Biomedical subjects

U Hansen

Publications and source records attributed to U Hansen.

6 recordsLinked to original sources

PubMed2000

Inhibition of the mammalian transcription factor LSF induces S-phase-dependent apoptosis by downregulating thymidylate synthase expression.

The thymidylate synthase (TS) gene, which is induced at the G(1)-S transition in growth-stimulated cells, encodes an enzyme that is essential for DNA replication and cell survival. Here we demonstrate that LSF (LBP-1c, CP2) binds to sites within the TS promoter and intronic regions that are required for this induction. Mutation of the LSF binding sites inhibits G(1)-S induction of mRNA derived from a TS minigene. Furthermore, expression of dominant-negative LSF (LSFdn) prevents the increase in TS enzyme levels during G(1)-S, and induces apoptosis in growth- stimulated mouse and human cell lines. Such apoptosis can be prevented either by circumventing the TS requirement through addition of low concentrations of thymidine, or by coexpression of the TS gene driven by a heterologous promoter. Induction of apoptosis by LSFdn parallels the process known as thymineless death, which is induced by the TS inhibitor and chemotherapeutic drug 5-fluorodeoxyuridine. Thus, LSF is a novel regulatory factor that supports progression through S-phase by targeting a single gene that is critical for cell survival.

Amino Acid Sequence↗

PubMed1992

Activation of RNA polymerase II transcription by the specific DNA-binding protein LSF. Increased rate of binding of the basal promoter factor TFIIB.

While the components of the initiation complex at an RNA polymerase II basal promoter have been well characterized, few mechanistic studies have focused on how upstream DNA-binding, transcriptional activators influence protein assembly at the initiation site. Our analysis of basal transcription on both the simian virus 40 and adenovirus major late promoters demonstrates that two slow steps in initiation of transcription are the assembly of the general transcription factors TFIID and TFIIB onto the template DNA. On the simian virus 40 major late promoter, the rate of initiation complex formation is dramatically increased in the presence of the cellular transcriptional activator LSF. Direct analysis by band mobility shift assays demonstrates that LSF has no effect on the rate of binding, or the stability of TFIID on the promoter, predicting that LSF would not affect the template commitment step. Rather, kinetic analyses demonstrate that LSF reduces the lag in the rate of initiation complex formation attributable to the slow addition of TFIIB and suggest that LSF increases the rate of association of TFIIB with the committed template. In addition, LSF increases the total number of transcription complexes in long term assays, which is also consistent with LSF increasing the rate of association of TFIIB, where TFIIB is not saturating. These results indicate a mechanism for the activation of the initiation of RNA polymerase II transcription by one upstream activating protein, LSF. This mechanism may also be applicable to other activators that function in cases where limiting concentrations of TFIIB in the cell dictate slow binding of TFIIB.

Base Sequence↗

PubMed1992

In vitro initiation of transcription by RNA polymerase II on in vivo-assembled chromatin templates.

We have studied the initiation of transcription in vitro by RNA polymerase II on simian virus 40 (SV40) minichromosomal templates isolated from infected cells. The efficiency and pattern of transcription from the chromatin templates were compared with those from viral DNA templates by using two in vitro transcription systems, either HeLa whole-cell extract or basal transcription factors, RNA polymerase II, and one of two SV40 promoter-binding transcription factors, LSF and Sp1. Dramatic increases in numbers of transcripts upon addition of transcription extract and different patterns of usage of the multiple SV40 initiation sites upon addition of Sp1 versus LSF strongly suggested that transcripts were being initiated from the minichromosomal templates in vitro. That the majority of transcripts from the minichromosomes were due to initiation de novo was demonstrated by the efficient transcription observed in the presence of alpha-amanitin, which inhibited minichromosome-associated RNA polymerase II, and an alpha-amanitin-resistant RNA polymerase II, which initiated transcription in vitro. The pattern of transcription from the SV40 late and early promoters on the minichromosomal templates was similar to the in vivo pattern of transcription during the late stages of viral infection and was distinct from the pattern of transcription generated from viral DNA in vitro. In particular, the late promoter of the minichromosomal templates was transcribed with high efficiency, similar to viral DNA templates, while the early-early promoter of the minichromosomal templates was inhibited 10- to 15-fold. Finally, the number of minichromosomes competent to initiate transcription in vitro exceeded the amount actively being transcribed in vivo.

Adolescent↗

PubMed1991

Salpingitis isthmica nodosa in female infertility and tubal diseases.

Salpingitis isthmica nodosa (SIN) is a condition of nodular thickening of the proximal Fallopian tube. The purpose of this study was to investigate the occurrence, distribution and frequency of SIN in Danish women salpingectomized because of tubal pregnancy or salpingitis and to correlate SIN with infertility, pregnancies, outcome of pregnancies, births, pelvic inflammatory disease and salpingitis. Sections from the isthmus were present in the specimens from 223 tubes from 193 patients and were analysed by the same pathologist. Originally, SIN was found in 12 patients but on re-examination, it was found in 24 patients. Ten women with SIN were bilaterally salpingectomized. Only one woman had SIN in both tubes. Women with SIN gave birth to as many children as women without SIN. After SIN had been diagnosed, no children were born, but this was not statistically different from the frequency of births in the non-SIN group after salpingectomy. Women with SIN had histological signs of salpingitis more often than women without SIN, but SIN complicated with salpingitis did not influence the number of children or tubal pregnancies. Women with SIN had a greater risk of two or more tubal pregnancies than women without SIN.

Adult↗

PubMed1991

trans activation of the simian virus 40 late promoter by large T antigen requires binding sites for the cellular transcription factor TEF-1.

Simian virus 40 (SV40) T antigen stimulates the level of transcription from several RNA polymerase II promoters, including the SV40 late promoter. The mechanism of trans activation appears to be indirect since binding of T antigen to specific DNA sequences is not required. However, specific promoter elements that respond to T antigen have not previously been defined. We identified DNA sequences from the SV40 late promoter whose ability to stimulate transcription is induced by the expression of T antigen. In particular, the Sph I + II motifs of the SV40 enhancer can confer T-antigen inducibility to the normally uninducible herpes simplex virus thymidine kinase gene promoter when multiple copies of the sequence are inserted 5' of the transcription initiation site and TATA sequence. Binding sites for the cellular transcription factor TEF-1 and octamer binding proteins are contained within the Sph I + II motifs, as well as at other positions in the SV40 promoter. To study the role of individual protein-binding sites in trans activation by T antigen, mutations were constructed in various TEF-1 and octamer protein-binding sites of the SV40 late promoter. These mutations did not significantly affect basal promoter activity. However, mutation of all three TEF-1 sites prevented detectable activation by T antigen. DNase I footprinting of the mutated promoters with purified proteins demonstrated that inducibility by T antigen correlated with binding affinity of TEF-1 for the DNA and not with binding affinity of an octamer binding protein.

Animals↗

PubMed1976

Formation of lipid-linked sugar compounds in Halobacterium salinarium. Presumed intermediates in glycoprotein synthesis.

The ability of bacitracin to inhibit the growth of Halobacterium salinarium suggested that glycosylation of the major envelope component, a high molecular weight glycoprotein, might occur via a pathway involving lipid intermediates. This report demonstrates that the cells have enzymatic activities for formation of lipid-linked sugar compounds having the expected properties of such intermediates. Whole cell homogenate catalyzed the transfer of sugar from UDP-glucose, GDP-mannose, and UDP-N-acetyglucosamine to endogenous lipid acceptors. Two lipid products were formed from UDP-glucose, two from GDP-mannose, and one from UDP-N-acetylglucosamine. Characterization of the partially purified lipids by ion exchange chromatography, thin layer chromatography, and mild acid and base hydrolysis showed the major product in each case to have the properties expected for polyisoprenyl phosphoglucose, polyisoprenyl phosphomannose, and polyisoprenyl pyrophospho-N-acetylglucosamine. Estimates of chain length by thin layer chromatography indicate that the lipid has 11 to 12 isoprene identity as a C55-60-polyisoprenyl pyrophospho-N-acetylglucosamine. The N-acetylglucosamine transferase, present in cell envelope preparations, was partially characterized. The enzyme was found to be extremely halophilic, specifically requiring a high concentration of KCl. Optimum activity was obtained at 4 m KCl and partial substitution of K+ by Na+ resulted in a decrease in activity.

Cell Membrane↗