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X M Yu

Publications and source records attributed to X M Yu.

44 records · Page 3Linked to original sources
PubMed1987

The telomeres of Tetrahymena ribosomal DNA are not sufficient for stabilizing linear DNA in Xenopus oocytes.

Restriction fragments that include the telomeres of ribosomal DNA from Tetrahymena thermophila (TtrDNA) were ligated to the ends of linearized simian virus 40 (SV40) DNA. The linear SV40 DNA with TtrDNA ends, circular SV40 DNA, linear SV40 DNA, and intact TtrDNA were injected into the nuclei of Xenopus laevis oocytes and assayed for stability. The intact linear 21-kb TtrDNA and circular SV40 DNA were maintained stably for at least 72 h after injection while the linearized SV40 DNA, either with or without telomeric ends, was degraded rapidly. Limited digestion with micrococcal nuclease revealed that neither the intact TtrDNA nor the SV40 DNA with telomeric ends reconstituted into chromatin containing regularly spaced nucleosomes. Another linearized plasmid DNA (pBamC), 14 kb in length, also was not stable in Xenopus oocytes with or without the addition of TtrDNA telomeres. Therefore, TtrDNA telomeres by themselves are not sufficient for stabilization of linear DNA in Xenopus oocytes. Rather, linear TtrDNA is maintained stably because of additional sequence or structural information encoded within the molecule.

Animals↗
PubMed1986

Deletion analysis of RNA polymerase interaction sites in the Escherichia coli lactose operon regulatory region.

Two sets of deletions produced in vitro by S1 nuclease were used to study the structure and function of promoters lacPUV5 and lacP115. The upstream boundary of the RNA polymerase binding site in lacPUV5 was determined by comparing the levels of beta-galactoside expression in vivo programmed from a set of deletions progressively extending into the -35 region of the lacPUV5 promoter. Sequences upstream from base-pair -37 are not necessary for the full functioning of lacPUV5. A deletion that removes base-pair -37 retains only half of the promoter activity. Deletion of the first T X A base-pair of the consensus -35 region sequence, 5' T-T-T-A-C-A 3', leads to a sixfold reduction of promoter activity. Deletion of the whole -35 region of lacPUV5 leads to at least a 20-fold reduction of its promoter activity. Abortive initiation assays were performed on the fully functional lacPUV5 and two lacPUV5 deletions, which removed part of the -35 consensus sequence, to study their effect on the kinetics of RNA polymerase-promoter open complex formation. These two deletions show a 3.5 to 7-fold reduction in KB. Analysis in vivo of lacP115 showed that sequence information upstream from the -35 region is important for the full functioning of lacP115. A deletion removing sequences upstream from -41 caused a three- to fourfold reduction in promoter activity, apparently due to reduced transcription initiation. lacP115 has a much lower k2 value than lacPUV5; its KB value is about threefold higher than that of lacPUV5.

Base Sequence↗
PubMed1985

Deletion analysis of the Escherichia coli lactose promoter P2.

The Escherichia coli lactose (lac) operon transcription control region includes at least two sequences which are recognized by RNA polymerase holoenzyme in vitro, the normal lac promoter (termed P1) and an overlapping upstream promoter (termed P2). The structure of the P2 and the effect of RNA polymerase interaction at P2 on the association of RNA polymerase with P1 was analyzed by the isolation and characterization of various mutations at P2. A set of deletions with varying lengths of DNA between the lac P2 -10 region and a "-35 region" contributed by the vector DNA were constructed. In vitro studies indicate that as the spacing between the -10 region and "-35 region" is increased from 16 to 22 base pairs (bp), the steady state occupancy as measured by exonuclease III protection experiments and the ability to initiate transcripts from P2 decrease. Studies were also conducted using a single base pair insertion and a two base pair deletion between the natural -35 and -10 regions of P2. The mutation which decreases the in vitro occupancy and transcription initiation potential of P2 does not significantly affect the steady state in vitro occupancy of P1 nor the in vivo expression of the lac operon. These results are not consistent with the model that RNA polymerase occupancy at P2 competes with the P1 expression and therefore that this competition plays a role in cAMP bound catabolite gene activator protein (CAP-cAMP) control of the lac operon.

Base Sequence↗
PubMed1984

Deletion analysis of the CAP-cAMP binding site of the Escherichia coli lactose promoter.

S1 nuclease was used to generate a series of deletions which extend into the CAP-cAMP binding site from upstream of the Escherichia coli lactose operon promoter (lacP). Deletion and insertion mutations were also created which changed the spacing region between the CAP-cAMP binding site and the lacP -35 region. The promoter activities of these mutations were compared by measuring the levels of beta-galactosidase gene expression in vivo. The results show that sequence information prior to 74 base pairs (-74) upstream from the transcription start site (designated as +1) is not necessary for the full activation of the lac promoter by the CAP-cAMP complex. However, the deletion which extends to the -71 position retains only one third of the promoter activity in the presence of the CAP-cAMP complex. Removal of one symmetrical element from the two fold symmetry in the CAP-cAMP binding site abolished the CAP-cAMP stimulation of the lac promoter. Spacer mutations which increase by one base pair or decrease by two base pairs the length of the spacing region between the CAP-cAMP binding site and the lacP -35 region drastically reduced the CAP-cAMP stimulation of the lac promoter. This suggests that the distance between the lac promoter transcription start site and CAP-cAMP binding site is crucial for the function of the lac promoter, despite the fact that this distance varies in other E. coli promoters positively regulated by CAP-cAMP. A deletion which extends to the -59 position results in a two fold enhanced expression of lac in the absence of CAP-cAMP. This is consistent with the existance of a competitive RNA polymerase binding site in this region which would normally act to inhibit RNA polymerase binding.

Base Sequence↗
PubMed1984

Molecular cloning and sequence analysis of trp-lac fusion deletions.

DNA fragments containing deletions that fuse the trp operon to the lac operon were cloned and the end-points of the fusions were determined. The results from DNA sequence analysis correlated well with those from genetic, biochemical and physiological studies previously reported. The sequence data from this study, in combination with the known properties of these fusion strains, provided information on: (1) the precise lac operon distal boundary of the lac operator; (2) the nature of the trp operon terminator; and (3) the messenger RNA sequences that result in inhibition of lacZ translation initiation in trp-lac fused mRNAs.

Amino Acid Sequence↗
PubMedDate not supplied

Vasopressin mRNA and neurophysin-related cell-surface antigen (NRSA) in small-cell carcinoma.

Production by small-cell carcinoma (SCCL) of neurophysins (HNPs) and neurophysin-related cell-surface antigen (NRSA) was examined for two cell lines, for mouse xenografts, and for a resected human tumor, using polyclonal and monoclonal antibodies to vasopressin-associated human neurophysin (VP-HNP) and polyclonal antibodies to vasopressin (VP). The nature of the mRNA responsible for giving rise to these neurophysin-related products was investigated by performing Northern analysis on preparations of poly A+RNA and cDNA probes complimentary to portions of the exon A, exon B, and exon C regions of the human VP gene. SDS-electrophoresis and Western analysis revealed two prominent proteins of 42,000 and 20,000 Da in acid extracts from all SCCL sources when the monoclonal anti-HNP or one of the two polyclonal anti-HNP preparations were used. These antibodies also disclosed the presence of a minor component of 10,000 Da. A second polyclonal anti-HNP preparation reacted with one prominent protein of 30,000 Da and, for one cell line and mouse xenografts, another protein of 32,000 Da. Both of two anti-VP preparations reacted with proteins of 42,000, 30,000, 25,000, and 20,000 Da in extracts from all SCCL source material. The immunoreactive proteins of 42,000, 30,000, and 20,000 Da were all components of a membrane fraction from SCCL cells and tissues. In Northern analysis, a single RNA of about 900 bases hybridized with exon A and exon B probes, but not with the cDNA probe complimentary to exon C of the VP gene.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMedDate not supplied

Biosynthesis of vasopressin by gastrointestinal cells of Brattleboro and Long-Evans rats.

The distribution of vasopressin, provasopressin, vasopressin-associated neurophysin, and vasopressin-associated glycopeptide was determined immunohistochemically in the gastrointestinal system of Brattleboro and Long-Evans rats. Cells containing immunoreactivity for vasopressin, provasopressin, neurophysin, and glycopeptide were detected in the same cell types of the stomach and duodenum, while selected cells in the duodenum contained only immunoreactive glycopeptide. Unlike that in the hypothalamus, staining for neurophysin in the gastrointestinal tract was sensitive to fixation. These findings indicate that vasopressin is produced by cells in the rat gastrointestinal system and suggest the existence of synthetic pathways different from those found in hypothalamic neurons.

Animals↗