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W Connor

Publications and source records attributed to W Connor.

17 recordsLinked to original sources
PubMed1996

Evaluation of selected two-dimensional segmentation techniques for computed tomography quantitation of lymph nodes.

RATIONALE AND OBJECTIVES: As contrast agents that selectively target normal lymph nodes are undergoing development and evaluation, it has become important to accurately and reproducibly determine nodal boundaries to study the agents and determine such values as lymph node area or mean nodal contrast concentration. This study was performed to evaluate the accuracy of different two-dimensional computer segmentation methods, tested on acrylic phantoms constructed to imitate the appearance of lymph nodes surrounded by fat. METHODS: Five segmentation techniques (manual tracing, semiautomatic local criteria threshold selection, Sobel/watershed technique, interactive deformable contour algorithm and thresholding) were evaluated using phantoms. Subsequently, the first three methods were applied to the images of enhanced lymph nodes in rabbits. RESULTS: Minimum errors in phantom area measurement (< 5%) and interoperator variation (< 5%) were seen with the Sobel/watershed technique and the interactive deformable contour algorithm. These two techniques were significantly better than thresholding and semiautomated thresholding based on local properties. CONCLUSION: Methods based on Sobel edge detection offer more objective tools than thresholding methods for segmenting objects similar to lymph nodes in computed tomography images. Both methods, Sobel/watershed and interactive deformable contour algorithm, are fast and have simple user interfaces.

Algorithms↗
PubMed1995

Expression of an avian protamine in transgenic mice disrupts chromatin structure in spermatozoa.

The objective of this study was to determine the consequences of disrupting spermatozoal chromatin condensation on spermatozoal development and function. The avian protamine, galline, was targeted to spermatids of transgenic mice using the mouse protamine 1 gene promoter. Three transgenic mouse lines were established that expressed galline mRNA at 65%, 120%, and 185% of the level found in rooster testis. Galline mRNA accumulated in round spermatids to levels similar to that of mouse protamine and, as with the mammalian counterpart, translation was delayed until the elongating spermatid stage. Protein gels revealed that galline accumulated in mature spermatozoa whereas mouse protamines were reduced, suggesting that galline competes with protamines for binding to spermatozoal DNA. Acridine orange binding analysis indicated that DNA of the transgenic spermatozoa was not as tightly packed as that of controls. This was corroborated by electron microscopy, which revealed disruption of the normal dense chromatin structure of spermatozoal heads. Despite these perturbations of chromatin condensation, the transgenic spermatozoa were functionally normal, as the majority of transgenic mice had normal fertility. However, in mice that expressed excessive galline, there was a gradual destruction of seminiferous tubules leading to infertility. Our findings suggest that very precise packaging of DNA in germ cells may not be essential for subsequent unpackaging in the pronucleus of fertilized eggs and for subsequent normal development of the embryo.

Animals↗
PubMed1993

Very low high-density lipoproteins without coronary atherosclerosis.

Epidemiological studies have established that concentrations of plasma high-density lipoproteins (HDL) are inversely associated with premature atherosclerosis, but the physiological basis of this relationship remains unknown. We investigated 5 probands with very low plasma HDL. None had clinical or biochemical findings typical of the known genetic disorders with low HDL nor had evidence of premature coronary atherosclerosis by sensitive diagnostic methods. All 5 probands and the son of 1 of them had rapid catabolism of the HDL apolipoproteins A-I and A-II. These results indicate that not all people with low HDL are necessarily at risk of premature coronary heart disease and that further investigation is required before decisions can be made about their management.

Adult↗
PubMed1993

Characterization of a marsupial sperm protamine gene and its transcripts from the North American opossum (Didelphis marsupialis).

A synthetic oligonucleotide primer, designed from marsupial protamine protein-sequence data [Balhorn, R., Corzett, M., Matrimas, J. A., Cummins, J. & Faden, B. (1989) Analysis of protamines isolated from two marsupials, the ring-tailed wallaby and gray short-tailed opossum, J. Cell. Biol. 107] was used to amplify, via the polymerase chain reaction, protamine sequences from a North American opossum (Didelphis marsupialis) cDNA. Using the amplified sequences as probes, several protamine cDNA clones were isolated. The protein sequence, predicted from the cDNA sequences, consisted of 57 amino acids, contained a large number of arginine residues and exhibited the sequence ARYR at its amino terminus, which is conserved in avian and most eutherian mammal protamines. Like the true protamines of trout and chicken, the opossum protamine lacked cysteine residues, distinguishing it from placental mammalian protamine 1 (P1 or stable) protamines. Examination of the protamine gene, isolated by polymerase-chain-reaction amplification of genomic DNA, revealed the presence of an intron dividing the protamine-coding region, a common characteristic of all mammalian P1 genes. In addition, extensive sequence identity in the 5' and 3' flanking regions between mouse and opossum sequences classify the marsupial protamine as being closely related to placental mammal P1. Protamine transcripts, in both birds and mammals, are present in two size classes, differing by the length of their poly(A) tails (either short or long). Examination of opossum protamine transcripts by Northern hybridization revealed four distinct mRNA species in the total RNA fraction, two of which were enriched in the poly(A)-rich fraction. Northern-blot analysis, using an intron-specific probe, revealed the presence of intron sequences in two of the four protamine transcripts. If expressed, the corresponding protein from intron-containing transcripts would differ from spliced transcripts by length (49 versus 57 amino acids) and would contain a cysteine residue.

Amino Acid Sequence↗
PubMed1988

Bovine protamine genes contain a single intron. The structures of the two alleles.

In a recent paper, we described the isolation of the first bovine protamine cDNA (BPK59), demonstrating that the gene was autosomal and single copy (Krawetz, S. A., Connor, W., and Dixon, G. H. (1987) DNA 6, 45-57). We have since utilized the BPK59 clone as a probe, to isolate both alleles of the protamine gene from a bovine genomic library constructed in Charon 28. The sequenced alleles are highly homologous and code for identical mRNAs. Unlike the trout protamine genes, the bovine gene is not contiguous as it possesses a single intron. This intron divides the highly variable mammalian carboxyl domain from the conserved protamine basic core. Three tandemly repeated CG-like ("enhancer") sequences upstream from the CAAT box have been identified, separated by a conserved spacer region. Their possible role in the transcription of this P1 gene is discussed. The bovine genome immediately surrounding (approximately equal to 20 kilobase pairs) the protamine gene has been mapped and reveals that the sequence flanking the 5' segment of the gene is unique, while the sequence flanking the 3' segment is repetitive.

Alleles↗
PubMed1987

Cloning of bovine P1 protamine cDNA and the evolution of vertebrate P1 protamines.

A bovine P1 protamine cDNA from a bull testis cDNA library was isolated utilizing a series of oligonucleotide probes. Sequence analysis showed that the cloned cDNA insert extended 317 bp to the poly(A) tail. The 51-residue 6750-dalton protamine primary translated protein is encoded within a 156-bp segment. The protamine sequence predicted from the cDNA sequence differs from that previously reported for the amino acid sequence of bovine protamine P1 by the insertion of the tripeptide Cys-Arg-Arg from residues 39-41 in the carboxy-terminal region of the mature protein. Consistent with previous hybridization analysis, nucleotide sequence comparisons showed that trout protamine cDNA was more closely related to that of bovine than to that of mouse. However, bovine P1 protamine cDNA shared greater sequence homology with mouse P1. A common nucleotide sequence of 30 bp is conserved among all three of these species. Primer extension analysis revealed that, as with trout protamine mRNAs, the majority of the untranslated portion of the mRNA lies 3' to the coding segment. Comparisons of their mRNA secondary structures by computer modeling indicate that the mRNAs fold back onto themselves, producing similar, extensively hydrogen-bonded, convoluted forms. These models support the view that translational regulation of protamine mRNA may be partially dependent on secondary structure. Southern analysis suggests that the bovine protamine P1 gene is not sex-linked and is present as one (or relatively few) copy within the bovine genome.

Amino Acid Sequence↗
PubMed1986

Oligonucleotides as probes for mammalian protamine mRNAs.

Protamine-like sequences have been identified in poly A(+)mRNAs from mammalian testes by the use of a common, complementary oligonucleotide (GCAGCANCKPTANCKNGCCAT; predicted from the common N-terminal amino acid sequence, MARYRCC, seen in several mammalian P1 protamines [D.J. McKay, B.S. Renaux and G.H. Dixon, Bioscience Reports 5:383-391 (1985)]). This oligonucleotide was utilized to prepare species-specific, primer-extended transcripts for use as Northern blotting probes. Analysis of the mRNA primer-extended transcripts revealed a discrete and similar set of products common to both bull and rat testis mRNAs which were distinct from those obtained from human testis mRNA. Northern analysis of total poly A(+) mRNAs from the corresponding mammalian testis was consistent with these results and suggests that bull and rat protamine mRNAs are more closely related to each other than to human protamine mRNA.

Animals↗
PubMed1986

A vector-primer-cloner-sequencer plasmid for the construction of cDNA libraries: evidence for a rat glyceraldehyde-3-phosphate dehydrogenase-like mRNA and a ferritin mRNA within testis.

We have developed a vector-primer-cloner-sequencer (VPCS), based on the pUC plasmids, which is easy to prepare. Stable cDNA libraries have been generated from human, rat, and bull testis. The inserts are of various size classes, including high-molecular-weight reverse transcripts which can be easily sequenced. The utility of this VPCS has been demonstrated by isolating a rat ferritin and glyceraldehyde-3-phosphate dehydrogenase-like clone.

Animals↗
PubMed1985

Histone H4 and H2B genes in rainbow trout (Salmo gairdnerii).

The complete nucleotide sequence of the 3.0-kb BamH I-Sst I restriction fragment contained within the rainbow trout genomic clone lambda TH2 has been determined. This region contains the rainbow trout H4 and H2B histone genes and 5' and 3' flanking and spacer sequences, and represents the 5' half of the histone-gene cluster; the remaining half has been characterized previously. The genes are uninterrupted, and are transcribed from the same strand. The protein sequence of H4, as determined from the nucleic acid sequence, is the same as that derived for other vertebrate H4 proteins, although comparison of nucleotide sequences shows a great deal of sequence divergence, especially in the third base position. The amino acid sequence of H2B, though largely homologous to those of other vertebrate H2B proteins, displays some characteristic differences in primary structure. Consensus sequences noted in many other eukaryotic genes, as well as histone-specific consensus sequences, have been identified. An unusual feature of the spacer region between the H4 and H2B genes is the presence of a duplicated sequence 87 bp in length. The 5' and 3' ends of each repeat are complementary, and each repeat contains smaller repeated sequences internally, as well as a possible cruciform structure.

Amino Acid Sequence↗
PubMed1984

Organization of the histone genes in the rainbow trout (Salmo gairdnerii).

Twelve clones containing histone genes were isolated from a genomic trout library constructed in the vector Charon 4A. Each of the clones was found to contain a conserved 10.2-kb Eco RI fragment that contained one copy of each of the histones in the order H4-H2B-H1-H2A-H3, all of which are transcribed from the same strand. Genomic Southern blots indicate that these clusters are representative of the vast majority of the histone genes in the trout. Tandemly linked clusters were not found. Approximately 145 copies of this cluster are present in a trout sperm cell. Sequence analysis has shown the genes to be without introns and to show strong selection for codons ending in C or G. Consensus signals similar to those found in other histone genes are present in the flanking regions.

Animals↗
PubMed1984

Organization and nucleotide sequence of rainbow trout histone H2A and H3 genes.

A 2.56-kbp fragment containing genes coding for histones H2A and H3 that forms a portion of the 10.2-kbp cluster containing all five histone genes isolated from a lambda-Charon 4A library of rainbow trout genomic DNA has been characterized in detail and its complete nucleotide sequence determined. The genes are arranged in tandem, being encoded on the same DNA strand. They are separated by 380 bp of intergenic spacer DNA that contains an alternating purine-pyrimidine stretch of 20 bp and a 46-bp stretch that has the potential of forming a triple cruciform structure. The histone genes contain no introns, have the RNA polymerase II promoter-associated signals known as CAAT and TATA boxes in their 5' flanking regions and contain a conserved inverted repeat sequence, similar to that found in histone genes of other species, capable of forming a hairpin structure at the 3' end of the transcription unit.

Amino Acid Sequence↗
PubMed1982

Nucleotide sequence of a protamine component CII gene of Salmo gairdnerii.

We have isolated, using nick-translated cloned protamine cDNA's as probes, several genomic clones containing protamine gene sequences from a Charon 4A library of Eco R1 digested rainbow trout (Salmo gairdnerii) DNA. One clone was chosen for detailed study and the 2.5 kbp Bam HI-Eco R1 restriction fragment containing the gene was subcloned in the plasmid pBR322. A 920 bp Bg1 II - Bam HI restriction fragment contains a sequence coding for protamine component CII as well as regions 5' and 3' to the mRNA coding portion. Present in the region 5' to the mRNA coding sequence are the promoter associated signals "TATA" box and "CAAT" box. The 5' untranslated region of the mRNA whose length and sequence were not established from the cDNA clones (1) was determined by nuclease mapping and starts within a sequence similar to the "capping signal" found in other genes. The protamine gene for CII contains no introns, a situation common to most histone genes, but, unlike the histone genes does not occur close to other protamine genes in a "cluster".

Animals↗
PubMed1981

Molecular analysis of the protamine multi-gene family in rainbow trout testis.

We have synthesized a family of double-stranded cDNAs (ds cDNAs) using as a template the family of highly purified protamine mRNAs from rainbow trout testis. Individual pure protamine cDNA components were isolated by cloning this family of protamine ds cDNAs in a plasmid vector (pMB9). Clones containing protamine sequences were characterized by restriction mapping and by a positive hybrid-selected translation assay, which allowed us to correlate particular cDNAs with particular protein components. To allow more detailed comparisons, complete nucleotide sequences were determined for selected protamine clones. We have detected at least 5 distinctly different coding sequences, which nevertheless show at least 82% homology, and which have probably arisen by repeated gene duplication. These very highly conserved coding sequences do however contain a distinctly variable region near the 5'-end of the mRNA (N-terminus of the protein), corresponding to the major sites of serine phosphorylation. Since the amino acid sequences predicted by our DNA sequences were slightly different from those previously published (1), we have independently determined the amino acid sequences of protamine components CI, CII, CIII from our own source of trout testis. These new peptide sequences are completely consistent with those predicted by our nucleotide sequences. The 3'-untranslated regions of the protamine mRNAs are, surprisingly almost as highly conserved as the coding regions. Both coding and 3'-noncoding portions appear to be under a similar degree of selective pressure and evolutionary constraint to remain constant.

Animals↗
PubMed1979

A subset of trout testis nucleosomes enriched in transcribed DNA sequences contains high mobility group proteins as major structural components.

Mononucleosomes greatly enriched in non-histone proteins were prepared by limited digestion of testis nuclei with micrococcal nuclease. Five to fifteen per cent of the chromatin was solubilized and could be separated by adjustment to 0.1 M NaCl, into a soluble fraction MN1, consisting of mononucleosomes containing the four inner histones and the small basic non-histone, H6, associated with a 140-base-pair DNA fragment. H1 was notably absent in MN1. The fraction insoluble in 0.1 M NaCl (MN2) comprised a mixture of mono-, di-, tri-, and oligosomes. MN2 monosome fraction contained the four inner histones plus H1 and lacked H6 and the length of its DNA was 170 base-pairs. Previous work had shown that limited micrococcal nuclease digestion of trout testis nuclei released a great proportion of the non-histone protein, high mobility group protein T (HMG-T). It seems likely that HMG-T is the major non-histone protein located in the linker regions of a subset of nucleosomes containing the non-histone protein H6 as a major structural component. Moreover, the presence of HMG-T renders this subset of nucleosomes very sensitive to micrococcal nuclease. Hybridization experiments were performed to demonstrate that the DNA from MN1 monosomes corresponds to a subset of the trout testis genome. This DNA subset is greatly enriched in sequences that are present in cytoplasmic RNA. Chromatin subunits enriched in their content of H6 and HMG-T could also be obtained by limited digestion of trout testis chromatin with DNase II followed by precipitation with MgCl2.

Animals↗
PubMed1978

Cholesterol turnover in hereditary crystalline corneal dystrophy of Schnyder.

A patient with hereditary crystalline corneal dystrophy of Schnyder required penetrating keratoplasty because of poor visual acuity. Blood cholesterol was labeled with 14C-cholesterol and 11 days later a penetrating keratoplasty was done. At the time of surgery, corneal levels of cholesterol were much higher than serum levels, showing that cornea is an active site for the uptake and storage of cholesterol in this disorder.

Adult↗
PubMedDate not supplied

An H1 histone gene from rainbow trout (Salmo gairdnerii).

A 1.7-kbp DNA region from the 10.2-kb cluster containing the five rainbow trout histone genes has been subcloned in pBR322 and completely sequenced. It contains a trout histone H1 gene together with its 5' and 3' flanking sequences. This H1 gene codes for a H1 variant different from the major trout testis H1 previously sequenced by Macleod et al. (1977). Northern blots of total RNA from trout testis, kidney, and liver indicate that this H1 gene is expressed in all three tissues but that the level of H1 mRNA is much higher in testis than in other tissues. The lack of heterogeneity in the sizes and 5' initiation sites of trout H1 mRNAs is surprising in view of the substantial heterogeneity of H1 variant proteins observed previously. The coding sequence of the H1 gene shows strong evidence of repeated partial duplications of a hexapeptide motif of the form Ala.Ala.Ala.Lys.Lys.Pro and of a pentapeptide phosphorylation-site sequence, Lys.Ser.Pro.Lys.Lys, during its evolution. Comparisons are drawn between this gene and the coding sequences of other vertebrate H1 genes from chicken and Xenopus, and a strong homology is seen in the region of amino acids 22-101, which form the hydrophobic "head" of the H1 molecule. The 5' and 3' regulatory signals in the trout H1 are also compared with those of H1 genes from other sequences.

Amino Acid Sequence↗