Human cellular retinol-binding protein gene organization and chromosomal location

The gene encoding the human cellular retinol-binding protein (CRBP) has been isolated from genomic libraries and its structure determined. Only one copy of the gene is present in the human genome. We have located the CRBP gene to segment 3p11-3qter on human chromosome 3 using hybridizations to mouse-human, rat-human and hamster-human cell hybrids. The gene harbors four exons encoding 24, 59, 33, and 16 amino acid residues respectively. The second intervening sequence alone occupies 19 kb of the 21 kb of the CRBP gene. The nucleotide sequence of the gene has been determined with the exception of the second intron. The positions of the introns agree with those in the rat CRBPII, the rat liver fatty-acid-binding protein and the mouse adipose P2 protein genes encoding molecules belonging to the same protein family as CRBP. In contrast to the other sequenced members of this family the promoter of the CRBP gene resembles those found in the 'housekeeping' genes in that it is (G + C)-rich, contains multiple copies of the CCGCCC sequence and lacks TATA box. A 9-bp homology containing the core sequence of the simian virus 40 enhancer repeat was found in the 5' upstream region. A genomic Southern blot probed with CRBP cDNA revealed hybridizing bands in restricted chicken and frog DNA.     

Structure and regulation of the sheep metallothionein-Ia gene

Screening of a sheep genomic lambda library with a sheep metallothionein-I cDNA clone resulted in the isolation of a 13,200-base-pair fragment containing a metallothionein gene which DNA sequence analysis identified as the gene encoding the cloned cDNA. The two introns occur at identical positions to those in other mammalian metallothioneins but are considerably larger. The first intron contains a DNA element that is present in a related but not identical form in many places in the sheep genome. Comparison of the promoter sequences of this gene (sMT-Ia) with the promoters of metallothionein genes from other species identified a number of conserved regions which may be important in the regulation of this gene by heavy metals, glucocorticoids and alpha-interferon. In sheep fibroblasts, the levels of sMT-Ia mRNA was found to be maximally elevated (95-fold) in the presence of zinc or cadmium and elevated 30-fold in the presence of copper. Dexamethasone had no effect upon mRNA levels. Thus this gene shows a pattern of regulation similar to the human MT-If gene, but distinct from the other human and mouse metallothionein genes so far reported.     

Mineral licks as a sodium source for Isle Royale moose

Summary Natural mineral licks and their use by moose (Alces alces) on Isle Royale National Park, Michigan, were studied during 1982–85. The distribution of known licks suggested that they occurred in association with glacial debris, primarily in the western portions of the island. Moose utilized mineral springs extensively during the spring-summer period, and at least 5 licks were used year-round. During summer, a pronounced diel pattern of moose visitation was apparent, with peak use occurring between 0400–0800 h. Although daytime lick use declined by late June, morning and evening use continued to be relatively high throughout the study period. Peak lick use coincided with leaf-emergence in spring. Moose continued to utilize mineral licks despite the availability of ponds containing aquatic plants. Sodium appeared to be the element attracting moose to licks where they ingest copious amounts of water. Observed sodium ingestion rates (0.35 g/min) at licks indicate that licks provide a more concentrated source of sodium compared to aquatic plants (0.023 g/min). Based on the data presented, we reject the conclusions of earlier workers that aquatic plants constitute the only significant source of sodium for Isle Royale moose.     

Molecular cloning of the c2 locus of Zea mays, the gene coding for chalcone synthase

Summary The c2 locus of Zea mays, identified as one of the genes affecting anthocyanin biosynthesis, was cloned using the transposable element En (Spm) as a gene tag. The Spm element present at the c2 locus in the autonomously mutating c2-m1 line was isolated using En1 element specific probes. Sequences flanking the element were identified as c2 locus specific and were used to clone the nonautonomous c2-m2 and wild-type alleles. The cloning and analysis of a cDNA complementary to the c2 locus provided evidence that this gene encodes the enzyme chalcone synthase.     

Changes in Insulin Binding to Developing Embryonic Chick Neural Retina Cells

Specific cell surface insulin binding to embryonic chick neural retina cells has been demonstrated in vivo. Kinetics of insulin binding as well as hormonal specificity were similar to those reported for other vertebrate cells and tissues, both neural and nonneural. When surface insulin binding to retinal cells was studied as a function of embryonic age, a developmental relationship was observed. Scatchard analysis revealed that the number of cell surface insulin receptors decreased approximately 75% between days 10 and 16 of embryonic development. Receptor affinities remained fairly constant for this period.     

Abrogation of cell surface expression of human class I transplantation antigens by an adenovirus protein in Xenopus laevis oocytes

Class I transplantation antigens form complexes with a virus protein encoded in the early region E3 of the adenovirus-2 genome. The interaction between this viral glycoprotein, E19, and nascent human class I antigens has been examined by microinjecting purified mRNA into Xenopus laevis oocytes. Both E19 and the two class I antigen subunits, the heavy chain and beta 2-microglobulin (beta 2M), were efficiently translated. The heavy chains did not become terminally glycosylated, as monitored by endoglycosidase H digestion, and were not expressed on the oocyte surface unless they were associated with beta 2M. The E19 protein did not become terminally glycosylated, and we failed to detect this viral protein on the surface of the oocytes. Co-translation of heavy chain and E19 mRNA demonstrated that the two proteins associate intracellularly. However, neither protein appeared to be transported to the trans-Golgi compartment. Similar observations were made in adenovirus-infected HeLa cells. Heavy chains bound to beta 2M became terminally glycosylated in oocytes in the presence of low concentrations of E19. At high concentrations of the viral protein, no carbohydrate modifications and no cell surface expression of class I antigens were apparent. Thus, beta 2M and E19 have opposite effects on the intracellular transport of the heavy chains. These data suggest that adenovirus-2 may impede the cell surface expression of class I antigens to escape immune surveillance.     

Recognition of Superior Photosynthetic Efficiency in the Field Using the CO(2)-Depletion Technique

The relationship between CO₂ exchange rate (CER) and growth of crops in the field was investigated in Connecticut Broadleaf tobacco (Nicotiana tabacum) using the CO₂-depletion technique. A particular objective was to determine if modest (i.e. <10%) varietal differences could be distinguished in mean CER. Statistical analysis of numerous CER values obtained over a wide range of irradiances during the course of the season indicated that differences of as little as 7% in the mean CER between varieties would be significant (n ~400). The usefulness of the CO₂-depletion technique in detecting modest differences in photosynthetic efficiency has thus been demonstrated. These results are discussed in relation to the prospects for introducing and detecting genetic traits which would diminish photorespiration and increase CER and growth.     

Isolation and characterization of an antithrombin III variant with reduced carbohydrate content and enhanced heparin binding

Two distinct forms of antithrombin III were isolated by chromatography of normal human plasma on heparin-Sepharose. The predominant antithrombin species present (AT-III alpha), which eluted from the affinity column in 1 M NaCl, was identified as the antithrombin III form which has been previously characterized. Ionic strength of the buffer was increased to elute a variant form of antithrombin III, designated as AT-III beta. The molecular weight of AT-III beta is less than that of AT-III alpha, but physicochemical studies do not indicate measureable differences in the polypeptide portion of the proteins. Carbohydrate determination revealed the sole detectable structural difference in the two antithrombins: levels of hexosamine, neutral sugars, and sialic acid in AT-III beta were all 25-30% less than in AT-III alpha. Kinetic studies of thrombin inactivation by both antithrombins, in the presence of nonsaturating amounts of heparin, indicated that AT-III beta inhibited thrombin more rapidly. AT-III beta is also distinguishable from AT-III alpha on the basis of heparin-binding affinity estimated from titration of protein fluorescence with heparin. Thus, antithrombin III exists as two molecular entities in human plasma which differ both structurally, in carbohydrate content, and functionally, in their heparin-binding behavior.