Gain of function by phosphorylation in Presenilin 1-mediated regulation of insulin signaling

During pregnancy, activation of the maternal immune system results in inflammation in the foetal nervous system. The causative agents are pro-inflammatory cytokines like interleukin-1β (IL-1β), produced by the foetus. In this study, we examine the effect of IL-1β on the proliferation and differentiation of neural progenitor cells (NPCs) to better understand its potential effects on the developing brain. We find that the IL-1β receptor (IL-1R1) is expressed in the ventral mesencephalon of the developing brain. Furthermore, IL-1R1 is expressed on Nestin-positive, Sox-2-positive NPCs. IL-1β treatment reduced the numbers of proliferating NPCs, an effect prevented by the IL-1R1 receptor antagonist. LDH and MTT assays, and western blot analysis for cleaved caspase 3 and poly(ADP-ribose) polymerase, confirmed that this was not due to an increase in cell death but rather an induction of differentiation. To further study the effects of IL-1β on cell fate determination, we differentiated NPCs in the presence and absence of IL-1β. Il-1β promoted gliogenesis and inhibited neurogenesis, an effect that required p38-MAPK kinase signalling. In summary, these data show that exposure of NPCs to IL-1β affects their development. This necessitates an examination of the consequences that maternal immune system activation during pregnancy has on the cellular architecture of the developing brain.     

Branched-chain amino acid-enriched nutrients stimulate antioxidant DNA repair in a rat model of liver injury induced by carbon tetrachloride

Oxidative stress (OS) plays an important role in the progression of chronic liver disease including organ injury and hypoalbuminemia. Long-term oral supplementation with branched-chain amino acids (BCAAs) can inhibit liver dysfunction but their role in the prevention of liver fibrosis and injury to the liver is unclear. The aim of this study was to assess how BCAAs preserve liver function from OS. To investigate how BCAAs specifically prevent OS, we evaluated the effect of oral supplementation with BCAAs on OS using a rat liver cirrhosis model. Liver cirrhosis was induced in ten male Sprague–Dawley rats by administering carbon tetrachloride for 12?weeks. Five of the ten carbon tetrachloride-treated rats were assigned to a control group and five to a BCAA group. BCAA-supplementation significantly preserved plasma albumin concentrations and significantly inhibited the occurrence of organ injury as determined by blood chemistry analysis. Hepatic expression of OGG1 mRNA was increased in the BCAA group compared to the control group. In the BCAA group, increased hepatic levels of OGG1 protein were found by western blot. On the other hand, the number of 8-OHdG-positive cells was significantly higher in liver sections taken 1?month after carbon tetrachloride treatment. Furthermore, OGG1-positive cells were significantly increased in the hepatocytes around the central vein. BCAA was found to reduce OS, which could possibly lead to a decrease in the occurrence of hypoalbuminemia and organ injury. Our results indicate that BCAA-enriched nutrients stimulate antioxidant DNA repair in a rat model of liver injury induced by carbon tetrachloride.     

Effect of thiazole orange doubly labeled thymidine on DNA duplex formation

Nucleic acid oligonucleotides are widely used in hybridization experiments for specific detection of complementary nucleic acid sequences. For design and application of oligonucleotides, an understanding of their thermodynamic properties is essential. Recently, exciton-controlled hybridization-sensitive fluorescent oligonucleotides (ECHOs) were developed as uniquely labeled DNA oligomers containing commonly one thymidine having two covalently linked thiazole orange dye moieties. The fluorescent signal of an ECHO is strictly hybridization-controlled, where the dye moieties have to intercalate into double-stranded DNA for signal generation. Here we analyzed the hybridization thermodynamics of ECHO/DNA duplexes, and thermodynamic parameters were obtained from melting curves of 64 ECHO/DNA duplexes measured by ultraviolet absorbance and fluorescence. Both methods demonstrated a substantial increase in duplex stability (ΔΔG°(37) ～ -2.6 ± 0.7 kcal mol(-1)) compared to that of DNA/DNA duplexes of the same sequence. With the exception of T·G mismatches, this increased stability was mostly unaffected by other mismatches in the position opposite the labeled nucleotide. A nearest neighbor model was constructed for predicting thermodynamic parameters for duplex stability. Evaluation of the nearest neighbor parameters by cross validation tests showed higher predictive reliability for the fluorescence-based than the absorbance-based parameters. Using our experimental data, a tool for predicting the thermodynamics of formation of ECHO/DNA duplexes was developed that is freely available at http://genome.gsc.riken.jp/echo/thermodynamics/ . It provides reliable thermodynamic data for using the unique features of ECHOs in fluorescence-based experiments.     

Control of potato aphids by the addition of barley strips in potato fields: a successful example of vegetation management

The densities of barley and potato aphids, their natural enemies and hyperparasitoids were assessed in three experimental potato fields as a case study to investigate the effectiveness of the addition of barley strips in potato fields for conservation biological control. These fields were located in a low plant-diversity landscape, but common aphid species and their natural enemies were present. The barley strips in the potato fields were found to support different species of aphids of potato, but these different sets of aphids shared a common set of natural enemies. The amount of time between peak aphid densities and peaks of their natural enemies' populations was shorter in the potato fields than in the barley strips. The levels of winged aphids in a potato monoculture field were significantly higher than those in a field with barley strips. The wingless and winged aphid populations in the field without barley strips was almost three times higher than in the fields with the barley strips, as measured at the peak aphid density. This result is one of few examples of the application of the conservation effect of greenhouse banker plants on outdoor crops.     

The ensemble of hetero-proteins in inorganic nanochannels

The assembly and proper alignment of two heterofluorescent proteins (sGFP and DsRed) in the mesoporous channels of ethanol-treated FSM6.2 (a folded-sheet mesoporous material with a pore diameter of 6.2 nm) was confirmed using a fluorescence resonance energy transfer (FRET) technique. The sGFP-DsRed-FSM6.2 conjugate showed a large decrease in the emission of donor (sGFP) fluorescence, indicating that the conjugate functions as an energy transfer system through the combination of the two heteroproteins, due to the successful encapsulation of the sGFP-DsRed pairs in the mesopores. Fluorescence spectral analysis demonstrated that the proteins were highly dispersed and homogeneously encapsulated in the mesopores of FSM6.2, even at high concentration, although they spontaneously aggregated and showed a red shift in solution at the concentration corresponding to that in the conjugate. Furthermore, an increase in the amount of sGFP and DsRed adsorbed to the pores of FSM6.2 led to a decrease in the distance between these proteins, resulting in enhancement of FRET efficiency.     

Identification of a second DNA binding site in the human Rad52 protein

Rad52 plays essential roles in homology-dependent double-strand break repair. Various studies have established the functions of Rad52 in Rad51-dependent and Rad51-independent repair processes. However, the precise molecular mechanisms of Rad52 in these processes remain unknown. In the present study we have identified a novel DNA binding site within Rad52 by a structure-based alanine scan mutagenesis. This site is closely aligned with the putative single-stranded DNA binding site determined previously. Mutations in this site impaired the ability of the Rad52-single-stranded DNA complex to form a ternary complex with double-stranded DNA and subsequently catalyze the formation of D-loops. We found that Rad52 introduces positive supercoils into double-stranded DNA and that the second DNA binding site is essential for this activity. Our findings suggest that Rad52 aligns two recombining DNA molecules within the first and second DNA binding sites to stimulate the homology search and strand invasion processes.     

Biosynthesis and secretion of salusin-alpha from human cells

Salusins originally identified using bioinformatics analyses have been shown to act on the cardiovascular and endocrine systems. Although the hypotensive activity of salusin-alpha is limited, it exerts a significant anti-atherosclerotic effect via suppression of foam cell formation in human monocyte-derived macrophages by down-regulating acyl-CoA:cholesterol acyltransferase-1. Furthermore, serum salusin-alpha levels show a close negative correlation with the severity of atherosclerotic diseases. However, biosynthesis and secretion of salusin-alpha peptide from cultured mammalian cells have not been demonstrated to date. We examined the expression, synthesis and release of salusin-alpha in human-derived cell lines. Preprosalusin mRNA and protein were detected ubiquitously in all cells tested, whereas the processing of preprosalusin into salusin-alpha peptide is dependent upon each cell type. Immunohistochemical study revealed the most abundant salusin-alpha-like immunoreactivity to be present in HeLa cells which released salusin-alpha-like immunoreactivity into the culture supernatant. Analysis of extracted conditioned media from HeLa cells by reverse-phase high performance liquid chromatography coupled with radioimmunoassay detection revealed a single immunoreactive component that co-eluted with authentic salusin-alpha. These results present the first evidence that salusin-alpha is biosynthesized and released from human-derived cells.     

Lim15/Dmc1 enhances DNA topoisomerase II catenation activity independent of sequence homology

We reported previously that Coprinus cinereus Lim15/Dmc1 (CcLim15), a meiosis-specific recA-like protein, could specifically activate C. cinereus DNA topoisomerase II (CcTopII). In particular, it enhanced the catenation activity of CcTopII in vitro at the meiotic prophase stage (Iwabata K, Koshiyama A, Yamaguchi T, Sugawara H, Hamada NF, Namekawa HS, Ishii S, Ishizaki T, Chiku H, Nara T, Sakaguchi K, Nucleic Acids Res, 33:5809–5818, 2005). In this study, the interaction between CcTopII and CcLim15, especially during catenation, was investigated in detail using atomic force microscopy. We demonstrated earlier that CcLim15 enhanced the catenation activity of CcTopII in a dose-dependent manner. When using two different-sized plasmid rings (5.4 and 3 kbp), which did not have any homologous sequence regions, equal proportions of homologous and heterologous catenanes were produced, suggesting that CcLim15 causes an increase in catenation activity irrespective of the presence of homologous sequences between the rings. We also showed that CcLim15 works as a DNA-condensing agent. Therefore, we speculate that CcLim15 may work as a DNA-condensing factor specific to the zygotene event and that CcTopII is likely to resolve tangles when the chromosomes initiate pairing at multiple sites by CcLim15.