Ultraviolet light-induced mutation in UV-resistant, thermosensitive derivatives of lexA-strains of Escherichia coli K-12

It was shown previously that a major class of UV-resistant derivatives of lexA- strains of E. coli K-12 is defective in cell division at 42.5 degrees. The thermosensitive mutations, judging by genetic mapping and complementation tests, are believed to be intragenic suppressor mutations that lower the activity of the diffusible product that results in the LexA- phenotype (Mount et al., 1973). Several thermosensitive derivatives have been characterized in regard to their susceptibility to mutation induction by UV at the permissive growth temperature (30 degrees). Although the strains tested are approximately as resistant to UV as lexA+ strains, they showed a level of mutation induction that was considerably lower. By means of genetic complementation tests it was demonstrated that the low levels of UV mutagenesis in lexA- strains and their thermosensitive derivatives result from the synthesis of a diffusible product. One possible interpretation of these results is that a diffusible product in lexA- strains prevents the induction of error-prone repair. Altering the activity of this product by tsl mutations can lead to increased, but not normal, levels of error-prone repair.     

Two motifs with different function regulate the anterograde transport of the adiponectin receptor 1

The anterograde trafficking of GPCR has been described as a tightly controlled process involving specific amino acid sequences that mediate the receptor transport. In this study, we investigated whether the cell surface delivery of the adiponectin receptor 1, a newly identified class of heptahelix receptors different from G protein-coupled receptors, is regulated. Sequential N-terminal deletion revealed that the export of the AdipoR1 from the endoplasmic reticulum (ER) is controlled by distinct parts of the receptor N-terminus. Strong evidence is provided that the ER exit is mediated by two specific sequences, a F(X)(3)F(X)(3)F and a D(X)(3)LL motif. Disruption of these motifs led to a substantial accumulation of the AdipoR1 in the ER. Mutation of similar motifs in the AdipoR1 C-terminus did not result in aberrant receptor localization, suggesting that these motifs are sequence and position specific to the AdipoR1 N-terminus. Further analysis of the regulation mechanism identified an interaction with the chaperone BiP and additionally, strong evidence is provided that both motifs exert different biological function in the AdipoR1 ER export. In conclusion, our data demonstrate that the receptor transport shares similar ER exit motifs although AdipoR are structurally different from GPCR. However, since even two specific sequences are identified, the anterograde trafficking of the AdipoR1 seems to be regulated in a more complex manner.     

Analysis of the enantiomers of citalopram and its demethylated metabolites using chiral liquid chromatography

A procedure using a chirobiotic V column is presented which allows separation of the enantiomers of citalopram and its two N-demethylated metabolites, and of the internal standard, alprenolol, in human plasma. Citalopram, demethylcitalopram and didemethylcitalopram, as well as the internal standard, were recovered from plasma by liquid–liquid extraction. The limits of quantification were found to be 5 ng/ml for each enantiomer of citalopram and demethylcitalopram, and 7.5 ng/ml for each enantiomer of didemethylcitalopram. Inter- and intra-day coefficients of variation varied from 2.4% to 8.6% for S- and R-citalopram, from 2.9% to 7.4% for S- and R-demethylcitalopram, and from 5.6% to 12.4% for S- and R-didemethylcitalopram. No interference was observed from endogenous compounds following the extraction of plasma samples from 10 different patients treated with citalopram. This method allows accurate quantification for each enantiomer and is, therefore, well suited for pharmacokinetic and drug interaction investigations. The presented method replaces a previously described highly sensitive and selective high-performance liquid chromatography procedure using an acetylated β-cyclobond column which, because of manufactural problems, is no longer usable for the separation of the enantiomers of citalopram and its demethylated metabolites.     

Protein kinase CK2 interacts with adiponectin receptor 1 and participates in adiponectin signaling

Adiponectin is an adipokine with anti-atherogenic, anti-diabetic and insulin sensitizing properties. Its effects on energy homeostasis, glucose and lipid metabolism are mediated by two ubiquitously expressed seven-transmembrane receptors, AdipoR1 and -R2. With the exception of APPL1 and RACK1, no intracellular binding partners of adiponectin receptors are reported and thus signaling pathways downstream of these receptors remain largely unknown. To incorporate adiponectins protective potential in drug development it is essential to understand adiponectin signaling cascades in detail. A yeast two-hybrid approach employing AdipoR1s cytoplasmatic N-terminus led to the identification of the regulatory subunit of protein kinase CK2. We confirmed the interaction in co-immunoprecipitation, ELISA experiments and co-localization analysis in mammalian cells. Furthermore we could localize the interaction site in an N-terminal basic region close to the transmembrane domain. In adiponectin stimulation experiments of C2C12 mouse myotubes and MCF7 cells incorporating CK2 inhibitor 2-dimethylamino-4,5,6,7-tetrabromo-1H-benz-imidazole (DMAT) we found a modulator role of CK2 in adiponectin signaling. Accordingly we identified the regulatory subunit of protein kinase CK2 as a novel intracellular partner of AdipoR1 and have strong evidence of CK2 as an effector molecule in adiponectin signaling. Since CK2 is involved in signaling cascades of other adipokines and hormones, e.g. leptin and insulin, our findings suggest a possible key function in crosstalk between adiponectin and insulin signaling pathways and could provide further insight into the anti-diabetic effects of adiponectin.     

Risk of Ovarian Cancer and Inherited Variants in Relapse-Associated Genes

Background

We previously identified a panel of genes associated with outcome of ovarian cancer. The purpose of the current study was to assess whether variants in these genes correlated with ovarian cancer risk.

Methods and Findings

Women with and without invasive ovarian cancer (749 cases, 1,041 controls) were genotyped at 136 single nucleotide polymorphisms (SNPs) within 13 candidate genes. Risk was estimated for each SNP and for overall variation within each gene. At the gene-level, variation within MSL1 (male-specific lethal-1 homolog) was associated with risk of serous cancer (p = 0.03); haplotypes within PRPF31 (PRP31 pre-mRNA processing factor 31 homolog) were associated with risk of invasive disease (p = 0.03). MSL1 rs7211770 was associated with decreased risk of serous disease (OR 0.81, 95% CI 0.66–0.98; p = 0.03). SNPs in MFSD7, BTN3A3, ZNF200, PTPRS, and CCND1A were inversely associated with risk (p<0.05), and there was increased risk at HEXIM1 rs1053578 (p = 0.04, OR 1.40, 95% CI 1.02–1.91).

Conclusions

Tumor studies can reveal novel genes worthy of follow-up for cancer susceptibility. Here, we found that inherited markers in the gene encoding MSL1, part of a complex that modifies the histone H4, may decrease risk of invasive serous ovarian cancer.     

Properties of strains of Escherichia coli K12 carrying mutant lex-1 and uvrA6 alleles

Summary Strains with both uvrA6 and the lex-1 mutations are more sensitive to ultraviolet light (UV) than isogenic strains with only one of the mutations. The lex ^- uvrA^-double mutant has the same sensitivity to methyl-methane-sulfonate as the lex ^- uvrA⁺single mutant. UV-irradiated cultures of lex ^- uvrA⁺and lex ^- uvrA^-strains do not produce more streptomycinresistant mutants per survivor than unirradiated cultures. UV-irradiated cultures of a lex ⁺ uvrA^-strain produce large yields of mutants at both low (4 ergs/mm²) and high (25 ergs/mm²) doses of UV compared with the lex ⁺ uvrA⁺ strain which produce an intermediate yield of mutants at 25 ergs/mm², and a small yield at 4 ergs/mm², not significantly greater than unirradiated cultures. A dose of UV which does not induce mutations in strains with the lex-1 mutation produces only a small decrease in DNA synthesis in the lex ^- uvrA⁺strain. The results are interpreted to mean that the lex-1 mutation probably does not affect the same pathway of DNA repair as the uvrA ⁺product (i.e. excision of thymine dimers), and that the absence of UV-induced mutations in irradiated cultures of lex ^-strains is probably not due to a cessation of DNA replication.     

Analysis of the Distribution of Magnetic Fluid inside Tumors by a Giant Magnetoresistance Probe

Magnetic fluid hyperthermia (MFH) therapy uses the magnetic component of electromagnetic fields in the radiofrequency spectrum to couple energy to magnetic nanoparticles inside tumors. In MFH therapy, magnetic fluid is injected into tumors and an alternating current (AC) magnetic flux is applied to heat the magnetic fluid- filled tumor. If the temperature can be maintained at the therapeutic threshold of 42°C for 30 minutes or more, the tumor cells can be destroyed. Analyzing the distribution of the magnetic fluid injected into tumors prior to the heating step in MFH therapy is an essential criterion for homogenous heating of tumors, since a decision can then be taken on the strength and localization of the applied external AC magnetic flux density needed to destroy the tumor without affecting healthy cells. This paper proposes a methodology for analyzing the distribution of magnetic fluid in a tumor by a specifically designed giant magnetoresistance (GMR) probe prior to MFH heat treatment. Experimental results analyzing the distribution of magnetic fluid suggest that different magnetic fluid weight densities could be estimated inside a single tumor by the GMR probe.     

Giant clams in shallow reefs: UV-resistance mechanisms of Tridacninae in the Red Sea

The photosymbiosis of tropical giant clams (subfamily Tridacninae) with unicellular algae (Symbiodiniaceae) restricts their distribution to the sunlit, shallow waters of the euphotic zone where organisms are additionally exposed to potentially damaging levels of solar UV radiation. Metabolic and physiological responses of Red Sea Tridacna maxima clams, including net calcification and primary production, as well as valvometry (i.e., shell gaping behavior) were assessed when exposed to simulated high radiation levels received at 3 and 5 m underwater. The two levels of radiation included exposure treatments to photosynthetically active radiation (PAR; 400–700 nm) alone and to both, PAR and ultraviolet-B radiation (UV-B; 280–315 nm). The valvometry data obtained using flexible magnetic sensors indicated that specimens under PAR + UV-B exposure significantly reduced the proportion of their exposed mantle area, a potential photo-protective mechanism which, however, reduces the overall amount of PAR received by the algal symbionts. Consequently, specimens under PAR + UV-B displayed a slight, although non-significant, reduction in primary production rates but no signs of additional oxidative stress, changes in symbiont densities, chlorophyll content, or levels of mycosporine-like amino acids. Net calcification rates of T. maxima were not affected by exposure to UV-B; however, calcification was positively correlated with incident PAR levels. UV-B exposure changes the valvometry, reducing the exposed mantle area which consequently diminishes the available PAR for the photosymbionts. Still, T. maxima maintains high rates of primary production and net calcification, even under high levels of UV-B. This provides experimental support for a recently described, effective UV-defensive mechanism in Tridacninae, in which the photonic cooperation of the associated algal symbionts and giant clam iridocytes is assumed to establish optimal conditions for the photosynthetic performance of the clams’ symbionts.