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Biomedical subjects

D W Schultz

Publications and source records attributed to D W Schultz.

At least 19 recordsLinked to original sources
PubMed1998

Age-related macular degeneration. Clinical features in a large family and linkage to chromosome 1q.

OBJECTIVES: To identify the chromosomal location of a disease-causing gene and to describe the clinical characteristics of a large family with age-related macular degeneration (ARMD). METHODS: An ARMD pedigree was identified, and the disease state of family members was documented by stereoscopic fundus photography and was classified using a modified version of the Wisconsin Age-Related Maculopathy Grading System. A genome-wide screen at approximately 6-centimorgan spacing using a DNA-pooling strategy combined with shared-segment analysis was used to identify likely chromosomal regions. The entire family was then screened at each likely locus, and 1 positive locus was refined by screening with markers at an average density of 0.5 centimorgan and subjected to parametric linkage analysis. RESULTS: In the 10 affected family members, ARMD was manifest by the presence of large, soft, confluent drusen accompanied by varying degrees of retinal pigment epithelial degeneration and/or geographic atrophy. Age-related macular degeneration segregated as an autosomal-dominant trait, with the disease locus mapping to chromosome 1q25-q31 between markers D1S466 and D1S413, with a multipoint lod score of 3.00. CONCLUSION: Age-related macular degeneration localized to chromosome 1q25-q31 (gene symbol, ARMD1) as a dominant trait in a large family with a predominantly dry phenotype. CLINICAL RELEVANCE: Identification of ARMD genes will facilitate early diagnosis and aid in understanding the molecular pathophysiological mechanisms of ARMD. This knowledge will contribute to the development of preventive and improved treatment strategies.

Aged↗
PubMed1997

Autosomal dominant cerulean cataract is associated with a chain termination mutation in the human beta-crystallin gene CRYBB2.

Congenital cataracts are a common major abnormality of the eye that frequently cause blindness in infants. At least a third of all cases are familial; autosomal dominant congenital cataract (ADCC) appears to be the most common familial form in the Western world. Cerulean cataracts have peripheral bluish and white opacifications in concentric layers with occasional central lesions arranged radially. Although the opacities may be observed during fetal development and childhood, usually visual acuity is only mildly reduced until adulthood, when lens extraction is generally necessary. We have been studying a family (ADCC-1) with cerulean blue ADCC, in which the affected daughter of a first cousin mating was presumed to be homozygous for the cataract gene. Recently, we mapped an ADCC gene in this family to a region of chromosome 22 containing three beta-crystallin genes. Here we report that a chain-termination mutation in CRYBB2 is associated with ADCC in this family.

Amino Acid Sequence↗
PubMed1996

On malleability in the genetic code.

To explain now-numerous cases of codon reassignment (departure from the "universal" code), we suggest a pathway in which the transformed codon is temporarily ambiguous. All the unusual tRNA activities required have been demonstrated. In addition, the repetitive use of certain reassignments, the phylogenetic distribution of reassignments, and the properties of present-day reassinged tRNAs are each consistent with evolution of the code via an ambiguous translational intermediate.

Codon↗
PubMed1994

Transfer RNA mutation and the malleability of the genetic code.

We propose that evolutionary reassignment of codons is facilitated by a translationally ambiguous intermediate. For example, recently discovered tRNA mutations that allow relatively efficient simultaneous cognate and near-cognate coding (sharing 2 contiguous nt) in vivo may speed reassignment of the near-cognate codon. As predicted by this notion, characterized codon reassignments are strikingly non-random, and half can be immediately explained by unusual tRNA activities already demonstrated. In addition, sequences of reassigned tRNAs contain sequences that promote ambiguity. tRNA structural change may provide a transitional pathway that allows rapid selection of a new specificity, rather than slow mutation toward a new codon-amino acid association.

Biological Evolution↗
PubMed1994

tRNA structure and ribosomal function. I. tRNA nucleotide 27-43 mutations enhance first position wobble.

Transfer RNA su7 G36 is a derivative of tRNA(Trp) with a 3'GUC anticodon complementary to the glutamine codon CAG. This tRNA requires a normally forbidden G-U wobble at the first codon position to suppress a UAG (amber) termination codon. Measurement of amber suppression by mutated su7 G36 tRNAs and correction for tRNA levels and aminoacylation allowed calculation of KUAG, a linearized index of in vivo ribosomal function. Following saturating mutagenesis of the anticodon arm of su7 G36, screening for UAG suppression using a lacZ reporter yielded tRNAs with up to 40-fold increased first position G-U wobble, judged from KUAG. The parental anticodon helix has minimized this type of miscoding, and virtually all changes in the top base-pair of the anticodon helix, nucleotides (nt) 27-43, increased the error. Thus, misincorporation of amino acids due to aberrant first position wobble is apparently prevented by normal tRNA structure, which is specifically altered by substitution at nt 27-43, the top base-pair of the anticodon helix. All 16 permutations of nt 27-43, the hotspot for increased wobble, were subsequently constructed and compared. Comparison of values for tRNA coding function, tRNA level, and aminoacylation for the 16 suggest that a tRNA conformational change, specifically involving both nt 27-43, differentially affects all these tRNA functions. This conformational alteration, which presumably occurs normally on the ribosome, appears more complex than simple breakage of the normal 27-43 base-pair. We suggest that the change is in the angle and/or flexibility of the tRNA L-shape. Among these 16 tRNAs, efficient wobble is strongly and inversely correlated with good aminoacylation and high tRNA levels; this quality may have been selected. Constraints on the sequences of natural tRNAs suggest that nt 27-43 have effects on function in many tRNAs.

Anticodon↗
PubMed1994

tRNA structure and ribosomal function. II. Interaction between anticodon helix and other tRNA mutations.

Using multiply mutated tRNA genes, we have studied unusual coding by tRNAs that have altered nucleotides (nt) 27-43, which normally form the top base-pair of the anticodon helix. In vivo, nt 27-43 mutations accelerate non-canonical C-A coding at the third (3') codon position 14-fold, similar to the 40-fold stimulation originally shown for first (5') codon position non-canonical G-U pairing. Thus the effects of nt 27-43 generalize to a second type of unusual coding. Nt 27-43 changes have a similar relative effect on tRNA level, aminoacylation, and ribosomal activity, despite concurrent changes of the 3' anticodon nucleotide which alter coding. However, under conditions of efficient aminoacylation, only a fraction of these (potential missense) anticodon changes can be recovered, suggesting toxicity. Available data support the idea that the effects of nt 27-43 are not particular to one codon. A previously isolated D-arm mutation (G24A) has a similar coding effect, enhancing both first position G-U wobble up to 130-fold, the third position C-A mispairing 40-fold. Anticodon helix mutations at 27-43 have little effect on 3' or 5' miscoding in the presence of the G24A D-arm mutation, and reciprocally, the D-arm's effects are much diminished in the presence of the anticodon helix mutations. Because these two tRNA loci alter both types of aberrant coding, and because they are highly interdependent, they may exploit a similar mechanism, dependent on a similar effect on tRNA conformation. We suggest a relatively non-specific decrease in the ribosomal rejection rates for tRNAs altered at anticodon helix nucleotides 27 and 43. Thus coding via non-canonical pairings at both 5' and 3' ends of the codon-anticodon helix has a measurable rate in vivo. However, we find that normal tRNA structure minimizes the efficiency of this aberrant translation. To put these same findings in another light, tRNAs bearing identical anticodons, if altered in structure elsewhere, may translate the genetic code differently.

Anticodon↗
PubMed1992

High dose oral methylprednisolone in patients with rheumatoid arthritis: pharmacokinetics and clinical response.

A commercially available 1.0 g intravenous (i.v.) dosage formulation of methylprednisolone, as the sodium hemisuccinate salt (Solu Medrol, Upjohn) was administered both parenterally and orally (pulse steroid therapy) on separate occasions, to eight elderly (mean 65 y) patients with active rheumatoid arthritis. The relative oral bioavailability of the sterol was 69.2%. Elimination of methylprednisolone was prolonged when given orally; the mean residence times were 7.23 h and 3.94 h for oral and i.v. administrations, respectively. Clinical response to pulse steroid therapy was no different with respect to route of administration. There were no significant differences in standard clinical and laboratory assessments of disease activity when the two therapies were compared. Oral administration of methylprednisolone in patients requiring high-dose pulse steroid therapy is convenient and avoids the discomfort and inconvenience associated with i.v. administration.

Administration, Oral↗
PubMed1990

A simple and sensitive in vivo luciferase assay for tRNA-mediated nonsense suppression.

We present a rapid assay for tRNA suppression in living Escherichia coli. An amber, ochre, or opal nonsense mutation in a cloned luxB gene from the bacterium Vibrio harveyi was suppressed. Because luciferase (Lux) activity depends completely on the appearance of the full-length luxB gene product, the amount of light produced was proportional to tRNA-mediated nonsense suppression in the cell. This luminometric assay was notably quicker, easier, and more sensitive than a traditional colorimetric assay employing beta-galactosidase. Assays required only one addition to a growing culture and were complete within 1 min. Light output was directly proportional to the amount of bacterial luciferase in a sample over a range of greater than or equal to 40,000-fold. Fewer than 100 cells were required for detection of Lux with ordinary instrumentation; assays were 80-fold more sensitive than simultaneous beta-galactosidase measurements. Assayed cells survived and could be recovered as colony formers. The beta-galactosidase colorimetric assay and the luciferase assay were similarly reproducible. Light from colonies expressing Lux was visible to the dark-adapted eye and useful for screening. A rapid assay that does not depend on the formation of permanent transformants can be based on electroporation followed by luminometry.

Base Sequence↗
PubMed1986

Conservation of Chi cutting activity in terrestrial and marine enteric bacteria.

Homologous recombination in Escherichia coli occurs at increased frequency near Chi sites, 5'G-C-T-G-G-T-G-G3'. Cutting of DNA close to the Chi sequence by the E. coli RecBC enzyme is essential to Chi's stimulation of recombination. We have detected Chi-dependent cutting activity in extracts of several genera of terrestrial enteric bacteria (family Enterobacteriaceae) and of two genera of marine enteric bacteria (family Vibrionaceae). More distantly related bacteria had no detectable Chi-dependent cutting activity. These results support the view that recognition of this specific nucleotide sequence as a signal activating recombination has been maintained during the evolution of certain groups of bacteria. We discuss the possibility that other sequences play a similar role in other groups of bacteria.

Autoradiography↗
PubMed1986

Activity of Chi recombinational hotspots in Salmonella typhimurium.

Chi sites have previously been shown to stimulate homologous recombination by the Escherichia coli RecBC pathway. To test the activity of Chi in another organism, bacteriophage lambda crosses were carried out in Salmonella typhimurium strains bearing the E. coli lambda receptor protein. Chi is active in these crosses in S. typhimurium, but is less active than in the same crosses carried out in E. coli. The lower Chi activity in S. typhimurium appears to be intrinsic to the S. typhimurium RecBC enzyme, since the Chi activity in E. coli-S. typhimurium hybrids depends on the species of origin of their RecBC enzyme. For these studies we constructed and F' factor and a pBR322-derived plasmid carrying the thyA+ recC+ recB+ argA+ region of the S. typhimurium chromosome.

Bacteriophage lambda↗
PubMed1985

Chi-dependent DNA strand cleavage by RecBC enzyme.

Chi sites enhance in their vicinity homologous recombination by the E. coli RecBC pathway. We report here that RecBC enzyme catalyzes Chi-dependent cleavage of one DNA strand, that containing the Chi sequence 5'G-C-T-G-G-T-G-G3'. Chi-specific cleavage is greatly reduced by single base pair changes within the Chi sequence and by mutations within the E. coli recC gene, coding for a RecBC enzyme subunit. Although cleavage occurs preferentially with double-stranded DNA, the product of the reaction is single-stranded DNA. These results demonstrate the direct interaction of RecBC enzyme with Chi sites that was inferred from the genetic properties of Chi and recBC, and they support models of recombination in which Chi acts before the initiation of strand exchange.

Base Sequence↗
PubMed1985

RecBC enzyme nicking at Chi sites during DNA unwinding: location and orientation-dependence of the cutting.

Homologous recombination by the E. coli RecBC pathway occurs at elevated frequency near Chi sites. We reported previously that Chi induces RecBC enzyme to cleave one DNA strand--that containing the Chi sequence 5'G-C-T-G-G-T-G-G3'. We report here that the Chi-dependent cleavage occurs four, five, or six nucleotides to the 3' side of the Chi octamer and produces nicks with 3'-OH and 5'-PO4 groups. Chi-dependent cleavage occurs if RecBC enzyme approaches the Chi sequence from the right, but not if it approaches only from the left, during unwinding of the duplex DNA substrate. A single RecBC enzyme molecule appears to cleave the DNA and to release part of it as a single-stranded fragment. These and previous results indicate that Chi-dependent cleavage is concomitant with DNA unwinding by RecBC enzyme and provide an enzymatic basis for the orientation-dependence of Chi recombinational hotspot activity. These observations demonstrate a key step of a proposed model of recombination in which RecBC enzyme produces a potentially invasive single-stranded DNA tail extending from Chi to its left. We discuss the relation between the action of Chi sites and that of special sites enhancing eukaryotic recombination.

Base Sequence↗
PubMed1984

Caries patterns in Head Start children in a fluoridated community.

Assessment of caries experience based on the person rather than on the tooth opens the possibility for qualitative descriptions of caries in a population, as well as for the study of specific factors associated with different caries experiences. The study of a Head Start population in adjacent fluoridated communities was divided into two parts. It was the purpose of part one of the study to determine the prevalence of specific caries patterns (presumably associated with different etiologies). Of the children, 39 percent were caries-free; 32 percent had carious lesions only in pit-and-fissure defects of molars; 6.5 percent had carious lesions in hypoplastic defects; 11 percent had facial-lingual lesions, compatible with "nursing caries"; and 11.5 percent had approximal lesions of molars; no child in the study had rampant caries. The second part compared specific lifestyle variables with specific caries patterns. Statistically significant differences or trends were found between caries-free children and those with smooth-surface lesions for mother's educational level, time spent with grandparents, mother's perceived primary reason for cavities, and mother's tendency to permit the child to eat sweets without restriction. No significant differences or trends were found for lifestyle variables between caries-free children and those having lesions associated only with tooth defects.

Child, Preschool↗
PubMed1983

Nucleotide sequence of the lysozyme gene of bacteriophage T4. Analysis of mutations involving repeated sequences.

The nucleotide sequence of the lysozyme (e) gene of bacteriophage T4 and approximately 130 additional nucleotides on each side has been determined. The 5'-end of the gene for internal protein III appears to be located about 70 base-pairs from the 3'-end of the lysozyme gene. Nucleotide sequence analysis of mutant e genes confirmed that three identified hotspots of frameshift mutations are runs of five A nucleotides in the wild-type gene. The endpoints of two deletions are direct repeats of eight base-pairs in the wild-type gene. Two frameshift mutations with high reversion frequencies are duplications of five or seven base-pairs. The cloning and nucleotide sequence determination of the lysozyme gene will facilitate further study of the molecular biology of T4 lysozyme.

Base Sequence↗