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C K Tuggle

Publications and source records attributed to C K Tuggle.

68 records · Page 4Linked to original sources
PubMed1993

Cloning and restriction fragment length polymorphism analysis of a cDNA for swine PIT-1, a gene controlling growth hormone expression.

A partial swine cDNA which encodes the functional domain of PIT-1 was isolated by the polymerase chain reaction (PCR). The swine PIT-1 cDNA clone is 95% identical at the protein level to the rat Pit-1 gene. Thus, Pit-1's known function in control of rat growth hormone and prolactin expression is likely to be conserved in swine. This swine cDNA clone was used to investigate genetic variability at PIT-1 in several American and Chinese breeds. Polymorphic BamHI fragments were found in pure-bred Meishan animals (n = 13), but only monomorphic fragments in five American breeds (n = 36).

Amino Acid Sequence↗
PubMed1990

Region-specific enhancers near two mammalian homeo box genes define adjacent rostrocaudal domains in the central nervous system.

To gain insight to the mechanisms underlying region-specific gene expression in mammalian development, we investigated the regulatory DNA associated with the proximal promoter of two homeo box genes, murine Hox-1.3 and human Hox-5.1. Using lacZ gene fusions in transgenic mice, we identified regulatory elements in the 5'-flanking sequences of the Hox-1.3 and the Hox-5.1 genes that specifically direct beta-galactosidase expression to the brachial and the upper cervical regions (respectively) of the central nervous system (CNS). These two elements act at the transcriptional level, are active in either orientation, and confer region-specific expression to unrelated promoters, satisfying the criteria for enhancer elements. The two spatial domains defined by these enhancers are directly adjoining, extend along the rostrocaudal axis for the same span of 6-7 metameres, and represent specific subsets of the overall CNS regions expressing all endogenous Hox-1.3 or Hox-5.1 transcripts. The adjacent domains in the developing murine CNS that express Hox-1.3 and Hox-5.1 gene fusions are strikingly reminiscent of the adjacent stripes of expression in Drosophila embryos seen with Sex combs reduced and Deformed, the two Drosophila homeotic genes most homologous to Hox-1.3 and Hox-5.1, respectively. These findings represent the first demonstration of region-specific mammalian enhancers and raise the possibility that the mammalian CNS may be subdivided into a series of rostrocaudal domains on the basis of the activity of enhancers near homeo box genes.

Animals↗
PubMed1990

Regulation of the operon encoding ribonucleotide reductase: role of the negative sites in nrd repression.

Expression of the nrd genes was previously shown to be controlled by both positive and negative regulation (C. K. Tuggle and J. A. Fuchs, EMBO J. 5:1077-1085, 1986). Two regions, one located 5' and one located 3' of the nrd promoter (nrdP), were identified as negative regulatory sites since deletion of these sequences increased nrd expression. These regions of DNA have sequence similarities, and a looping mechanism was proposed to explain the requirement for two distinct sites in nrd repression. To investigate the role of these sequences in regulating nrd, a gel electrophoresis assay was used to detect the proteins that bind to the nrd regulatory sites. A protein that bound to restriction fragments containing the negative regulatory sites but not to other DNA fragments was identified in cell extracts and was partially purified. DNase I footprinting experiments showed that the binding protein protects the 5' negative site previously identified in vivo. The 3' negative site also identified in vivo was not required in vitro for high-affinity protein binding to the 5' site, but lower-affinity binding to this site could be detected. Specific binding to the 5' site was found to be elevated approximately 10-fold in crude extracts from thymine-starved cells as compared with that in extracts from unstarved cells. This higher activity was also evident in purified preparations, suggesting that thymine starvation increases the expression of the negative regulatory protein. The finding that a purified protein preparation binds both negative regulatory sites indicates that this preparation contains the nrd repressor protein or proteins. Insertion of 37 base pairs (3.5 helix turns) of DNA at a HpaII site or 35 base pairs (3.3 turns) at a MnlI site between the 5' regulatory sites and nrdP abolished the increase in nrd expression resulting from thymine starvation in vivo, but negative regulation appeared to be less affected than when either negative site was deleted. Insertion of DNA in these constructs was shown not to affect repressor binding in vitro, indicating either that a simple model of DNA looping to bring equivalent operator sites into physical proximity does not explain repression at nrd or that the distance between sites is sufficient that helical turns are of little importance.

Base Sequence↗
PubMed1990

The use of lacZ gene fusions in the studies of mammalian development: developmental regulation of mammalian homeobox genes in the CNS.

1. Mammalian cells can produce bacterial beta-galactosidase when they carry the lacZ gene under the control of mammalian regulatory elements. The single cell resolution of the beta-galactosidase histochemical detection method makes this molecule an excellent marker in studies of development at the cellular and molecular level. Different lacZ fusion genes can be engineered to study the histological diversification of cell lineages, the developmental regulation of isolated genes, or to recognize and clone genes with new expression profile. 2. Transgenic mice carrying lacZ gene fusions provide information on the cell type, developmental stage and spatial specificity of cis-acting regulatory regions linked to a mammalian homeobox gene. We describe our strategy for designing the gene fusions. 3. The scord region of the Hox 1.3 gene is sufficient to determine spatially restricted expression of a heterologous protein in the midgestational spinal cord. We propose to use this region to alter the expression pattern of other homeobox gene products. Developmental alterations due to a variant expression pattern would point to the function of the misexpressed gene.

Animals↗
PubMed1988

Spatial regulation of homeobox gene fusions in the embryonic central nervous system of transgenic mice.

The spatially restricted expression of mammalian homeobox genes in teh embryonic central nervous system (CNS) provides an opportunity to study the basis of spatial gene regulation in mammalian development. Here, we define a regulatory region of the murine Hox 1.3 gene that mediates such a region-specific expression pattern. The Hox 1.3 gene contains two exons, encodes a putative protein of 270 amino acids, and is expressed preferentially in the spinal cord at midgestation. We have analyzed transgenic mice containing various Hox 1.3 DNA fragments fused to reporter sequences, such as a human growth hormone gene fragment or the E. coli lacZ structural gene. As shown by RNAase protection assays or by in situ analyses of beta-galactosidase activity, several hybrid genes are expressed in the embryonic central nervous system in a spatially restricted manner, along both the rostrocaudal and dorsoventral axes. A 912 nucleotide sequence located immediately upstream of the Hox 1.3 coding sequence is sufficient to direct expression to the dorsolateral cells of the brachial spinal cord.

Amino Acid Sequence↗
PubMed1986

Regulation of the operon encoding ribonucleotide reductase in Escherichia coli: evidence for both positive and negative control.

The ribonucleotide reductase genes (nrd) are induced by thymine starvation. Deletion analysis of the sequences upstream of the cloned nrd genes was used to identify several regulatory regions. The start of transcription (nrdP) was mapped 110 bp upstream of nrdA, the first structural gene. A site required for positive regulation of nrd was mapped 135 bp upstream of nrdP in a region with two direct repeat sequences as well as potential secondary structure. Two other sites (one upstream of nrdP, the other downstream) were identified as sequences whose deletion markedly increase expression. These latter sites show sequence homology and probably interact since the effects of their individual deletion are not additive when combined.

Base Sequence↗
PubMed1985

Glutathione reductase is not required for maintenance of reduced glutathione in Escherichia coli K-12.

Seven independently isolated glutathione reductase-deficient (gor) Escherichia coli mutants were found to have an in vivo glutathione redox state that did not significantly differ from that of the parental strain, 98 to 99% reduced. Strains containing both a gor mutation and either a trxA mutation (thioredoxin deficient) or a trxB mutation (thioredoxin reductase deficient) were able to maintain a 94 to 96% reduced glutathione pool, suggesting that glutathione can be reduced independently of glutathione reductase and thioredoxin reductase.

Escherichia coli↗