study demonstrates that both PGE1 and PGE2 powerfully inhibit quantal ATP release in the guinea-pig vas deferens.     

Antibody binding to a peptide but not the whole protein by recognition of the C-terminal carboxy group

Factor V and protein S are cofactors of activated protein C (APC) which accelerate APC-mediated factor VIII inactivation. The effects of factor V and protein S were quantitated in a reaction system in which plasma factor VIII was inactivated by APC and the loss of factor VIII activity was monitored in a factor X-activating system in which a chromogenic substrate was used to probe factor Xa formation. Factor V increased the rate of APC-mediated factor VIII inactivation in a dose-dependent manner in representative plasma samples with protein S or factor V deficiency, abnormal factor V (heterozygous or homozygous for factor VR506Q), or a combination of heterozygous protein S deficiency and heterozygous factor VR506Q. This effect was much less pronounced in the plasma samples with a decreased protein S level, but the impaired response in these plasmas was corrected by addition of protein S, indicating that both factor V and protein S are required for optimal inactivation of factor VIII by APC. The effects of factor V and protein S were also studied in a reaction system with purified proteins. APC-catalysed factor VIII inactivation was enhanced 3.7-fold in the presence of 1.1 nM factor V and 1.5-fold in the presence of 2.4 nM protein S. When both 1.1 nM factor V and 2.4 nM protein were present the rate enhancement was 11-fold. Factor V is a more potent cofactor than protein S, as can be concluded from the fact that 0.04 nM factor V gave the same stimulation as 2.4 nM protein S. Protein S lost its cofactor function after complexation with C4b binding protein, which indicates that it is free protein S that acts as a cofactor. To investigate the effect of the R506Q mutation in factor V on APC-mediated factor VIII inactivation, factor V was purified from the plasma of patients homozygous for factor VR506Q. In the absence of protein S, factor VR506Q did not enhance factor VIII inactivation by APC, but in the presence of 2.4 nM protein S a slight enhancement was observed. The APC cofactor activity of factor V was lost when factor V was activated with thrombin or with the factor V activator from Russell's viper venom. These data indicate that optimal inactivation of factor VIII by APC requires the presence of an intact factor V molecule and free protein S.     

The molecular basis of beta thalassaemia in Punjabi and Maharashtran Indians includes a multilocus aetiology involving triplicated alpha-globin loci

We have analysed 201 beta-thalassaemia (beta-thal) genes from natives of the Punjab (156) and Maharashtra states of India and found the causative mutation in 200 of them. The most common beta-globin gene mutations differed significantly between these two groups and between these groups and Indian immigrants in the U.S.A. and the U.K. In the Punjabi Indians the IVS-1, nt 1 (G-T) mutation accounted for nearly one-quarter of beta-thal genes, whereas it was 5% or less in the other groups. Likewise, the cap + 1 mutation was much more prevalent in the Punjabis, whereas the nonsense codon 15 allele had a higher frequency in the Maharashtrans of the Bombay region. The common IVS-1, nt5 allele had a frequency of 60% of beta-thal genes in the Maharastrans, 35% in North American immigrants, and only 23% in the Punjabis. Two-thirds of all beta-thal genes in Punjab were found in the merchant caste (Khatri-Arora), whereas the menial caste (Shudra) was highly represented among those with beta-thal genes in Maharashtra. Two novel beta-globin alleles were each found once; a frameshift codon 55 (+A) in Maharashtrans and a frameshift codons 47-48 (+ATCT) in Punjabis. Of three Punjabi patients with beta-thal intermedia in whom only a single severe beta-globin gene mutation was found, two had six alpha-globin genes (homozygosity for a triplicated alpha-globin locus) instead of the normal alpha-globin gene number of four. Thus, these two individuals had a multilocus aetiology of beta-thal and their parents have the unusual recurrence risk of 1 in 8 for conceiving a third with beta-thal intermedia. Since 15% of 126 alpha-globin clusters studies in Punjabis contained either single (10%) or triplicated (5%) alpha-globin genes, the alpha-globin gene number is a frequent modifier of the phenotype of beta-thal in this ethnic group.     

Bridging student health risks and academic achievement through comprehensive school health programs

In the National Action Plan for Comprehensive School Health Education, representatives for over 40 health, education, and social service organizations viewed education and health as independent systems. Participation concluded that healthy children learn better, and they cautioned that no curriculum can compensate for deficiencies in student health status. While literature confirms the complexity of health issues confronting today's students, schools face enormous pressure to improve academic skills. Local school leaders and stakeholders often remain unconvinced that improving student health represents a means to achieving improved academic outcomes. A rich body of literature confirms a direct link between student health risk behavior and education outcomes, education behavior, and student attitudes about education. This article summarizes relevant information concerning the health risk behavioral categories of intentional injuries; tobacco; alcohol, and other drugs; dietary, physical activity, and sexual risk behaviors.     

Assessment of platelet activation by coronary sinus blood sampling during balloon angioplasty and directional coronary atherectomy

Three markers of platelet activation (platelet-derived microparticles, fibrinogen binding and expression of P-selectin) were assessed by flow cytometry during diagnostic coronary angiography and therapeutic coronary interventions. In 24 patients undergoing diagnostic angiography, blood was collected to determine if our sampling techniques or coronary angiography caused platelet activation. Changes during diagnostic angiography were used to establish baseline values and interpret changes during coronary interventions. In 21 patients, blood samples were obtained at 5 time points during percutaneous transluminal coronary angioplasty (PTCA) (n = 17) or directional coronary atherectomy (DCA) (n = 4). During coronary interventions, mean values for the percentage of platelets expressing P-selectin or binding fibrinogen increased, but with considerable variation among patients. Individual responses for platelet activation markers in each patient were characterized using a twofold increase to indicate elevation related to the intervention. Patients were classified as having complicated or uncomplicated procedures based on the presence of acute closure, dissection, or thrombus observed by angiography. There were no differences in the percentage of elevated markers between patients with uncomplicated (12.5%) and complicated (19%) PTCA procedures. However, patients treated with DCA had more elevated markers (38%) than those treated with PTCA (15%) (p = 0.04). Our data suggest that the extent of platelet activation in individual patients cannot be predicted by common angiographic findings or complications. More markers of platelet activation were present after DCA and may reflect a greater degree of vascular trauma associated with this procedure.