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G Westin

Publications and source records attributed to G Westin.

48 records · Page 3Linked to original sources
PubMed1983

Loci for human U1 RNA: structural and evolutionary implications.

Three clones U1-1, U1-6, and U1-8 containing sequences related to human U1 RNA have been studied by sequence analysis. The results show that each of the three clones represents a distinct locus. The U1-6 locus is closely related to the HU1-1 locus, which is believed to represent a functional U1 gene. The U1-1 and U1-8 loci are pseudogenes by definition, since they contain sequences that are closely related to but not identical with the human U1 RNA sequence. The U1-6 locus contains the sequence T-A-T-A-T close to the 5'-end of the U1 sequence but it is unclear if this represents the promoter. When the U1-8 locus was compared to the U1-6 locus, it was observed that the 5'-flanking sequences, except in the immediate vicinity of the pseudogene, are as well-conserved as the U1-related sequence itself, at least up to position -220. The high degree of homology in the 5'-flanking region suggests that U1 genes have a much more strict sequence requirement with regard to 5'-flanking sequences than most other eukaryotic genes. The U1-6 and U1-8 loci contain the sequence T-A-T-G-T-A-G-A-T-G-A between positions -211 and -221. An identical sequence is present in the equivalent position in the HU1-1 locus, and may represent the promoter. The high degree of conservation in the postulated promoter region indicates that pseudogenes like U1-8 possibly could be expressed. A truncated U1-related sequence is present between 106 to 150 nucleotides upstream from the U1 gene/pseudogene in the U1-6, the U1-8 and the HU1-1 loci, suggesting that the U1 genes may have been clustered early in evolution. The U1-1 locus has a strikingly different structure from the U1-8 locus; the pseudogene itself is as closely related to the U1 RNA sequence as is the U1-8 pseudogene but the flanking sequences, both on the 5' and the 3' side, share no detectable homology with the corresponding regions in the U1-6 or U1-8 loci. It may therefore be postulated that small nuclear RNA pseudogenes are created by several different mechanisms.

Base Sequence↗
PubMed1982

In vitro transcription of phage T4 late genes on purified DNA by partially purified RNA polymerase from T4-infected Escherichia coli b cells.

RNA polymerase was purified from 'late' phage T4-infected Escherichia coli B cells by DNA-cellulose affinity chromatography and high salt agarose filtration. The DNA-cellulose-purified RNA polymerase preparation contained T4-coded DNA endonuclease activity and several proteins, some with sizes comparable with the known T4 maturation factors, essential for late RNA synthesis. Some of these proteins, and the DNA endonuclease utilizing native, parental T4 DNA and supercoiled phi X 174 DNA as substrates, were partially separated from the RNA polymerase as a complex during agarose filtration. In vitro RNA was made by the DNA-cellulose-purified RNA polymerase using native, parental T4 DNA as template. About 26% of the in vitro RNA was transcribed from the DNA r-strand; 75% from the same r-strand region as in vivo late after infection. Both the abundancy and specificity of the in vitro r-strand transcription were markedly reduced after agarose filtration of the enzyme. Addition of the proteins separated from the RNA polymerase during agarose filtration caused a restoration of in vitro r-strand transcription abundance, but not its specificity. These results show that partially purified RNA polymerase from T4-infected E. coli B cells was able to transcribe late T4 genes in vitro with some abundancy and specificity on purified, parental T4 DNA, but further purification of the enzyme caused an irreversible reduction of this ability.

Bacterial Proteins↗
PubMed1982

A candidate gene for human U1 RNA.

Clones containing sequences complementary to the small nuclear RNA U1 were isolated from the human DNA library of Lawn et al. (1978). Three clones were studied by hybridization and restriction enzyme cleavage. The results showed that the inserts in all three clones were different and that each clone contains one single copy of a sequence which hybridizes to U1 RNA. The results revealed moreover that only one of the three clones contains all the cleavage sites which can be predicted from the known sequence of human U1 RNA, suggesting that the three clones comprise one candidate U1 gene and two pseudogenes. A fragment from the recombinant with the candidate U1 gene was subcloned in the pPR322 plasmid and part of its sequence was determined. The results showed that the subclone contains a sequence which matches that of the human U1 RNA perfectly. The sequence "TATAT" which often is found adjacent to RNA polymerase II start sites, was identified 33-37 base pairs upstream from the beginning of the U1 sequence. Two ten base pairs long, nearly perfect, direct repeats were also identified in the vicinity of the U1 sequence and an imperfect inverted repeat follows immediately after the U1 gene.

Base Sequence↗
PubMed1982

A pseudogene for human U4 RNA with a remarkable structure.

The human DNA library of Lawn et al. (1978) was screened for sequences complementary to the small nuclear (sn) RNA U4. Several positive clones were identified by screening 100 000 recombinants, indicating that U4 sequences like other snRNA sequences are dispersed in the human genome. One recombinant was characterized in detail by subcloning a Bg/II fragment 1.9 kilobases (kb) long in the pBR322 plasmid. The subcloned fragment was partially sequenced and the results revealed a pseudogene for U4 RNA. The pseudogene was found to have a remarkable structure; it contains a sequence that, except in two positions, matches the first 68 nucleotides of the human U4 RNA sequence and the pseudogene is, moreover, flanked by perfect direct repeats 20 bp long. The results support the model of van Arsdell et al. (1981) suggesting that certain snRNA pseudogenes arise by reverse transcription of the RNA followed by integration of the cDNA copy at a new chromosomal locus.

Base Composition↗
PubMed1982

Sequence organization of a viral DNA insertion present in the adenovirus-type-5-transformed hamster line BHK268-C31.

The hamster cell line BHK268-C31 contains two large viral inserts which both include sequences from the left-hand end of adenovirus type 5 (Ad5) DNA. One of these viral inserts has been cloned in the lambda vector Charon 4A. Electron microscopic analysis and restriction enzyme mapping shows that the recombinant carries a 4.4-kb-long colinear segment of viral DNA, which is located between map positions 1.5 and 14.2 in the Ad5 genome. The junctions between viral DNA and flanking sequences have been sequenced and found not to show any specific features. One of the junctions is located in the E1a coding region, 573 bp from the left-hand end of the Ad5 genome, whereas the other junction is situated in the coding region for polypeptide IVa2. The promoter region as well as the cap site for the mRNAs from region E1a are thus missing from this insert and its role in viral transformation is unclear.

Animals↗
PubMed1981

Human DNA sequences complementary to the small nuclear RNA U2.

Clones containing sequences complementary to the small nuclear RNA U2 were isolated from a human DNA library (1). Three clones, designated U2/4, U2/6 and U2/7 were purified and characterized by restriction enzyme cleavage, hybridization and heteroduplex analysis. Hybridization showed that the three clones each contained one single region which is complementary to U2 RNA. Restriction enzyme cleavage revealed furthermore that the inserted fragments in the three recombinants are different. Heteroduplex analysis identified a 240-380 bp long duplex region in each heteroduplex which includes sequences complementary to U2 RNA. Heteroduplexes between clones U2/4 and U2/7 as well as between U2/4 and U2/6 revealed two additional approximately 200 bp long homologies. The remainder of the inserts were found to lack measurable sequence homology. Two fragments from clone U2/4 were subcloned in the pBR322 vector and the subclones were used to determine the nucleotide sequence of a region in clone U2/4 which is complementary to U2 RNA. A comparison between the established sequence and the sequence for rat U2 RNA (2) reveals several discrepancies.

Base Sequence↗
PubMedDate not supplied

Identification of a CpG island in the human LRP-2 gene and analysis of its methylation status in parathyroid adenomas.

Methylation of CpG residues in mammalian genomes is a mechanism of vital importance for many cellular functions, which all relate to gene expression. In this study we describe the identification of a CpG island in the 5'-region of the gene encoding human megalin/LRP-2, a receptor capable of binding multiple ligands, which is involved in the regulation of calcium metabolism. Southern blot analysis and genomic bisulfite sequencing revealed that the CpG island is methylated in a non-expressing cell line, largely unmethylated in an expressing cell line and unmethylated in human parathyroid tissue. In addition, we show that artificial methylation of LRP-2 promoter reporter plasmids leads to strong transcriptional repression, in vitro as well as in transfected cells. No evidence for aberrant LRP-2 gene methylation in parathyroid adenomas, in which the LRP-2 protein is generally down-regulated, was found.

Adenoma↗