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Cerebrovascular disease is a common cause of morbidity in sickle cell anemia (HbSS): approximately 10% of patients have a clinical stroke before 20 years of age, and another 22% have silent infarction on magnetic resonance imaging. The phenotypic variation among patients with HbSS suggests a role for modifier genes and/or environmental influences. To assess the familial component of clinical stroke in HbSS, we estimated the prevalence of clinical stroke among all patients and among HbSS sibling pairs at 9 pediatric centers. The sample included 3425 patients with sickle cell disease who were younger than 21 years, including 2353 patients with HbSS. The stroke prevalence was 4.9% for all genotypes; 7.1% for patients with HbSS; 1.1% for patients with HbSβothalassemia; 0.6% for patients with Sβ+ thalassemia; and 0% for patients with HbSC. In 207 sibships, more than 1 child had HbSS. There were 42 sibships in which at least 1 sibling had a stroke, and in 10 of the 42, 2 siblings had a stroke. A permutation test indicated that the number of families in which 2 children had strokes was larger than the number expected if strokes were randomly distributed among children in sibships (P = .0012). There was no difference in stroke prevalence based on sex, nor was the mean age at stroke presentation significantly different between singletons and sibships with stroke. We conclude that there is a familial predisposition to stroke in HbSS. Attempts to identify genetic modifiers should be initiated with family-based studies.

The analysis of trade-restricting policies under uncertainty has typically argued in favour of the specific tariff over alternative commercial policies such as the ad valorem tariff and the quota, when the raising of revenue and maintenance of consumer's welfare are the policy objectives. This paper reconsiders this result. It is demonstrated that, when the shape of the welfare probability distribution is considered explicitly, the quota may be the dominant trade-restricting policy. In addition, the analysis of trade-restricting instruments is analysed for the case where the principle of safety-first is applied to the revenue-raising objective.

Chicken ovalbumin upstream promoter-transcription factor (COUP-TF) was identified as a low abundance protein in bovine uterus that co-purified with estrogen receptor (ER) in a ligand-independent manner and was separated from the ER by its lower retention on estrogen response element (ERE)-Sepharose. In gel mobility shift assays, COUP-TF bound as an apparent dimer to ERE and ERE half-sites. COUP-TF bound to an ERE half-site with high affinity, Kd = 1.24 nM. In contrast, ER did not bind a single ERE half-site. None of the class II nuclear receptors analyzed, i.e. retinoic acid receptor, retinoid X receptor, thyroid receptor, peroxisome proliferator-activated receptor, or vitamin D receptor, were constituents of the COUP-TF.DNA binding complex detected in gel mobility shift assays. Direct interaction of COUP-TF with ER was indicated by GST "pull-down" and co-immunoprecipitation assays. The nature of the ER ligand influenced COUP-TF-ERE half-site binding. When ER was liganded by the antiestrogen 4-hydroxytamoxifen (4-OHT), COUP-TF-half-site interaction decreased. Conversely, COUP-TF transcribed and translated in vitro enhanced the ERE binding of purified estradiol (E2)-liganded ER but not 4-OHT-liganded ER. Co-transfection of ER-expressing MCF-7 human breast cancer cells with an expression vector for COUP-TFI resulted in a dose-dependent inhibition of E2-induced expression of a luciferase reporter gene under the control of three tandem copies of EREc38. The ability of COUP-TF to bind specifically to EREs and half-sites, to interact with ER, and to inhibit E2-induced gene expression suggests COUP-TF regulates ER action by both direct DNA binding competition and through protein-protein interactions.

Restriction mapping of the globin genes from a homozygous delta beta thalassemia patient from Israel indicates that at least a 10-kilobase deletion is present extending 3' from within the large intervening sequence (IVS 2) of the delta globin gene and including the entire beta globin gene. Unique bands are seen when cellular DNA from this patient is digested with a variety of restriction endonucleases and hybridized with a probe specific for the delta IVS 2. Extensive analysis of the Israeli delta beta thalassemia DNA as well as material from an Italian delta beta thalassemia homozygote with enzymes which cleave more frequently in delta IVS 2 has localized the 5' end of the deletion to a 107-base pair region within delta IVS 2. This region contains a unique repetitive sequence (TG)4 which has been reported to be a specific recognition signal for recombination and may be involved in the formation of these mutant genes. Two homozygous delta(0) thalassemia DNA samples from Japan were also analyzed for gene rearrangements or other changes by restriction enzyme mapping. No changes from normal were seen using 14 different enzymes indicating the absence of large deletions in the region around the delta globin gene. More specifically, both the 5' and 3' splice junctions of the IVS 2 appear to be normal from hybridization of restriction fragments generated by HphI and AluI, respectively, with a delta IVS 2 specific probe. We have also shown that point mutations which could lead to termination codons are not present at codons 35, 37, 43, 61, and 121, since restriction enzymes which recognize these sites produce normal patterns. The delta(0) thalassemia phenotype in these two subjects is most likely due to a point mutation either at one of the other 24 potential termination codons not accessible to restriction analysis or to other single nucleotide changes which could either decrease delta globin gene transcription or lead to abnormal processing or transport of delta globin mRNA.

Benzimidazole anti-microtubule drugs, such as benomyl, induce paralysis and slow the growth of the nematode Caenorhabditis elegans. We have identified 28 mutations in C. elegans that confer resistance to benzimidazoles. All resistant mutations map to a single locus, ben-1. Virtually all these mutations are genetically dominant. Molecular cloning and DNA sequence analysis established that ben-1 encodes a beta-tubulin. Some resistant mutants are completely deleted for the ben-1 gene. Since the deletion strains appear to be fully resistant to the drugs, the ben-1 product appears to be the only benzimidazole-sensitive beta-tubulin in C. elegans. Furthermore, since animals lacking ben-1 are viable and coordinated, the ben-1 beta-tubulin appears to be nonessential for growth and movement. The ben-1 function is likely to be redundant in the nematode genome.

gamma delta beta-Thalassemia is a rare disorder of hemoglobin biosynthesis, characterized molecularly by partial or complete deletions of the beta-globin gene complex of 100 kilobases (kb) or greater. Common to all mutants described has been the deletion of the most-5' sequences of the beta-globin complex. We have used the techniques of pulsed-field gel electrophoresis and polymerase chain reaction to study a patient with a clinical gamma delta beta-thalassemia phenotype. This subject developed a de novo deletion on a maternally inherited beta-globin gene chromosome involving approximately 30 kb of sequences 5' to the epsilon gene; the deletion extends from -9.5 kb to -39 kb 5' of epsilon and includes three of the four DNase I hypersensitive sites (at -10.9 kb, -14.7 kb, and -18 kb 5' of epsilon). The remaining sequences of the beta-globin complex, including the DNase I hypersensitive sites at -6.1 kb and all structural genes in cis to the deletion are physically intact, but presumably nonfunctional, as evidenced by the presence of a beta S-globin gene that is not expressed as a sickle hemoglobin. Deletion of DNase I hypersensitive sites on a previously functional beta-globin gene complex confirms the significance of these sites in regulating globin gene expression.

In Saccharomyces cerevisiae, 3-amino-1,2,4-triazole (aminotriazole) competitively inhibits the activity of imidazoleglycerolphosphate dehydratase, the product of the HIS3 gene. Wild-type strains are able to grow in the presence of 10 mM aminotriazole because they induce the level of imidazoleglycerolphosphate dehydratase. However, strains containing gcn4 mutations are unable to grow in medium containing aminotriazole because they lack the GCN4 transcriptional activator protein necessary for the coordinate induction of HIS3 and other amino acid biosynthetic genes. Here, we isolated a new gene, designated ATR1, which when present in multiple copies per cell allowed gcn4 mutant strains to grow in the presence of aminotriazole. In wild-type strains, multiple copies of ATR1 permitted growth at extremely high concentrations of aminotriazole (80 mM), whereas a chromosomal deletion of ATR1 caused growth inhibition at very low concentrations (5 mM). When radioactive aminotriazole was added exogenously, cells with multiple copies of ATR1 accumulated less aminotriazole than wild-type cells, whereas cells with the atr1 deletion mutation retained more aminotriazole. Unlike the mammalian mdr or yeast PDR genes that confer resistance to many drugs, ATR1 appears to confer resistance only to aminotriazole. Genetic analysis, mRNA mapping, and DNA sequencing revealed that (i) the primary translation product of ATR1 contains 547 amino acids, (ii) ATR1 transcription is induced by aminotriazole, and (iii) the ATR1 promoter region contains a binding site for the GCN4 activator protein. The deduced amino acid sequence suggests that ATR1 protein is very hydrophobic with many membrane-spanning regions, has several potential glycosylation sites, and may contain an ATP-binding site. We suggest that ATR1 encodes a membrane-associated component of the machinery responsible for pumping aminotriazole (and possibly other toxic compounds) out of the cell.

The development of the sequential traits of seed yield (heads/area, seeds/head, and seed wt) of safflower, Carthamus tinctoriusL., and the breeding for high and low yield were studied. The original data for each of the three traits were standardized by two transformations and compared. Of the sequential traits, genotype had a large effect on seed wt and smaller effects on seeds/head and heads/ area. Location effects were generally highly significant for each trait; dates of planting were less important. The sequential traits show independence in a correlation analysis. Together the traits account for 97% of the variation in yield, with head numbers and seeds in the head accounting for most of the variation. For each trait separately, seed wt accounts for most of the variation in yield, followed by seeds/head and head numbers. Standardized partial regression coefficients indicate that for selection, one should give more weight to head numbers and seeds/head when all these traits are considered simultaneously and to head numbers when one trait is considered at a time. In general, head numbers or seeds/head or both traits could be responsible for high yielding lines. Seed wt was generally inflexible in different environments, but heads/ area and seeds/head were more flexible. Line A1186‐1 was among the five highest yielding lines in nearly all locations and for all planting dates, indicating the highest adaptation to the range of environments in the experiments.

The estrogen receptor (ER) is a transcription factor that binds to a specific DNA sequence found in the regulatory regions of estrogen-responsive genes, called the estrogen response element (ERE). Many genes that contain EREs have been identified, and most of these EREs contain one or more changes from the core consensus sequence, a 13-nucleotide segment with 10 nucleotides forming an inverted repeat. A number of genes have multiple copies of these imperfect EREs. In order to understand why natural EREs have developed in this manner, we have attempted to define the basic sequence requirements for ER binding. To this end, we measured the binding of homodimeric ER to a variety of nonconsensus EREs. We discovered that an ERE containing even a single change from the consensus may be unable to bind ER. However, an ERE with two changes from the consensus may be capable of binding avidly to ER in the context of certain flanking sequences. We found that changes in the sequences flanking a nonconsensus ERE can greatly alter ER-ERE affinity, either positively or negatively. Careful study of sequences flanking a series of EREs made it possible to develop rules that predict whether ER binds to a given natural ERE and also to predict the relative amounts of binding when comparing two EREs.

Achieving strain-level resolution is a major obstacle for source tracking and temporal studies of microbiomes. In this study, we describe a novel deep-sequencing approach that provides species- and strain-level resolution of the neonatal microbiome.