Biomedical subjects

H C Gorham

Publications and source records attributed to H C Gorham.

5 recordsLinked to original sources

PubMed1998

Mutation of yeast Ku genes disrupts the subnuclear organization of telomeres.

The mammalian Ku70 and Ku86 proteins form a heterodimer that binds to the ends of double-stranded DNA in vitro and is required for repair of radiation-induced strand breaks and V(D)J recombination [1,2]. Deletion of the Saccharomyces cerevisiae genes HDF1 and HDF2--encoding yKu70p and yKu80p, respectively--enhances radiation sensitivity in a rad52 background [3,4]. In addition to repair defects, the length of the TG-rich repeat on yeast telomere ends shortens dramatically [5,6]. We have shown previously that in yeast interphase nuclei, telomeres are clustered in a limited number of foci near the nuclear periphery [7], but the elements that mediate this localization remained unknown. We report here that deletion of the genes encoding yKu70p or its partner yKu80p altered the positioning of telomeric DNA in the yeast nucleus. These are the first mutants shown to affect the subnuclear localization of telomeres. Strains deficient for either yKu70p or yKu80p lost telomeric silencing, although they maintained repression at the silent mating-type loci. In addition, the telomere-associated silencing factors Sir3p and Sir4p and the TG-repeat-binding protein Rap1p lost their punctate pattern of staining and became dispersed throughout the nucleoplasm. Our results implicate the yeast Ku proteins directly in aspects of telomere organization, which in turn affects the repression of telomere-proximal genes.

Animals↗

PubMed1997

Chromosome ends: all the same under their caps.

The recent characterisation of subtelomeric regions from a variety of organisms from yeast to man has led to the realisation that all chromosome ends are similar in structure although maintenance of the terminus varies. The mosaic of repeats and proteins associated with telomeres has an architectural role which divides the genome into two domains, allowing for the adaptive use of the region as well as the evolution of non-telomerase-mediated telomere maintenance.

Adaptation, Biological↗

PubMed1996

Light-induced carotenogenesis in Myxococcus xanthus: light-dependent membrane sequestration of ECF sigma factor CarQ by anti-sigma factor CarR.

Light-induced carotenogenesis in Myxococcus xanthus is under the control of the carQRS operon. CarQ, a proposed extracytoplasmic (ECF) RNA polymerase sigma factor, is required for expression of the operon and the carC gene that encodes phytoene dehydrogenase. CarR, an inner membrane protein in Escherichia coli, is essential for carQRS promoter inactivation in the dark. CarS is required for the light-dependent expression of the promoter of the carB gene cluster that encodes the rest of the structural genes for carotenogenesis. Regulation of carQRS is dependent on the stoichiometry of CarQ and CarR. Increasing the copy number of carQ over carR led to constitutive carotenogenesis, as did loss of translational coupling between carQ and carR. The severity of the constitutive phenotype depended on the distance between the uncoupled genes. When expressed in M. xanthus, a CarR:beta-galactosidase fusion protein disappeared in the light. We propose that anti-sigma factor CarR sequesters CarQ to the membrane in the dark, but, in the light, loss of CarR leads to release of the sigma factor.

Bacterial Proteins↗

PubMed1994

Ricin B chain fragments expressed in Escherichia coli are able to bind free galactose in contrast to the full length polypeptide.

Deleted forms of ricin B chain (RTB) containing only one of the two galactose binding sites were produced in E. coli and targeted to the periplasm by fusion to the ompA or ompF signal sequences. The proteins were then isolated from the periplasm and their sugar binding properties assessed. Previous studies investigating the properties of such proteins produced in Xenopus laevis oocytes suggested that deleted forms of RTB, when not glycosylated, retain their ability to bind simple sugars, unlike the full-length unglycosylated proteins. When produced in E. coli however we found that only one, EB733, of a number of deleted forms of RTB closely related to those previously produced in Xenopus laevis oocytes, bound to simple sugars. All of the deletion forms of RTB were found to bind in the asialofetuin binding assay; an assay which has been previously utilized to measure binding of lectins to the terminal galactose residues of glycoprotein oligosaccharides. However, in contrast to glycosylated RTB, binding of the deletion mutants could be competed to only a small degree or not at all with galactose. The only deletion mutant observed to bind to free galactose when produced in E. coli corresponded closely to the complete domain 2 of RTB. It is assumed that this mutant forms a stable structure similar to that of the C-terminal domain in the full-length protein. The structural integrity of EB733 was not only suggested by its sugar binding properties and solubility but also by its consistently higher level of expression and the absence of any apparent susceptibility to E. coli proteases.

Base Sequence↗

PubMed1993

Light-induced carotenogenesis in Myxococcus xanthus: DNA sequence analysis of the carR region.

The carR region encodes a light-inducible promoter, a negative regulator of the promoter and a trans-acting activator that controls the light-inducible Myxococcus xanthus carotenoid biosynthesis regulon. DNA sequence analysis revealed, downstream of the promoter, three translationally coupled genes, carQ, carR and carS. Sequencing of mutations demonstrated that carR encoded the negative regulator and was an integral membrane protein. Mutant construction and sequencing revealed that carS was the trans-acting activator and that carQ was a positive regulator of the promoter. Neither gene encodes proteins with known sequence-specific DNA-binding motifs. The sequence of the light-inducible promoter region, identified by primer extension analysis, showed similarity to the consensus sequence of the Escherichia coli stress response ('heat-shock') promoters.

Amino Acid Sequence↗