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F H Ruddle

Publications and source records attributed to F H Ruddle.

At least 145 records · Page 8Linked to original sources
PubMed1986

The homeo domain of a murine protein binds 5' to its own homeo box.

Nuclear protein extracts from day 12.5 mouse embryos were used to study protein binding to DNA sequences 5' of the Hox 1.5 homeo box. Embryos of this developmental stage are known to express this gene. DNA binding protein blotting and retardation gel techniques show that murine embryonic nuclear proteins specifically bind a 753-base pair (bp) DNA fragment from the region upstream of the Hox 1.5 homeo box. A fusion protein containing the Hox 1.5 homeo domain constructed in lambda gt11 also binds the same 753-bp DNA fragment. Specific binding of the fusion protein to the upstream DNA fragment shows that the homeo box contains the sequences required for specific protein-DNA interactions, and the 753-bp fragment contains a homeo domain binding site. These results support the hypothesis that murine homeo boxes are DNA binding domains of proteins involved in the regulation of embryonic development.

Amino Acid Sequence↗
PubMed1986

Cell proliferation and expression of the transferrin receptor gene: promoter sequence homologies and protein interactions.

A 365-bp fragment from the 5' region of the human transferrin receptor gene has been subcloned and sequenced. This fragment contains 115 bp of flanking sequence, the first exon, and a portion of the first intron. It contains a TATA box, several GC-rich regions, and is able to efficiently promote expression of the bacterial CAT gene in mouse 3T3 cells. Sequence comparisons demonstrate that this DNA segment has homology to the promoter regions of the human dihydrofolate reductase gene and the mouse interleukin 3 gene, as well as to a monkey DNA sequence that has homology to the SV40 origin and promotes expression of an unidentified gene product. Several high molecular mass proteins that interact with the transferrin receptor gene promoter have been identified. The activity of these proteins is transiently increased in 3T3 cells that have been stimulated by serum addition. This increase precedes a rise in transferrin receptor mRNA levels in the cytoplasm, which in turn precedes entry of the cells into S phase. DNase I footprinting of the transferrin receptor promoter reveals several protein binding sites. Two of the sites are within the conserved GC-rich region of the promoter. One of these binding sites probably interacts with Spl, while the second interacts with an uncharacterized protein.

Binding Sites↗
PubMed1986

Transcriptional regulation by iron of the gene for the transferrin receptor.

Treatment of K562 cells with desferrioxamine, a permeable iron chelator, led to an increase in the number of transferrin receptors. Increasing intracellular iron levels by treatment of cells with either human diferric transferrin or hemin lowered the level of the transferrin receptors. By using a cDNA clone of the human transferrin receptor, we showed that the changes in the levels of the receptor by iron were accompanied by alterations in the levels of the mRNA for the receptor. The rapidity of these changes indicated that the mRNA had a very short half-life. By using an in vitro transcriptional assay with isolated nuclei, we obtained evidence that this regulation occurred at the transcriptional level.

Cell Line↗
PubMed1985

Regional location of T cell receptor gene Ti alpha on human chromosome 14.

The chromosomal location of Ti alpha was determined by hybridization of a radiolabeled cDNA for the alpha chain of human T cell receptor with 12 human X mouse cell hybrid DNAs cleaved with BamHI. Seven hybrids contained human Ti alpha, while the remaining five lacked it. Only human chromosome 14 matched the distribution of human Ti alpha signal across the mapping panel. Hybrids segregating a chromosome 14 translocation were used to demonstrate that Ti alpha is in the region 14pter greater than 14q21. Thus, the alpha and beta chain genes that contribute structural components to the Ti moiety of the human T cell receptor lie on different chromosomes. In humans, the immunoglobulin heavy chain locus and Ti alpha are in different regions of chromosome 14, with Ti alpha more proximal and the immunoglobulin heavy chain locus more distal.

Animals↗
PubMed1985

Chromosomal locations of the murine T-cell receptor alpha-chain gene and the T-cell gamma gene.

Two independent methods were used to identify the mouse chromosomes on which are located two families of immunoglobulin (Ig)-like genes that are rearranged and expressed in T lymphocytes. The genes coding for the alpha subunit of T-cell receptors are on chromosome 14 and the gamma genes, whose function is yet to be determined, are on chromosome 13. Since genes for the T-cell receptor beta chain were previously shown to be on mouse chromosome 6, all three of the Ig-like multigene families expressed and rearranged in T cells are located on different chromosomes, just as are the B-cell multigene families for the Ig heavy chain, and the Ig kappa and lambda light chains. The findings do not support earlier contentions that genes for T-cell receptors are linked to the Ig heavy chain locus (mouse chromosome 12) or to the major histocompatibility complex (mouse chromosome 17).

Animals↗
PubMed1985

Proximity of thyroglobulin and c-myc genes on human chromosome 8.

The human thyroglobulin structural gene (TG) was mapped to the long arm of chromosome 8 by blot hydridization of a TG cDNA probe to DNA from 21 human X mouse somatic cell hybrids containing overlapping subsets of human chromosomes. In situ hybridization of the TG probe to metaphase chromosomes from a karyotypically normal human lymphoblastoid cell line, JS, localized the TG gene to within the region 8q23----q24.3. Thus, the TG and c-myc genes map to the same chromosome band in normal human cells. In a human colon carcinoma cell line (COLO 320 DM) which contains amplified c-myc, the TG gene is not amplified and hence it lies outside the amplification domain.

Animals↗
PubMed1985

Interferon-beta-related DNA on human chromosome 4.

A DNA subclone (pPE-4000) derived from the lambda B4 interferon-beta-related human genomic DNA clone was used as a probe in blot-hybridization experiments of DNA from a panel of human-rodent somatic cell hybrids containing overlapping subsets of human chromosomes. The DNA hybridization experiments showed that the lambda B4 IFN-beta locus is localized to human chromosome 4. A provisional regional assignment to 4q12-qter was also obtained. Thus available hybridization data implicate human chromosomes 2, 4, and 9 in the human IFN-beta system while the available biological data also implicated human chromosome 5.

Animals↗
PubMed1985

Chromosomal distribution of genes coding for fast twitch skeletal muscle myosin light chains.

The mouse fast twitch skeletal muscle myosin light chains are encoded by a multigene family which comprises the gene coding for the myosin light chain 2 (Myl2f), and the gene coding for both myosin light chains 1 and 3 (Myl1f/Myl3f). In addition, a Myl1f/Myl3f-related pseudogene is present in the domestic mouse Mus musculus. The members of this gene family were assigned to chromosomes by molecular hybridization, using DNA extracted from a panel of cloned mouse-Chinese hamster somatic hybrid cells and specific DNA probes. The genes coding for the light chains of the myosin molecule are dispersed on several chromosomes, while genes coding for the heavy chain of myosin are located on a single, different chromosome.

Animals↗
PubMed1985

Isolation and regional mapping of random X sequences from distal human X chromosome.

Chromosome-mediated gene transfer (CMGT) lines were shown to be convenient donors of genomic sequences from specific regions of the genome adjacent to selectable markers. Two libraries were prepared from CMGT lines carrying sequences spanning the long arm of the human X chromosome from HPRT (Xq26) to G6PD (Xq28). A series of 22 CMGT lines sharing the same selectable marker (HPRT) were used in conjunction with five standard translocation hybrids to provide fine-resolution regional mapping of the nonrepetitive X specific probes isolated from the libraries. The order of three human recombinant sequences with respect to known X-linked markers is: PGK (Xq13), 05-02 (DXS78); HPRT (Xq26), 07-03 (DXS79); surface antigen S11 (Xq27), 07-14 (DXS80); and G6PD (Xq28).

Animals↗
PubMed1985

In vitro activation of human sperm induced by amphibian egg extract.

In this paper we characterize a system for the activation of human sperm using cell-free extracts from Xenopus laevis eggs. We characterize the kinetics of sperm activation in terms of morphological and DNA synthetic parameters. We have optimized some of the components which are necessary for chromatin decondensation and present data to demonstrate that the in vitro process is efficient over a wide range of salt concentrations and amounts of supplementary reducing agents.

Animals↗
PubMed1985

Homeo box gene complex on mouse chromosome 11: molecular cloning, expression in embryogenesis, and homology to a human homeo box locus.

The homeo box is a 180 bp protein-coding domain found within homeotic genes of Drosophila and conserved in a variety of invertebrate and vertebrate species. It has been suggested that the mammalian homeo box sequences may play a role in controlling pattern formation during embryogenesis. We report findings that support this hypothesis. We have cloned three overlapping recombinant phage clones that cover a region of mouse chromosome 11 that contains a cluster of four homeo boxes (the Hox-2 locus). This locus encodes multiple transcripts that are expressed during embryogenesis. Forty kilobases of the Hox-2 region is devoid of repetitive elements and shows extensive homology with the human Hox-2 locus. These results provide direct evidence for genetic expression during embryonic development, a conserved organization in comparison to the cognate human locus, and a complexity of organization and transcript expression similar to that found in Drosophila.

Animals↗
PubMed1985

DNA-mediated genetic transformation of mouse embryos and bone marrow--a review.

In recent years, new gene transfer systems have been developed which allow molecularly cloned genetic material to be introduced into whole organisms. These systems include the microinjection of DNA into mammalian embryos, transfection of DNA into mouse bone marrow cells, and the infection of early embryos with retroviruses. Exogenous DNA appears to integrate randomly into the host genome. The production of transgenic mice by injection of DNA into mouse embryos has rapidly gained importance as an experimental tool for the study of gene regulation during development. Through this technique, recombinant molecules of any type can be introduced into one-celled embryos, and thus can be used to study development from its earliest stages. DNA sequences have been shown to integrate and transmit through the germ line to subsequent generations as mendelian traits. Transgenic mice carrying various gene constructs have been successfully exploited for the elucidation of factors which determine tissue specificity of gene expression as well as the level of gene control. Phenotypic changes related to expression of foreign genes have also been observed. This experimental approach thus promises to rapidly solve many of the heretofore most challenging problems in developmental genetics. Insertion of foreign genes has also made possible the creation of insertional mutants which manifest themselves most frequently as recessives. Such mutations can be readily studied at the molecular level by using the transferred material as a probe for recovery of the affected host sequence from genomic libraries. Many of these same problems have been addressed by introducing retroviral DNA into mouse embryos. Here, the sequences used for transfer have been limited to retroviral genes, but nonetheless these experiments have been profitably exploited for studies both of gene regulation and mutagenesis. Gene transfer systems are being developed allowing the experimenter to transfer DNA into bone marrow cells of mice, after which the recipient cells can be reintroduced into lethally irradiated histocompatible animals. This system has the advantage that selection can be applied during the gene transfer process such that the expression of the foreign material is assured. In addition, these experiments have created a model system for production of animals carrying a subpopulation of cells which is highly resistant to a toxic agent. This system has the potential for therapeutic application to man.

Animals↗
PubMed1985

Regional mapping of the phenylalanine hydroxylase gene and the phenylketonuria locus in the human genome.

Phenylketonuria (PKU) is an autosomal recessive disorder of amino acid metabolism caused by a deficiency of the hepatic enzyme phenylalanine hydroxylase (PAH; phenylalanine 4-monooxygenase, EC 1.14.16.1). A cDNA clone for human PAH has previously been used to assign the corresponding gene to human chromosome 12. To define the regional map position of the disease locus and the PAH gene on human chromosome 12, DNA was isolated from human-hamster somatic cell hybrids with various deletions of human chromosome 12 and was analyzed by Southern blot analysis using the human cDNA PAH clone as a hybridization probe. From these results, together with detailed biochemical and cytogenetic characterization of the hybrid cells, the region on chromosome 12 containing the human PAH gene has been defined as 12q14.3----qter. The PAH map position on chromosome 12 was further localized by in situ hybridization of 125I-labeled human PAH cDNA to chromosomes prepared from a human lymphoblastoid cell line. Results of these experiments demonstrated that the region on chromosome 12 containing the PAH gene and the PKU locus in man is 12q22----12q24.1. These results not only provide a regionalized map position for a major human disease locus but also can serve as a reference point for linkage analysis with other DNA markers on human chromosome 12.

Chromosome Mapping↗
PubMed1985

Use of a protein-blotting procedure and a specific DNA probe to identify nuclear proteins that recognize the promoter region of the transferrin receptor gene.

We describe a procedure for detecting high-affinity, sequence-specific DNA-binding proteins from crude nuclear extracts. The technique utilizes electrophoretic transfer of NaDodSO4/PAGE-fractionated proteins onto nitrocellulose filters. Incubation of the filters with a 5% (wt/vol) solution of nonfat dry milk effectively blocks nonspecific and low-affinity DNA-binding sites. Incubation of the blocked filters with radiolabeled DNA under optimal binding conditions and subsequent autoradiography reveals high-affinity DNA-protein interactions. We have used this procedure to identify proteins that bind specifically to the promoter region of the transferrin receptor gene.

Base Sequence↗