Detection of human blood by cloth-based enzyme immunoassay of immunoglobulin G

A specific and sensitive assay for the detection of human blood was developed using polyester cloth coated with goat anti-human IgG antibody to capture human IgG, an abundant and stable protein in blood. The captured IgG was detected by the reaction between goat anti-human IgG antibody-peroxidase conjugate and a chromogenic peroxidase substrate. Because the assay is simple and rapid, and permits simultaneous analysis of multiple samples, it has the potential to be used as a forensic test for human blood.     

Kinetics of bacteriophage lambda repressor synthesis directed by the PRE promoter: influence of temperature, multiplicity of infection, and mutation of PRM or the cro gene

Summary Expression of the P _RE (establishment) pathway for repressor synthesis is regulated both by phage-specific genetic elements and by physiological conditions. Here we describe the effects of temperature, multiplicity of infection, mutations in the cro gene, and a mutation in P _RM on P _RE-directed repressor synthesis. As Reichardt (1975a) has shown, repressor synthesis begins 5–15 min after infection by wildtype phage, and is shut off at 20–30 min after infection, depending on the temperature. At 43°, synthesis starts sooner, shuts off earlier, and leads to lower repressor levels than are attained at lower temperatures. Experiments with the temperature sensitive mutant crots20 demonstrate that, as had been shown previously in experiments at 30° and 37° C, cro protein is responsible for the shut-off of repressor synthesis at 43°. In addition to the effects of temperature, the kinetics of repressor synthesis are strongly affected by multiplicity of infection (moi). At mois greater than 10, repressor synthesis after infection by wildtype at 30° is dramatically inhibited. Unexpectedly, the P _RM mutation prm116, under certain conditions, can alleviate both cro-mediated shutoff and the inhibition of P _RE-directed repressor synthesis at high moi. These effects of prm116 are observed only at low temperature (30°–32° C) and at mois of about 6–10 or greater; they also appear to be cis-specific. Possible mechanisms for the effects of the prm116 mutation are discussed. Finally, these studies demonstrate that crots20, which was isolated as a temperature-sensitive lethal mutation in the cro gene (Herskowitz, unpublished), is temperature-sensitive with respect to the ability to shutoff P _RE-directed repressor synthesis; however, even at low temperature (30° C), the crots20 gene product is only partially active.     

Antioxidant actions of du-zhong (Eucommia ulmoides Oliv.) toward oxidative damage in biomolecules

This study aimed to investigate the antioxidant effect of water extracts of Du-zhong (WEDZ) on oxidative damage in biomolecules such as deoxyribose, DNA, and 2'-deoxyguanosine (2'-dG) as induced by Fenton reaction. The WEDZ used included leaves, raw cortex, and roasted cortex. All of the WEDZ inhibited the oxidation of deoxyribose induced by Fe(3+)-EDTA/H2O2/ascorbic acid in a concentration dependent manner. At a concentration of 1.14 mg/mL, the inhibitory effect of the extracts of leaves, roasted cortex, and raw cortex was 85.2%, 68.0% and 49.3%, respectively. The extract of leaves inhibited the strand-breaking of DNA induced by the Fenton reaction at concentrations of 5 and 10 micrograms/microL. This inhibitory effect was similar to mannitol whereas the extracts of raw cortex and roasted cortex had no inhibitory effect at all. WEDZ also inhibited the oxidation of 2'-dG to 8-OH-2'-dG induced by Fe(3+)-EDTA/H2O2/ascorbic acid. Gallic acid had a prooxidant effect, but trolox and mannitol had an antioxidant effect. The leaf extract had a marked inhibitory effect on Fenton reaction induced oxidative damage in biomolecules. The extract of roasted cortex exhibited modest inhibition while the extract of raw cortex had the least inhibitory effect on oxidative damage in biomolecules. This is in contrast to gallic acid in the same reaction system, whose higher reducing power and weaker chelating ability may contribute to its prooxidant effect. In the present study, leaf extract of Du-zhong had inhibitory effect on oxidative damage in biomolecules. Therefore, drinking of Du-zhong tea (leaf extract) over a long period of time may have anticancer potential.     

Autoregulation of tubulin expression is achieved through specific degradation of polysomal tubulin mRNAs

We have utilized protein synthesis inhibitors to investigate the autoregulatory mechanism that uses the concentration of unpolymerized tubulin subunits to specify tubulin mRNA content in animal cells. Puromycin and pactamycin, both of which remove RNAs from polysomes, completely unlink tubulin RNA content from the level of free subunits, whereas pretreatment of cells with cycloheximide, which traps mRNAs onto stalled polyribosomes, enhances the specific degradation of tubulin RNAs in response to increases in the subunit content. Moreover, in the absence of protein synthesis inhibitors, the tubulin RNAs that are lost from cells with elevated free tubulin subunit levels are those that are associated with polyribosomes. Further, beta-tubulin mRNAs encoding a truncated translation product of only 26 amino acids (and that cannot be polyribosomal) are not substrates for autoregulation. We conclude that autoregulation of tubulin synthesis is achieved by specifically altering the stability of tubulin RNAs that are bound to polyribosomes.     

Mechanistic control of carcinoembryonic antigen-related cell adhesion molecule-1 (CEACAM1) splice isoforms by the heterogeneous nuclear ribonuclear proteins hnRNP L, hnRNP A1, and hnRNP M

Carcinoembryonic antigen-related cell adhesion molecule-1 (CEACAM1) is expressed in a variety of cell types and is implicated in carcinogenesis. Alternative splicing of CEACAM1 pre-mRNA generates two cytoplasmic domain splice variants characterized by the inclusion (L-isoform) or exclusion (S-isoform) of exon 7. Here we show that the alternative splicing of CEACAM1 pre-mRNA is regulated by novel cis elements residing in exon 7. We report the presence of three exon regulatory elements that lead to the inclusion or exclusion of exon 7 CEACAM1 mRNA in ZR75 breast cancer cells. Heterologous splicing reporter assays demonstrated that the maintenance of authentic alternative splicing mechanisms were independent of the CEACAM1 intron sequence context. We show that forced expression of these exon regulatory elements could alter CEACAM1 splicing in HEK-293 cells. Using RNA affinity chromatography, three members of the heterogeneous nuclear ribonucleoprotein family (hnRNP L, hnRNP A1, and hnRNP M) were identified. RNA immunoprecipitation of hnRNP L and hnRNP A1 revealed a binding motif located central and 3' to exon 7, respectively. Depletion of hnRNP A1 or L by RNAi in HEK-293 cells promoted exon 7 inclusion, whereas overexpression led to exclusion of the variable exon. By contrast, overexpression of hnRNP M showed exon 7 inclusion and production of CEACAM1-L mRNA. Finally, stress-induced cytoplasmic accumulation of hnRNP A1 in MDA-MB-468 cells dynamically alters the CEACAM1-S:CEACAM1:L ratio in favor of the l-isoform. Thus, we have elucidated the molecular factors that control the mechanism of splice-site recognition in the alternative splicing regulation of CEACAM1.     

Tea Consumption and Risk of Head and Neck Cancer

Background

The current study evaluated the association between tea consumption and head and neck cancer (HNC) in Taiwan, where tea is a major agricultural product and a popular beverage.

Methods

Interviews regarding tea consumption (frequency, duration, and types) were conducted with 396 HNC cases and 413 controls. Unconditional logistic regression was performed to estimate the odds ratio (OR) and 95% confidence interval (CI) of HNC risk associated with tea drinking, adjusted for sex, age, education, cigarette smoking, betel quid chewing, and alcohol drinking.

Results

A reduced HNC risk associated with tea drinking (OR for every cup per day = 0.96, 95% CI: 0.93–0.99; OR for ≧5 cups per day = 0.60, 95% CI: 0.39–0.94) was observed. The association was especially significant for pharyngeal cancer (OR for every cup per day = 0.93, 95% CI: 0.88–0.98; OR for ≧5 cups per day = 0.32, 95% CI: 0.16–0.66). A significant inverse association between HNC and tea consumption was observed particularly for green tea.

Conclusions

This study suggests that tea drinking may reduce the risk of HNC. The anticancer property of tea, if proven, may offer a natural chemopreventive measure to reduce the occurrence of HNC.     

Histone deacetylase 4 interacts with 53BP1 to mediate the DNA damage response

Anumber of proteins are recruited to nuclear foci upon exposure to double-strand DNA damage, including 53BP1 and Rad51, but the precise role of these DNA damage-induced foci remain unclear. Here we show in a variety of human cell lines that histone deacetylase (HDAC) 4 is recruited to foci with kinetics similar to, and colocalizes with, 53BP1 after exposure to agents causing double-stranded DNA breaks. HDAC4 foci gradually disappeared in repair-proficient cells but persisted in repair-deficient cell lines or cells irradiated with a lethal dose, suggesting that resolution of HDAC4 foci is linked to repair. Silencing of HDAC4 via RNA interference surprisingly also decreased levels of 53BP1 protein, abrogated the DNA damage-induced G2 delay, and radiosensitized HeLa cells. Our combined results suggest that HDAC4 is a critical component of the DNA damage response pathway that acts through 53BP1 and perhaps contributes in maintaining the G2 cell cycle checkpoint.