P-Element Insertion Alleles of Essential Genes on the Third Chromosome of Drosophila Melanogaster: Correlation of Physical and Cytogenetic Maps in Chromosomal Region 86e-87f

We have established a collection of 2460 lethal or semi-lethal mutant lines using a procedure thought to insert single P elements into vital genes on the third chromosome of Drosophila melanogaster. More than 1200 randomly selected lines were examined by in situ hybridization and 90% found to contain single insertions at sites that mark 89% of all lettered subdivisions of the Bridges' map. A set of chromosomal deficiencies that collectively uncover ~25% of the euchromatin of chromosome 3 reveal lethal mutations in 468 lines corresponding to 145 complementation groups. We undertook a detailed analysis of the cytogenetic interval 86E-87F and identified 87 P-element-induced mutations falling into 38 complementation groups, 16 of which correspond to previously known genes. Twenty-one of these 38 complementation groups have at least one allele that has a P-element insertion at a position consistent with the cytogenetics of the locus. We have rescued P elements and flanking chromosomal sequences from the 86E-87F region in 35 lines with either lethal or genetically silent P insertions, and used these as probes to identify cosmids and P1 clones from the Drosophila genome projects. This has tied together the physical and genetic maps and has linked 44 previously identified cosmid contigs into seven ``supercontigs' that span the interval. STS data for sequences flanking one side of the P-element insertions in 49 lines has identified insertions in the αγ element at 87C, two known transposable elements, and the open reading frames of seven putative single copy genes. These correspond to five known genes in this interval, and two genes identified by the homology of their predicted products to known proteins from other organisms.     

Cardiovascular adaptations to 10days of cycle exercise

Mier, Constance M., Michael J. Turner, Ali A. Ehsani, andRobert J. Spina. Cardiovascular adaptations to 10 days of cycleexercise. J. Appl. Physiol. 83(6):1900-1906, 1997.We hypothesized that 10 days of training wouldenhance cardiac output (CO) and stroke volume (SV) during peak exerciseand increase the inotropic response to -adrenergic stimulation. Tensubjects [age 26 ± 2 (SE) yr] trained on a cycleergometer for 10 days. At peak exercise, training increasedO₂ uptake, CO, and SV(P < 0.001). Left ventricular (LV)size and function at rest were assessed with two-dimensional echocardiography before (baseline) and after atropine injection (1.0 mg) and during four graded doses of dobutamine. LV end-diastolic diameter increased with training (P < 0.02), whereas LV wall thickness was unchanged. LV contractileperformance was assessed by relating fractional shortening (FS) to theestimated end-systolic wall stress(_ES). Training increased theslope of the FS-_ES relationship (P < 0.05), indicating enhancedsystolic function. The increase in slope correlated with increases inCO (r = 0.71,P < 0.05) and SV(r = 0.70,P < 0.05). The increase in bloodvolume also correlated with increases in CO(r = 0.80, P < 0.01) and SV (r = 0.85, P < 0.004). These datashow that 10 days of training enhance the inotropic response to-adrenergic stimulation, associated with increases in CO and SVduring peak exercise.

     

Ataxia with Vitamin E Deficiency: Refinement of Genetic Localization and Analysis of Linkage Disequilibrium by Using New Markers in 14 Families

Ataxia with vitamin E deficiency (AVED) is an autosomal recessive disease characterized clinically by neurological symptoms with often striking resemblance to those of Friedreich ataxia. This disorder has been reported previously as familial isolated vitamin E deficiency. We have mapped recently the AVED locus to a 5-cM confidence interval on chromosome 8q by homozygosity mapping in six Mediterranean families. We have now analyzed six new and two previously described families and demonstrate genetic homogeneity despite important clinical variability and wide geographic origins. Analysis of nine new tightly linked microsatellite markers, including four characterized in this study, revealed a predominant but not unique mutation in northern African populations, where this condition is more frequent. Haplotype analysis but also classical recombinations allowed us to refine the AVED position to a 1-cM interval. A YAC contig over this interval was constructed from marker STSs and YAC fingerprint data, in order to facilitate the search of the AVED gene.     

Prothrombin cleavage by human vascular smooth muscle cells: A potential alternative pathway to the coagulation cascade

Thrombin is a potent mitogen for human vascular smooth muscle cells (HVSMC) and its enzymatic activity is required for this function. The present study demonstrates that prothrombin is also mitogenic for HVSMC due to the generation of enzymatically active thrombin which occurs upon incubation of prothrombin with the cells. Analysis by SDS-PAGE, immunoblotting, and amino acid sequencing revealed that prothrombin incubated with HVSMC undergoes limited proteolysis. Prethrombin 1 was formed through cleavage at R¹⁵⁵-S¹⁵⁶. Cleavage at R²⁷¹-T²⁷² generated fragment 1.2 and prethrombin 2 whilst cleavage at R²⁸⁴-T²⁸⁵ yielded truncated prothrombin 2 (prethrombin 2′). However, cleavage at R³²⁰-I³²¹ which, during prothrombin activation produces two-chain α-thrombin, was not detectable. Studies on HVSMC-conditioned medium revealed that a similar pattern of prothrombin cleavage occurred by a cell-secreted factor(s). Amidolytic activity analysis indicated that 1–3% catalytically active thrombin-like activity was generated upon incubation of prothrombin with HVSMC-conditioned medium. By treating conditioned medium with various classes of proteinase inhibitors or hirudin, it was determined that prothrombin is cleaved by a cell-derived serine proteinase-like factor(s) at R²⁷¹-S²⁷² and by α-thrombin at R¹⁵⁵-S¹⁵⁶ and R²⁸⁴-T²⁸⁵. Antibodies neutralising the activity of either urokinase, tissue plasminogen activator, or factor Xa failed to alter the prothrombin cleaving activity of conditioned medium. This activity which may catalyse an alternative pathway for the generation of thrombin, was eluted from a gel filtration column as a single peak with apparent molecular mass of 30–40 kDa. © 1995 Wiley-Liss, Inc.     

Benthic foraminiferal fauna during the time of the Indian-Asian contact, in southern Balochistan, Pakistan

Cretaceous and early Paleocene benthic foraminifera were studied from one section along the western Gaj River, southern Balochistan, Pakistan, to reconstruct the paleoenvironment of the Tethys Sea during the Indian-Asian contact. We recognize three lithostratigraphic units in ascending order: the Mughal Kot Formation, the Pab Sandstone, and the Jamburo Group. Both the Maastrichtian Mughal Kot Formation, which consists of shale with grey marly limestone, and the Maastrichtian Pab Sandstone, which consists of quartzose sandstone, indicate an open ocean environment as they have diversified planktic and benthic foraminiferal assemblages. The Maastrichtian-Paleocene Jamburo Group, consisting of dark grey, calcareous shale and marlstone with some sulfide grains, is characterized by low diversities of benthic assemblages. The change to the lower diversities may be associated with the development of poor circulation of deeper water that was caused by narrowing of the Tethys Sea.The Trochammina spp. Assemblage from the Jamburo Group, which can be correlated with flysch-type agglutinated foraminiferal assemblages, has a low benthic species diversity, indicating an unfavorable condition for calcareous foraminifera because of the development of oxygen-depleted water. The absolute abundance of agglutinated specimens shows a remarkable change from low numbers in the Maastrichtian to high ones in the Paleocene. The benthic foraminiferal evidence supports the hypothesis that the collision of the Asian and Indian plates occurred near the end of the Cretaceous.     

Drug-induced circling preference in rats

Drugs of abuse, such as phencyclidine (PCP), methamphetamine (METH), and cocaine (COC) are known to affect several behaviors in rats, such as motor activity, stereotypy, and circling. In this study, we evaluated whether these drugs produce circling preferences in the presence or absence of unilateral 6-hydroxydopamine (6-OHDA)-induced lesions of the caudate nucleus. Adult male CD rats were lesioned with 10 μg 6-OHDA/site. Animals were dosed with PCP (15 mg/kg, ip), its congener, (+) MK-801 (0.15 mg/kg, ip), METH (2 mg/kg, ip), COC (60 mg/kg, ip), or apomorphine (0.2 mg/kg, ip). circling preference was recorded in control and lesioned rats for 2 h before animals were sacrificed to determine monoamine levels by HPLC/EC. In control animals, administration of these drugs produced 60–70% left circling. In, lesioned animals, these drugs produced 78–90% ipsilateral (toward the lesion) circling, except apomorphine, which produced 60–80% contralateral (away from the lesion) circling. Dopamine (DA) and its metabolites 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) concentrations significantly decreased ipsilaterally in lesioned caudate nucleus (CN) and substantia nigra (SN). However, no significant changes were observed in nucleus accumbens (NA) and olfactory tubercles (OT). These data demonstrate that drugs of abuse like PCP, its congener (+) MK-801, METH, and COC produce a greater preference to turn toward the left than the right, a finding similar to that found in human psychosis. Since 6-OHDA lesions enhanced the circling bias and depleted DA and its metabolites DOPAC and HVA, it also suggests that the dopaminergic system may be involved in the circling behavior.     

Effect of neonatal monosodium glutamate on the activities of glutamate dehydrogenase and aminotransferases in the circumventricular organs of rat brain

Summary Glutamate (Glu) the major amino acid in mammalian brain and most dietary proteins possesses neurotransmitter as well as neurotoxic properties. We administered monosodium glutamate (MSG) 4 mg/g bwt, sc on postnatal day (PND) 1 through 10 to rats on alternate days or daily and sacrificed them on PND 45 or PND 90 respectively. The activities of glutamate dehydrogenase and aminotransferases were evaluated in the circumventricular organs of brain. Results show that neonatal MSG produces alterations in glutamate metabolism in blood-brain-barrier deficient regions.     

A beta-linked mannan inhibits adherence of Pseudomonas aeruginosa to human lung epithelial cells

Adherence through carbohydrate-binding adhesins is an earlystep in colonization of the lung by gram-negative organisms,and because published data indicate that binding involves mannosegroups, we tested the ability of a ß-linked acetylmannan(acemannan) to inhibit adherence of Pseudomonus aeruginosa tocultures of human lung epithelial cells. Adherence of radiolabelledP.aeruginosa to A549 cells (a type II-like pneurnocyte line)increased linearly with the duration of the incubation. Acemannaninhibited adherence of bacteria, and the extent of inhibitionwas related to the concentration of the mannan. Inhibition requiredcontinued contact between acemannan and the target epithelialcells; cells washed free of acemannan no longer discouragedbacterial binding. Comparison of binding between seven strainsof P.aeruginosa indicated that fewer mucoid than non-mucoidbacteria adhered, but binding of either phenotype was inhibitedby acemannan. Mannose methyl -D-mannopyranoside, methyl ß-D-mannopyrannosideand dextran did not affect adherence of any of the non-mucoidstrains. Mannose inhibited adherence by one mucoid strain, butnot the other, indicating differences between strains of thesame phenotype. Since prior treatment of epithelial cells withconcanavalin A did not affect acemannan-induced inhibition ofbacterial adherence, we concluded that the inhibitory effectof acemannan probably does not involve mannose-containing receptors. bacterial-host interactions lung epithelium mucoid strains non-mucoid strains     

Genetic regulation of gibberellin deactivation in Pisum

The regulation of gibberellin (GA) deactivation was examined using the sin (slender) mutation in the garden pea (Pisum sativum L.). This mutation blocks the deactivation of GA₂₀, the precursor of the bioactive GA₁. Firstly, crosses were made to combine sin with the GA biosynthesis mutations na, lhⁱ and le-3. The combination sin na produced a novel phenotype, with long (‘slender’) basal internodes and extremely short (‘nana’) upper internodes. In contrast, the double mutant sin lhⁱ was phenotypically dwarf. The mutation sin causes an accumulation of GA₂₀ in maturing seeds, and this was unaffected by na, since the na mutation is not expressed in seeds. In contrast, lhⁱ seeds did not accumulate GA₂₀, since lhⁱ imposes an early block on GA biosynthesis. Secondly, the effects of sin on several steps in GA deactivation were investigated. In maturing seeds, the mutation sin blocks two steps in GA₂₀ metabolism, namely, GA₂₀ to GA₂₉, and GA₂₉ to GA₂₉-catabolite. In the vegetative plant, on the other hand, sin blocked the step GA₂₀ to GA₂₉, but not GA₂₉ to GA₂₉-catabolite; the steps GA₂₀ to GA₈₁ and GA₂₀ to GA₁ were also not impaired in this mutant. It is clear that the effects of sin, like those of na, are strongly organ-specific. The presence of separate enzymes for the steps GA₂₀ to GA₂₉ and GA₂₉ to GA₂₉-catabolite was suggested by the observation that GA₈ inhibited the latter step, but not the former, and by the inability of GA₂₀ and GA₂₉ to inhibit each other's metabolism. It is suggested that the Sin gene may be a regulatory gene controlling the expression of two structural genes involved in GA deactivation.     

Hb J-Singa (alpha-78 Asn leads to Asp), a newly discovered hemoglobin variant with the same amino acid substitution as one of the two present in Hb J-Singapore (alpha-78 Asn leads to, alpha-79 Ala leads to Gly)

A new fast-moving alpha-chain Hb variant with an Asn leads to Asp substitution at position alpha-78 was found in a French-Acadian family living in Eastern Canada. The identical substitution was reported in Hb J-Singapore, which also had an additional Ala leads to Gly substitution at position alpha-79. The new variant, which did not result in any clinical symptoms, was named accordingly, Hb J-Singa.