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Biomedical subjects

R Aasland

Publications and source records attributed to R Aasland.

32 records · Page 2Linked to original sources
PubMed1991

Human uracil-DNA glycosylase complements E. coli ung mutants.

We have previously isolated a cDNA encoding a human uracil-DNA glycosylase which is closely related to the bacterial and yeast enzymes. In vitro expression of this cDNA produced a protein with an apparent molecular weight of 34 K in agreement with the size predicted from the sequence data. The in vitro expressed protein exhibited uracil-DNA glycosylase activity. The close resemblance between the human and the bacterial enzyme raised the possibility that the human enzyme may be able to complement E. coli ung mutants. In order to test this hypothesis, the human uracil-DNA glycosylase cDNA was established in a bacterial expression vector. Expression of the human enzyme as a LacZ alpha-humUNG fusion protein was then studied in E. coli ung mutants. E. coli cells lacking uracil-DNA glycosylase activity exhibit a weak mutator phenotype and they are permissive for growth of phages with uracil-containing DNA. Here we show that the expression of human uracil-DNA glycosylase in E. coli can restore the wild type phenotype of ung mutants. These results demonstrate that the evolutionary conservation of the uracil-DNA glycosylase structure is also reflected in the conservation of the mechanism for removal of uracil from DNA.

Amino Acid Sequence↗
PubMed1991

Presence of cfaD-homologous sequences and expression of coli surface antigen 4 on enterotoxigenic Escherichia coli; relevance for diagnostic procedures.

We examined the ability of a colonization factor antigen I (CFA/I) polynucleotide probe to identify coli-surface antigen 4 producing (CS4+) strains of enterotoxigenic Escherichia coli (ETEC). At low stringency (LS) the probe hybridized to colony lysates of strains previously shown to produce CS4 or CFA/I fimbriae. Only DNA from CFA/I+ strains maintained a stable probe-target hybrid under high stringency (HS) conditions. On examination of several clones from three previous CS4 producers, identified as positive in LS and negative in HS colony hybridization, spontaneous loss of nucleotide sequences homologous to a gene encoding a positive CFA/I regulator, CfaD, was found to be associated with lacking expression of CS4. Our findings indicate that, on stored or subcultured isolates of ETEC, identification of CS4 strains may benefit from applying gene probe technology.

Antigens, Bacterial↗
PubMed1990

Co-expression of the genes encoding transforming growth factor-alpha and its receptor in papillary carcinomas of the thyroid.

Transforming growth factor-alpha (TGF-alpha) is frequently coexpressed with its receptor, epidermal growth factor receptor (EGF-R), in several types of carcinoma and sarcoma. It is believed that this results in an autocrine stimulation of tumor growth in these tumors. We have found that TGF-alpha and EGF-R/c-erbB RNAs were co-expressed at significantly higher levels in papillary thyroid carcinomas and their lymph-node metastases than in non-neoplastic thyroid tissues. We also observed a low level of expression of RNA specific for insulin-like growth factor I in these tumors, which was highest in a lymph-node metastasis. Autocrine stimulation by TGF-alpha may thus be a common feature of papillary carcinomas of the thyroid. Since EGF is known to induce proliferation and dedifferentiation of normal thyroid cells in culture, TGF-alpha and its receptor may play an important role in thyroid carcinogenesis.

Adolescent↗
PubMed1990

Absence of abnormalities of the c-erbB-1 and c-erbB-2 proto-oncogenes in human thyroid neoplasia.

The c-erbB-1 and c-erbB-2 proto-oncogenes are frequently activated by gene amplification and overexpression in a variety of human cancers. In an analysis of a large series of benign and malignant thyroid tumours, no abnormalities of structure or expression of either of c-erbB-1 or c-erbB-2 were found. Activation of these oncogenes is not a necessary event in neoplasia of this epithelial system.

DNA, Neoplasm↗
PubMed1989

Molecular cloning of human uracil-DNA glycosylase, a highly conserved DNA repair enzyme.

Uracil-DNA glycosylase is the DNA repair enzyme responsible for the removal of uracil from DNA, and it is present in all organisms investigated. Here we report on the cloning and sequencing of a cDNA encoding the human uracil-DNA glycosylase. The sequences of uracil-DNA glycosylases from yeast, Escherichia coli, herpes simplex virus type 1 and 2, and homologous genes from varicella-zoster and Epstein-Barr viruses are known. It is shown in this report that the predicted amino acid sequence of the human uracil-DNA glycosylase shows a striking similarity to the other uracil-DNA glycosylases, ranging from 40.3 to 55.7% identical residues. The proteins of human and bacterial origin were unexpectedly found to be most closely related, 73.3% similarity when conservative amino acid substitutions were included. The similarity between the different uracil-DNA glycosylase genes is confined to several discrete boxes. These findings strongly indicate that uracil-DNA glycosylases from phylogenetically distant species are highly conserved.

Amino Acid Sequence↗
PubMed1988

Optimal conditions for hybridization with oligonucleotides: a study with myc-oncogene DNA probes.

We present a study on the refinement of filter-hybridization conditions for a series of synthetic oligonucleotides in the range from 17 to 50 base residues in length. Experimental conditions for hybridization and the subsequent washing steps of the filter were optimized for different lengths of the synthetic oligonucleotides by varying the formamide concentration and washing conditions (temperature and monovalent cation concentration). Target DNA was immobilized to the nitrocellulose filter with the slot blot technique. The sequences of the synthetic oligonucleotides are derived from the third exon of the human oncogene c-myc and the corresponding viral gene v-myc and the G + C content was between 43 and 47%. Optimal conditions for hybridization with a 82% homologous 30-mer and 100% homologous 17-, 20-, 25-, 30-, and 50-mers were found to be a concentration of formamide of 15, 15, 30, 30, 40, and 50%, respectively. Optimal conditions for washing were 0.5X standard sodium citrate (SSC) at 42 degrees C for 2 X 15 min. The melting temperature for these optimal hybridization and washing conditions was calculated to be up to 11 degrees C below the hybridization temperature actually used. This confirms that the duplexes are more stable than expected. The melting points for 17-, 20-, and 30-mers were measured in the presence of 5X SSC and found to be 43, 58, and 60 degrees C, respectively. Competition between double- and single-stranded DNA probes to the target DNA was investigated. The single-stranded DNA probes were about 30- to 40-fold more sensitive than the double-stranded DNA probes.

Base Sequence↗
PubMed1988

Expression of oncogenes in thyroid tumours: coexpression of c-erbB2/neu and c-erbB.

The receptor-type oncogenes c-erbB2/neu and c-erbB have been found amplified and/or overexpressed in a number of tumours of epithelial origin. We have studied the expression of oncogenes in biopsies from human thyroid tumours. The c-erbB2/neu and c-erbB oncogenes showed two- to three-fold higher levels of RNA in papillary carcinomas and lymph node metastases as well as in one adenoma when compared to non-tumour tissue. The nuclear oncogenes c-myc and c-fos were found to be expressed at varying levels in both non-tumour and tumour tissue. RNA transcripts specific for the platelet-derived growth factor A and B chains and the N-ras oncogene were detected in one anaplastic carcinoma. Neither rearrangements nor amplifications of oncogenes were observed in the thyroid tumours. These data are particularly interesting in light of the recent findings that epidermal growth factor induces proliferation and dedifferentiation of normal thyroid epithelial cells in vitro. We suggest that the epidermal growth factor or other ligands for the c-erbB and c-erbB2/neu receptors may contribute to the development and/or maintenance of the malignant phenotype of papillary carcinomas of the thyroid.

Adenoma↗
PubMed1988

Identity of the 17-kilodalton protein, a DNA-binding protein from Escherichia coli, and the firA gene product.

The 17-kilodalton protein, a DNA-binding protein encoded by the skp gene of Escherichia coli, was found to be identical to histonelike protein I, the product of the firA gene. This conclusion was reached after chromosomal localization, using the recently constructed high- and low-resolution E. coli restriction maps, and by direct comparison of the N-terminal amino acid sequence of histonelike protein I and the 17-kilodalton protein.

Chromosome Mapping↗
PubMed1987

Purification and characterization of the 17 K protein, a DNA-binding protein from Escherichia coli.

A basic protein of molecular mass 17 kDa (protein 17 K) which binds to relaxed DNA has been isolated and purified to homogeneity from Escherichia coli cells. The protein behaves as a tetramer in solution and there are 4800 monomers per cell in exponentially growing cells. The amino-acid composition and N-terminal sequence were determined. No effect of the protein on in vitro transcription was observed. The protein was shown to be different from the Ssb protein (Sigal, N. et al. (1972) Proc. Natl. Acad. Sci. USA 69, 3537-3541), protein H1 (Cukier-Kahn et al. (1972) Proc. Natl. Acad. Sci. USA 69, 3643-3647) and the HLP-1 protein (Lathe, R. et al. (1980) Proc. Natl. Acad. Sci. USA 77, 3548-3552).

Amino Acid Sequence↗
PubMed1987

DNA- and RNA-binding proteins of chromatin from Escherichia coli.

The different proteins present in chromatin of Escherichia coli have been analyzed by a variety of techniques. The chromatin was isolated using a previously published procedure (Sjåstad, K., Fadnes, P., Krüger, P.G. Lossius, I. and Kleppe, K. (1982) J. Gen. Microbiol. 128, 3037) and solubilized by the action of micrococcal nuclease or DNAase I. The DNA-protein and RNA-protein complexes thus obtained were purified by sucrose gradient centrifugation and isopycnic gradient centrifugation in metrizamide in low ionic strength. The protein: DNA ratio of the DNA-protein complexes was estimated from the latter method and found to be approx. 1.75. The protein components were analyzed further by one- and two-dimensional gel electrophoresis. Approx. 15 major polypeptides were detected in the DNA-protein complex, whereas 10 were present in the RNA-protein complex. The majority of the polypeptides in both complexes had acidic isoelectric pH. The polypeptides in the two complexes differed markedly and only two polypeptides, having molecular weights of 57,000 and 37,000, respectively, were found to be common in both complexes. In agreement with earlier studies, the basic protein HU was not present in the DNA-protein complex. Affinity studies of the proteins from chromatin using DNA- and RNA-Sepharose columns in general confirmed the above conclusions. The two-dimensional gel electrophoretic patterns of the proteins in the different complexes were compared with those of proteins in the inner and outer membranes. Only one of the major polypeptides present in the inner membrane, having a molecular weight of 57,000, was enriched in the DNA-protein complex.

Carrier Proteins↗