Search PubMed⌕ Search

Biomedical subjects

J A Ashley

Publications and source records attributed to J A Ashley.

At least 19 recordsLinked to original sources
PubMed1997

TCOF1 gene encodes a putative nucleolar phosphoprotein that exhibits mutations in Treacher Collins Syndrome throughout its coding region.

Treacher Collins Syndrome (TCS) is the most common of the human mandibulofacial dysostosis disorders. Recently, a partial TCOF1 cDNA was identified and shown to contain mutations in TCS families. Here we present the entire exon/intron genomic structure and the complete coding sequence of TCOF1. TCOF1 encodes a low complexity protein of 1,411 amino acids, whose predicted protein structure reveals repeated motifs that mirror the organization of its exons. These motifs are shared with nucleolar trafficking proteins in other species and are predicted to be highly phosphorylated by casein kinase. Consistent with this, the full-length TCOF1 protein sequence also contains putative nuclear and nucleolar localization signals. Throughout the open reading frame, we detected an additional eight mutations in TCS families and several polymorphisms. We postulate that TCS results from defects in a nucleolar trafficking protein that is critically required during human craniofacial development.

Base Sequence↗
PubMed1997

The structure of the human multiple exostoses 2 gene and characterization of homologs in mouse and Caenorhabditis elegans.

Hereditary multiple exostoses (EXT) is an autosomal dominant disorder characterized by multiple cartilage-capped outgrowths from the epiphyses of long bones. In some cases, these osteochondromas progress to malignant chondrosarcomas. Alterations in at least three genes (EXT1, EXT2, and EXT3) can cause this disorder. Two of these have been isolated (EXT1 and EXT2) and encode related members of a putative tumor suppressor family. We report here the genomic structure of the human EXT2 gene consisting of 14 exons (plus 2 alternative exons) covering an estimated 108 kb of chromosome 11p11-13. We have derived the DNA sequences at all exon/intron boundaries throughout this gene-information that is important for the detailed study of mutations in EXT2. We have also characterized the mouse EXT2 cDNA and have mapped the mouse locus to chromosome 2 between D2Mit15 and Pax6. This mouse homolog should enable transgenic knockout experiments to be initiated to further elucidate gene function. Interestingly, sequence comparisons reveal that the human and mouse EXT genes have at least two homologs in the invertebrate Caenorhabditis elegans, indicating that they do not function exclusively as regulators of bone growth. This observation opens the way for a functional analysis of these genes in nematodes and other lower organisms.

Amino Acid Sequence↗
PubMed1996

Identification of the homologous beige and Chediak-Higashi syndrome genes.

Vesicular transport to and from the lysosome and late endosome is defective in patients with Chediak-Higashi syndrome (CHS) and in mutant beige (bg) mice. CHS and bg cells have giant, perinuclear vesicles with characteristics of late endosomes and lysosomes that arise from dysregulated homotypic fusion. CHS and bg lysosomes also exhibit compartmental missorting of proteins, such as elastase, glucuronidase and cathepsin G. Lyst, a candidate gene for bg, was identified by direct complementary DNA selection from a yeast artificial chromosome (YAC) clone containing a 650-kilobase segment of the bg-critical region on mouse chromosome 13. Lyst is disrupted by a 5-kilobase deletion in bg mice, and Lyst messenger RNA is markedly reduced in bg homozygotes. The homologous human gene, LYST, is highly conserved with mouse Lyst, and contains a frame-shift mutation at nucleotides 117-118 of the coding domain in a CHS patient. Thus bg mice and human CHS patients have homologous disorders associated with Lyst mutations. Lyst encodes a protein with a carboxy-terminal prenylation motif and multiple potential phosphorylation sites. Lyst protein is predicted to form extended helical domains, and has a region of sequence similar to stathmin, a coiled-coil phosphoprotein thought to act as a relay integrating cellular signal response coupling.

Amino Acid Sequence↗
PubMed1995

Reactive immunization.

For almost 200 years inert antigens have been used for initiating the process of immunization. A procedure is now described in which the antigen used is so highly reactive that a chemical reaction occurs in the antibody combining site during immunization. An organophosphorus diester hapten was used to illustrate this concept coined "reactive immunization." The organophosphonate recruited chemical potential from the immune response that resembled the way these compounds recruit the catalytic power of the serine hydrolases. During this recruitment, a large proportion of the isolated antibodies catalyzed the formation and cleavage of phosphonylated intermediates and subsequent ester hydrolysis. Reactive immunization can augment traditional immunization and enhance the scope of catalytic antibody chemistry. Among the compounds anticipated to be effective are those that contain appropriate reactive functionalities or those that are latently reactive, as in the mechanism-based inhibitors of enzymes.

Animals↗
PubMed1995

Suppression of psychoactive effects of cocaine by active immunization.

Cocaine is a powerfully addictive substance and new strategies are needed to treat its abuse. Generating an active immunization to cocaine offers a means of blocking the actions of the drug by preventing it from entering the central nervous system, and should have fewer side effects than treatments based on manipulation of central neurotransmitter function. The design and preparation of a cocaine immunogen requires special regard for the stability of cocaine both free and as a haptenic determinant. Immunochemistry and a well defined behavioural model were brought together to address the problem of inactivation of the psychostimulant actions of cocaine. We report here that active immunization with a new, stable cocaine conjugate suppressed locomotor activity and stereotyped behaviour in rats induced by cocaine but not by amphetamine. Moreover, following acute injection of cocaine, levels of cocaine in the striatum and cerebellum of the immunized animals were lower than those of control animals. These results suggest that immunopharmacotherapy may be a promising means by which to explore new treatments for cocaine abuse.

Animals↗
PubMed1995

Human chromosome-specific cDNA libraries: new tools for gene identification and genome annotation.

To date, only a small percentage of human genes have been cloned and mapped. To facilitate more rapid gene mapping and disease gene isolation, chromosome 5-specific cDNA libraries have been constructed from five sources. DNA sequencing and regional mapping of 205 unique cDNAs indicates that 25 are from known chromosome 5 genes and 138 are from new chromosome 5 genes (a frequency of 79.5%). Sequence complexity estimates indicate that each library contains -20% of the approximately 5000 genes that are believed to reside on chromosome 5. This study more than doubles the number of genes mapped to chromosome 5 and describes an important new tool for disease gene isolation.

Base Sequence↗
PubMed1994

Antibody catalyzed cationic cyclization.

Two major goals for the design of new catalysts are the facilitation of chemical transformations and control of product outcome. An antibody has been induced that efficiently catalyzes a cationic cyclization in which an acyclic olefinic sulfonate ester substrate is converted almost exclusively (98 percent) to a cyclic alcohol. The key to the catalysis of the reaction and the restriction of the product complexity is the use of antibody binding energy to rigidly enforce a concerted mechanism in accord with the design of the hapten. Thus, the ability to direct binding energy allows the experimenter to dictate a reaction mechanism which is an otherwise difficult task in chemistry. New catalysts for cationic cyclization may be of general use in the formation of carbon-carbon and carbon-heteroatom bonds leading to multi-ring molecules including steroids and heterocyclic compounds.

Animals↗
PubMed1994

Competition and position-dependent targeting in the development of the Drosophila R7 visual projections.

The R7 photoreceptor neuron projections form a retinotopic map in the medulla of the Drosophila optic lobe. The more inner photoreceptors mutation, an allele of gap1, results in the differentiation of excess R7s in the eye, whose axons invade the brain and establish functional connections. We have used this hyperinnervation phenotype to explore the roles of photoreceptor-target regulation, competitive interactions, and chemoaffinity in map formation. We show that the extra axons are supported in a wild-type brain, with all R7s from a single ommatidium sharing a single termination site, and thus there is no evidence that the target regulates the size of the presynaptic population. In mosaic eyes, in which ommatidia containing extra R7s are surrounded by ommatidia lacking all R7 cells, R7 axons still target to appropriate retinotopic locations in a largely empty R7 terminal field. Axons at the edges of the projection, however, send collaterals into vacant areas of the field, suggesting they are normally restrained to share single termination sites by competitive interactions. In contrast, no sprouts are seen when the vacant sites are juxtaposed with singly innervated sites. In the third instar, R7 and R8 axons transiently display halos of filopodia that overlap adjacent terminals and provide a means to assess occupancy at adjacent sites. Finally, in sine oculis larvae in which only a small number of ommatidia develop, the R7/R8 axons target to predicted dorsoventral portions of the medulla despite the absence of their neighbors, suggesting that position in the eye field determines their connectivity in the brain. We suggest that the mechanisms used to set up this insect map are formally similar to strategies used by vertebrates. The availability of a genetic model for these events should facilitate studies aimed at understanding the molecular bases of retinotopic map development.

Animals↗
PubMed1991

An unexpectedly efficient catalytic antibody operating by ping-pong and induced fit mechanisms.

A transition state analogue was used to produce a mouse antibody that catalyzes transesterification in water. The antibody behaves as a highly efficient catalyst with a covalent intermediate and the characteristic of induced fit. While some features of the catalytic pathway were programmed when the hapten was designed and reflect favorable substrate-antibody interactions, other features are a manifestation of the chemical potential of antibody diversity. The fact that antibodies recapitulate mechanisms and pathways previously thought to be a characteristic of highly evolved enzymes suggests that once an appropriate binding cavity is achieved, reaction pathways commensurate with the intrinsic chemical potential of proteins ensue.

Acylation↗