C/EBPß deficiency enhances the keratinocyte innate immune response to direct activators of cytosolic pattern recognition receptors.

The skin is the first line of defense to cutaneous microbes and viruses, and epidermal keratinocytes play a critical role in preventing infection by viruses and pathogens through activation of the type I interferon (IFN) response. Using RNAseq analysis, here we report that the conditional deletion of C/EBPß transcription factor in mouse epidermis (CKOß mice) resulted in the upregulation of IFNß and numerous keratinocyte interferon-stimulated genes (ISGs). The expression of cytosolic pattern recognition receptors (cPRRs), that recognize viral RNA and DNA, were significantly increased, and enriched in the RNAseq data set. cPRRs stimulate a type I IFN response that can trigger cell death to eliminate infected cells. To determine if the observed increases in cPRRs had functional consequences, we transfected CKOß primary keratinocytes with the pathogen and viral mimics poly(I:C) (dsRNA) or poly(dA:dT) (synthetic B-DNA) that directly activate PRRs. Transfected CKOß primary keratinocytes displayed an amplified type I IFN response which was accompanied by increased activation of IRF3, enhanced ISG expression, enhanced activation of caspase-8, caspase-3 and increased apoptosis. Our results identify C/EBPß as a critical repressor of the keratinocyte type I IFN response, and demonstrates that the loss of C/EBPß primes keratinocytes to the activation of cytosolic PRRs by pathogen RNA and DNA to induce cell death mediated by caspase-8 and caspase-3.

Plasma 1-carbon metabolites and academic achievement in 15-yr-old adolescents.

Academic achievement in adolescents is correlated with 1-carbon metabolism (1-CM), as folate intake is positively related and total plasma homocysteine (tHcy) negatively related to academic success. Because another 1-CM nutrient, choline is essential for fetal neurocognitive development, we hypothesized that choline and betaine could also be positively related to academic achievement in adolescents. In a sample of 15-yr-old children (n= 324), we measured plasma concentrations of homocysteine, choline, and betaine and genotyped them for 2 polymorphisms with effects on 1-CM, methylenetetrahydrofolate reductase (MTHFR) 677C>T, rs1801133, and phosphatidylethanolamineN-methyltransferase (PEMT), rs12325817 (G>C). The sum of school grades in 17 major subjects was used as an outcome measure for academic achievement. Lifestyle and family socioeconomic status (SES) data were obtained from questionnaires. Plasma choline was significantly and positively associated with academic achievement independent of SES factors (paternal education and income, maternal education and income, smoking, school) and of folate intake (P= 0.009,R(2)= 0.285). With the addition of thePEMTrs12325817 polymorphism, the association value was only marginally changed. Plasma betaine concentration, tHcy, and theMTHFR677C>T polymorphism did not affect academic achievement in any tested model involving choline. Dietary intake of choline is marginal in many adolescents and may be a public health concern.-Nilsson, T. K., Hurtig-Wennlöf, A., Sjöström, M., Herrmann, W., Obeid, R., Owen, J. R., Zeisel, S. Plasma 1-carbon metabolites and academic achievement in 15-yr-old adolescents.

Induction of expression of a 14-3-3 gene in response to copper exposure in the marine alga, Fucus vesiculosus.

The macro-alga Fucus vesiculosus has a broad global and estuarine distribution and exhibits exceptional resistance to toxic metals, the molecular basis of which is poorly understood. To address this issue a cDNA library was constructed from an environmental isolate of F. vesiculosus growing in an area with chronic copper pollution. Characterisation of this library led to the identification of a cDNA encoding a protein known to be synthesised in response to toxicity, a full length 14-3-3 exhibiting a 71% identity to human/mouse epsilon isoform, 70-71% identity to yeast BMH1/2 and 95 and 71% identity to the Ectocarpus siliculosus 14-3-3 isoforms 1 and 2 respectively. Preliminary characterisation of the expression profile of the 14-3-3 indicated concentration- and time-dependent inductions on acute exposure of F. vesiculosus of copper (3-30 µg/l). Higher concentrations of copper (≥150 µg/l) did not elicit significant induction of the 14-3-3 gene compared with the control even though levels of both intracellular copper and the expression of a cytosolic metal chaperone, metallothionein, continued to rise. Analysis of gene expression within environmental isolates demonstrated up-regulation of the 14-3-3 gene associated with the known copper pollution gradient. Here we report for the first time, identification of a gene encoding a putative 14-3-3 protein in a multicellular alga and provide preliminary evidence to link the induction of this 14-3-3 gene to copper exposure in this alga. Interestingly, the threshold exposure profile may be associated with a decrease in the organism's ability to control copper influx so that it perceives copper as a toxic response.

Role of benzo[alpha]pyrene in generation of clustered DNA damage in human breast tissue.

Complex DNA damage may manifest in double-strand breaks (DSBs) and non-DSB, bistranded, oxidatively induced clustered DNA lesions (OCDLs). Although the carcinogen benzo[alpha]pyrene (B[alpha]P) has been shown to induce chromosomal aberrations and transformation of mammary cells, it is not known whether this compound engenders clustered DNA damage. Normal primary breast tissue-derived cells were treated with B[alpha]P, and the levels of DNA lesions, chromosomal aberrations, total antioxidant capacity (TAC), and reactive oxygen species (ROS) were determined. DNA from cells treated with 2 and 8 microM B[alpha]P exhibited increases of 3- and 4-fold in APE1 (p<0.001), 11- and 19-fold in Endo III (p<0.001), and 8- and 15-fold in hOGG1 (p<0.001) OCDLs, respectively, compared to the 0 microM B[alpha]P-treated (control) group. Mammary cells treated with 8 microM B[alpha]P produced 0.12 aberrations per cell (p<0.05) and there was a strong positive correlation (r=0.91) between the levels of OCDLs and those of chromosomal aberrations. Finally, TAC was decreased by 25% (p<0.02), whereas ROS production increased by 2-fold (p<0.02) in cells treated with 8 microM B[alpha]P compared to the control group. In conclusion, oxidatively induced clustered DNA damage mediated through differential expression of APE1, reduced TAC, and increased ROS may play a significant role in the chemically induced transformation of normal primary mammary cells.