The mendelian inheritance of a human X chromosome-specific DNA sequence polymorphism and its use in linkage studies of genetic disease

Summary A recombinant DNA sequence, RB6, was isolated from a human X chromosome library and shown to be X-specific by hybridisation to DNA from a human-mouse somatic cell hybrid containing X as the only human chromosome. The cloned sequence was located on the long arm distal to Xq13 using a human-mouse somatic cell hybrid containing a partial human X chromosome. DNA samples isolated from control human females were digested with the restriction enzyme MspI, and analysed by blotting and hybridisation to the radioactive cloned DNA. Eight of 14 individuals from a random population showed a single hybridising band 7.5 kilobase pairs (kb) in length, but six showed an additional band 10.1 kb in length. DNA from 12 members of a family with X-linked thyroxine-binding globulin deficiency was analysed for the segregation of this polymorphism. The results show that the polymorphism is inherited in a Mendelian fashion, and that the disease locus is not closely linked to the polymorphic site. Such polymorphisms will be useful as markers for chromosome mapping and for the antenatal diagnosis of genetic diseases.     

Organization of a family of highly repetitive sequences within the human genome

Recombinant clones containing the highly repetitive human DNA sequence approximately 340 base-pairs in length obtained after EcoRI digestion (αRI-DNA) were cloned in plasmid pAT153. Two clones contained a single copy of the αRI-DNA sequence, and the third had an insert with two copies of the sequence in tandem. When radioactive recombinant DNA was hybridized to total human DNA partially digested with EcoRI, a series of multiple bands was obtained up to 22 repeats in length, demonstrating that the αRI-DNA sequences occur in tandem arrays in the genomic DNA. A reassociation analysis using isolated insert DNA from one of the recombinant clones showed that the family of sequences is repeated 22,000 times in the human genome. Clones containing the αRI-DNA sequence were also isolated from a library of human genomic DNA in bacteriophage λ. Using these clones it was shown that, in at least some cases, the repetitive element is bounded by DNA less abundant than the αRI sequence.     

Localization of phosphatases in Trypanosoma rhodesiense

Phosphatase activity in Trypanosoma rhodesiense has been examined histochemically by light and electron microscopy and by enzymatic assay in homogenate fractions. Using a method with lead as capture ion, acid phosphatase was found in lysosome-like vesicles and in the flagellar pocket. No alkaline adenosine triphosphatase (ATPase) was detectable by this method. Direct assay of p-nitrophenylphosphatase activity in homogenate fractions showed that acid phosphatase activity was strongly membrane-bound, but that activity at pH 9 was minimal in both soluble and particulate fractions. "Endogenous" ATPase activity was localized specifically and reproducibly in the mitochondrial membranes and under the plasma membrane of he flagellum. This nonenzymic reaction product could not be eradicated by glycerol extraction or glucose depletion. Unlike the membrane staining, which was manifest only after lead treatment, heat-resistant electron-dense material was found in the matrix of lysosomal vesicles in trypanosomes fixed in glutaraldehyde only and not subjected to further treatment with heavy metal reagents. X-ray emission analysis showed the presence of calcium and phosphorus, indicating that the matrix might have a phosphate storage function.     

A physico-chemical study of heparin: Evidence for a calcium-induced co-operative conformational transition

The conformations of heparin in aqueous solution in the presence of sodium, potassium, magnesium and calcium cations were studied using circular dichroism, optical rotation, nuclear magnetic resonance and equilibrium dialysis. Potassium and magnesium cations, when added to sodium heparinate solutions, cause small chiroptical changes. Binding of calcium ions gives rise to large changes in both optical rotation and circular dichroism. This is indicative of a major change in chain conformation, which is also manifest in ¹³C and ¹H n.m.r.⁴Equilibrium dialysis suggests one mole of calcium bound per mole of tetrasaccharide, which n.m.r. indicates to be appropriately sulphated iduronateglucosamine-iduronate-glucosamine. The calcium is chelated by two iduronate carboxyl groups. Proton-proton coupling constants, determined by convolution difference spectroscopy and Carr-Purcell sequences, indicate that, over the temperature range 285 to 353 K, the iduronate ring is best described as ¹C₄(l) and the glucosamine residue as ⁴C₁(d) for both sodium and calcium forms.The conformational change induced by calcium is ascribed to rotation around the glycosidic linkages. The binding process is co-operative and the binding constant of 10³ to 10⁴m^?1 is biologically significant. The findings are consistent with intramolecular binding. Hence, this study represents the first report of a polysaccharide undergoing a cation-induced intramolecular disorder-order process. The authors postulate that a function of the post-polymerization epimerization of d-glucuronate to l-iduronate is the attainment of the precise geometry required for co-operative calcium binding with consequent modulation of the flexibility of the tetrasaccharide units.     

Variables affecting body burdens of lead,zinc and cadmium in a roadside population of the snailCepaea hortensis Müller

Summary Application of multiple regression analysis to the lead, zinc and cadmium contents of 985Cepaea hortensis collected from a suburban roadside site over two years showed that body weight, age and daylength were the major factors affecting soft tissue body burdens. Other highly significant influences were physiological changes at the juvenile-adult transition, vapour pressure deficit for the two days before sampling, and mean rainfall for the ten days before. Together these factors accounted for 69%, 79% and 87% of the total variance in lead, zinc and cadmium body burdens respectively. Unexplained variance was provisionally ascribed to individual differences in recent diet and digestive activity. The digestive gland contined the majority of the total body burden of each metal and played an important role in lead and zinc elimination. Whereas levels of lead inC. hortensis fluctuated rapidly, zinc was exchanged more slowly, perhaps by active regulation, and cadmium was effectively immobile, accumulating progressively with age. The implications of these results to the predators of land snails, and to terrestrial monitoring programmes, are discussed.     

Diel vertical migration in Mesocyclops edax: implications for predation rate estimates

The diel migration patterns of Mesocyclops edax and its preyin a small lake were followed in two studies separated by approximatelyone year. Gut contents of the predators were examined and selectivityindices calculated at each depth at 0100 h during 1980. Thethree principal zooplankton prey found in the guts of M. edaxwere Keratella, Kellicottia, and Bosmina. The predator and allthree major prey species exhibited unique and different dielvertical distribution and migration patterns. The complex natureof the spatio-temporal variation in prey density to which M.edax is exposed, demonstrates the dangers of using selectivityindices without knowledge of the distribution patterns of bothpredator and prey. An increase in vertebrate predation pressurefrom one year to the next is thought to be responsible for anincrease in the abundance of small zooplankton species, thedisappearance of two out of three of the large zooplankton species,and the onset of a pronounced nocturnal migration pattern inthe third large species. ¹Present address: Biology Department, Williams Hall #31, LehighUniversity, Bethlehem, PA 18015, USA     

The utilization of ketone bodies by the interscapular brown adipose tissue of the rat

The activities of 3-oxo acid-CoA transferase (EC 2.8.3.5, 13-15 micromol/min per g) and acetoacetyl-CoA thiolase (EC 2.3.1.9, 18-21 micromol/min per g) in interscapular brown adipose tissue of the rat are comparable to the activities reported for heart and kidney. The incorporation of D-3-hydroxy[3-14C]butyrate into lipid in vivo was about 30-fold higher in interscapular brown adipose tissue than in white adipose tissue of virgin rats. In lactating rats, the mammary gland was the major site of ketone body incorporation into lipid and incorporation of D-3-hydroxy-[3-14C]butyrate into lipid in brown adipose tissue was lower than in virgin rats. After an oral load of medium chain triacylglycerol, which inhibits lipogenesis in lactating mammary gland, the incorporation of ketone bodies into lipid was decreased in mammary gland but increased in brown adipose tissue. The rate of oxidation of D-3-hydroxy[3-14C]butyrate by brown adipose tissue slices in vitro was higher than the rate of incorporation into lipid.     

Radiosensitization of hypoxic mammalian cells in vitro by some 5-substituted-4-nitroimidazoles

The efficiencies of various 5-substituted-4-nitroimidazoles as radiation sensitizers have been determined in hypoxic Chinese hamster cells irradiated in vitro. Compared with published data on the sensitizing properties of substituted 2-nitro- and 5-nitroimidazoles, some of the 4-nitro derivatives show unusually high sensitizing efficiencies defined as the concentrations required to give an enhancement ratio of 1.6. The equilibrium one-electron reduction potentials of the compounds (E17) were measured by a pulse radiolysis technique and the results show that although sensitizing efficiencies are unexpectedly high, based on considerations of electron affinity, they still increase with increasing values of E17. Enhancement ratios were determined in two V79 cell lines for combinations of one of these compounds (a 4-nitroimidazole containing the group SO2.O.phenyl in the 5-position, NSC 38087) with various concentrations of misonidazole. The sensitization observed suggests that the two compounds may be operating by different mechanisms.