GTP, an inhibitor of transglutaminases, is hydrolyzed by tissue-type transglutaminase (TGase 2) but not by epidermal-type transglutaminase (TGase 3)

Epidermal-type transglutaminase (TGase 3) is devoid of GTPase activity, but its TGase activity is inhibited by GTP as in the case of tissue-type TGase (TGase 2). In addition, the inhibition was not affected by the presence of higher concentrations of Ca ion. These results indicate that GTP interacts with TGase 3 in a manner different from its action on TGase 2.     

Structure and function of the fourth subunit (Dpb4p) of DNA polymerase epsilon in Saccharomyces cerevisiae

DNA polymerase (Pol) of Saccharomyces cerevisiae is purified as a complex of four polypeptides with molecular masses of >250, 80, 34 (and 31) and 29 kDa as determined by SDS–PAGE. The genes POL2, DPB2 and DPB3, encoding the catalytic Pol2p, the second (Dpb2p) and the third largest subunits (Dpb3p) of the complex, respectively, were previously cloned and characterised. This paper reports the partial amino acid sequence of the fourth subunit (Dpb4p) of Pol. This protein sequence matches parts of the predicted amino acid sequence from the YDR121w open reading frame on S.cerevisiae chromosome IV. Thus, YDR121w was renamed DPB4. A deletion mutant of DPB4 (Δdpb4) is not lethal, but chromosomal DNA replication is slightly disturbed in this mutant. A double mutant haploid strain carrying the Δdpb4 deletion and either pol2-11 or dpb11-1 is lethal at all temperatures tested. Furthermore, the restrictive temperature of double mutants carrying Δdpb4 and dpb2-1, rad53-1 or rad53-21 is lower than in the corresponding single mutants. These results strongly suggest that Dpb4p plays an important role in maintaining the complex structure of Pol in S.cerevisiae, even if it is not essential for cell growth. Structural homologues of DPB4 are present in other eukaryotic genomes, suggesting that the complex structure of S.cerevisiae Pol is conserved in eukaryotes.     

8-Methyladenosine-substituted analogues of 2-5A: synthesis and their biological activities.

8-Methyladenosine-substituted analogues of 2-5A, p5'A2'p5'A2'p5'(me8A), p5'A2'p5'(me8A)2'p5'(me8A), p5'(me8A)2'p5'(me8A)2'p5'(me8A), and p5'(me8A) 2'p5'A2'p5'A, were prepared via a modification of a lead ion-catalyzed ligation reaction. These 2-5A monophosphates were converted into the corresponding 5'-triphosphates. Substitution of an 8-methyladenosine residue at the third position (2'-terminus) of the oligonucleotides increased the stability to snake venom phosphodiesterase digestion. Both binding and activation of mouse liver 2-5A dependent ribonuclease (RNase L) by the various 8-methyladenosine-substituted 2-5A analogues were examined. Among the 8-methyladenosine-substituted trimer analogues, the analogues with 8-methyladenosine residing in the 2'-terminal position showed the strongest binding affinity and were several times more effective than 2-5A itself as an inhibitor of translation.     

Correlation between limitation of growth ofPachysolen tannophilus on D-xylose with the formation of ethanol and other products

The formation of ethanol, xylitol, ribitol, arabitol and acetic acid from D-xylose byPachysolen tannophilus correlated with the limitation of growth. The correlation was consistent with these products being secondary metabolites.Issued as NRCC Publication Number 24259.     

The interactive effect of the cholinergic system and acute ovarian suppression on the brain: an fMRI study

Recent evidence suggests that loss of ovarian function following ovariectomy is a risk factor for Alzheimer's disease (AD); however, the biological basis of this risk remains poorly understood. We carried out an fMRI study into the interaction between loss of ovarian function (after Gonadotropin Hormone Releasing Hormone agonist (GnRHa) treatment) and scopolamine (a cholinergic antagonist used to model the memory decline associated with aging and AD). Behaviorally, cholinergic depletion produced a deficit in verbal recognition performance in both GnRHa-treated women and wait list controls, but only GnRHa-treated women made more false positive errors with cholinergic depletion. Similarly, cholinergic depletion produced a decrease in activation in the left inferior frontal gyrus (LIFG; Brodmann area 45) - a brain region implicated in retrieving word meaning - in both groups, and activation in this area was further reduced following GnRHa treatment. These findings suggest biological mechanisms through which ovarian hormone suppression may interact with the cholinergic system and the LIFG. Furthermore, this interaction may provide a useful model to help explain reports of increased risk for cognitive decline and AD in women following ovariectomy.     

Determination of antifungal activities in serum samples from mice treated with different antifungal drugs allows detection of an active metabolite of itraconazole

To establish an in vitro method of predicting in vivo efficacy of antifungal drugs against Candida albicans and Aspergillus fumigatus, the antifungal activities of fluconazole, itraconazole, and amphotericin B were determined in mouse serum. The minimum inhibitory concentration (MIC) of each drug was measured using mouse serum as a diluent. For C. albicans, the assay endpoint of azoles was defined as inhibition of mycelial extension (mMIC) and for A. fumigatus, as no growth (MIC). The MICs of amphotericin B for both pathogens were defined as the MIC at which no mycelial growth occurred. Serum MIC or mMIC determinations were then used to estimate the concentration of the drugs in serum of mice treated with antifungal drugs by multiplying the antifungal titer of the serum samples by the serum (m)MIC. The serum drug concentrations were also determined by HPLC. The serum concentrations estimated microbiologically showed good agreement with those determined by HPLC, except for itraconazole. Analysis of the serum samples from itraconazole-treated mice by a sensitive bioautography revealed the presence of additional spots, not seen in control samples of itraconazole. The bioautography assay demonstrated that the additional material detected in serum from mice treated with itraconazole was an active metabolite of itraconazole. The data showed that the apparent reduction in the itraconazole serum concentration as determined by HPLC was the result of the formation of an active metabolite, and that the use of a microbiological method to measure serum concentrations of drugs can provide a method for prediction of in vivo efficacy of antifungal drugs.     

MDA-MB-231 produces ATP-mediated ICAM-1-dependent facilitation of the attachment of carcinoma cells to human lymphatic endothelial cells

We examined the effects of supernatants of culture media of MDA-MB-231 and MCF-7 cells on the expression of adhesion molecules on human lymphatic endothelial cells (LECs) and evaluated whether the overexpression of adhesion molecules facilitated the attachment of carcinoma cells to LECs. The 48-h stimulation of MDA-MB-231, but not MCF-7, supernatant produced a significant expression of ICAM-1 on human LECs but little or no expression of E-selectin. Chemical treatment with dialyzed substances of <1,000 molecular weight (MW) caused a complete reduction of the supernatant-mediated response. In contrast, pretreatment with heating, digestion with protease, or chemical treatment with dialyzed substances of <500 MW produced no significant effect on the supernatant-mediated response. ATP (10(-7) M) caused overexpression of ICAM-1 on human LECs similar to that produced by the supernatant of MDA-MB-231. The ATP- and MDA-MB-231 supernatant-mediated responses were significantly reduced by treatment with 10(-6) M suramin (a purinergic P2X and P2Y receptor antagonist). In attachment assays, 10(-7) M ATP or MDA-MB-231 supernatant produced a significant increase in the attachment of carcinoma cells to human LECs. The treatment with 10(-6) M suramin caused a significant reduction of ATP- and supernatant-mediated facilitation of the attachment responses. Additional treatment with anti-ICAM-1 antibody also caused a significant reduction of ATP- and supernatant-mediated facilitation of the attachment responses. The experimental findings suggest that MDA-MB-231 may release or leak ATP, which produces the overexpression of ICAM-1 on human LECs through activation of purinergic P2X and/or P2Y receptors and then facilitates ICAM-1-mediated attachment of carcinoma cells to LECs.     

Pyridoxine stimulates filaggrin production in human epidermal keratinocytes

Molecular Biology Reports - Pyridoxine (PN), one of the vitamers of vitamin B6, plays an important role in the maintenance of epidermal function and is used to treat acne and rough skin. Clinical...     

The Arabidopsis pex12 and pex13 mutants are defective in both PTS1- and PTS2-dependent protein transport to peroxisomes

Peroxisome biogenesis requires various complex processes including organelle division, enlargement and protein transport. We have been studying a number of Arabidopsis apm mutants that display aberrant peroxisome morphology. Two of these mutants, apm2 and apm4, showed green fluorescent protein fluorescence in the cytosol as well as in peroxisomes, indicating a decrease of efficiency of peroxisome targeting signal 1 (PTS1)-dependent protein transport to peroxisomes. Interestingly, both mutants were defective in PTS2-dependent protein transport. Plant growth was more inhibited in apm4 than apm2 mutants, apparently because protein transport was more severely decreased in apm4 than in apm2 mutants. APM2 and APM4 were found to encode proteins homologous to the peroxins PEX13 and PEX12, respectively, which are thought to be involved in transporting matrix proteins into peroxisomes in yeasts and mammals. We show that APM2/PEX13 and APM4/PEX12 are localized on peroxisomal membranes, and that APM2/PEX13 interacts with PEX7, a cytosolic PTS2 receptor. Additionally, a PTS1 receptor, PEX5, was found to stall on peroxisomal membranes in both mutants, suggesting that PEX12 and PEX13 are components that are involved in protein transport on peroxisomal membranes in higher plants. Proteins homologous to PEX12 and PEX13 have previously been found in Arabidopsis but it is not known whether they are involved in protein transport to peroxisomes. Our findings reveal that APM2/PEX13 and APM4/PEX12 are responsible for matrix protein import to peroxisomes in planta.     

Shell productivity of the large benthic foraminifer Baculogypsina sphaerulata,based on the population dynamics in a tropical reef environment

Carbonate production by large benthic foraminifers is sometimes comparable to that of corals and coralline algae, and contributes to sedimentation on reef islands and beaches in the tropical Pacific. Population dynamic data, such as population density and size structure (size-frequency distribution), are vital for an accurate estimation of shell production of foraminifers. However, previous production estimates in tropical environments were based on a limited sampling period with no consideration of seasonality. In addition, no comparisons were made of various estimation methods to determine more accurate estimates. Here we present the annual gross shell production rate of Baculogypsina sphaerulata, estimated based on population dynamics studied over a 2-yr period on an ocean reef flat of Funafuti Atoll (Tuvalu, tropical South Pacific). The population density of B. sphaerulata increased from January to March, when northwest winds predominated and the study site was on the leeward side of reef islands, compared to other seasons when southeast trade winds predominated and the study site was on the windward side. This result suggested that wind-driven flows controlled the population density at the study site. The B. sphaerulata population had a relatively stationary size-frequency distribution throughout the study period, indicating no definite intensive reproductive period in the tropical population. Four methods were applied to estimate the annual gross shell production rates of B. sphaerulata. The production rates estimated by three of the four methods (using monthly biomass, life tables and growth increment rates) were in the order of hundreds of g CaCO₃ m⁻² yr⁻¹ or cm⁻³ m⁻² yr⁻¹, and the simple method using turnover rates overestimated the values. This study suggests that seasonal surveys should be undertaken of population density and size structure as these can produce more accurate estimates of shell productivity of large benthic foraminifers.