Identification of a highly conserved domain in the EcoRII methyltransferase which can be photolabeled with S-adenosyl-L-[methyl-3H]methionine. Evidence for UV-induced transmethylation of cysteine 186

DNA methyltransferases can be photolabeled with S-adenosyl-L-methionine (AdoMet). Specific incorporation of radioactivity has been demonstrated after photolabeling with either [methyl-3H]AdoMet or [35S]AdoMet (Som, S., and Friedman, S. (1990) J. Biol. Chem. 265, 4278-4283). The labeling is believed to occur at the AdoMet binding site. With the purpose of localizing the site responsible for [methyl-3H]AdoMet photolabeling, we cleaved the labeled EcoRII methyltransferase by chemical and enzymatic reactions and isolated the radiolabeled peptides by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and high pressure liquid chromatography. The labeled peptides were identified by amino-terminal sequencing. A common region was localized which accounted for 65-70% of the total label. This region includes a highly conserved core sequence present in all DNA (cytosine 5)-methyltransferases. One such fragment was digested further with chymotrypsin, and amino acid analysis of the resulting 3H-labeled peptide was consistent with the sequence Ala-Gly-Phe-Pro-(Cys)-Gln-Pro-Phe-Ser-Leu. However, the cysteine residue was not recovered as carboxymethylcysteine. The Pro-Cys bond was found to be protected from cleavage at cysteine residues after cyanylation. These results suggest that the cysteine residue is modified by the labeling reaction. The chymotryptic fragment was hydrolyzed enzymatically to single amino acids, and the labeled amino acid was identified as S-methylcysteine by thin layer chromatography. These results indicate that the cysteine residue is located at or close to the AdoMet binding site of EcoRII methyltransferase.     

Predicting the distributions of predator (snow leopard) and prey (blue sheep) under climate change in the Himalaya

Future climate change is likely to affect distributions of species, disrupt biotic interactions, and cause spatial incongruity of predator–prey habitats. Understanding the impacts of future climate change on species distribution will help in the formulation of conservation policies to reduce the risks of future biodiversity losses. Using a species distribution modeling approach by MaxEnt, we modeled current and future distributions of snow leopard (Panthera uncia) and its common prey, blue sheep (Pseudois nayaur), and observed the changes in niche overlap in the Nepal Himalaya. Annual mean temperature is the major climatic factor responsible for the snow leopard and blue sheep distributions in the energy‐deficient environments of high altitudes. Currently, about 15.32% and 15.93% area of the Nepal Himalaya are suitable for snow leopard and blue sheep habitats, respectively. The bioclimatic models show that the current suitable habitats of both snow leopard and blue sheep will be reduced under future climate change. The predicted suitable habitat of the snow leopard is decreased when blue sheep habitats is incorporated in the model. Our climate‐only model shows that only 11.64% (17,190 km²) area of Nepal is suitable for the snow leopard under current climate and the suitable habitat reduces to 5,435 km² (reduced by 24.02%) after incorporating the predicted distribution of blue sheep. The predicted distribution of snow leopard reduces by 14.57% in 2030 and by 21.57% in 2050 when the predicted distribution of blue sheep is included as compared to 1.98% reduction in 2030 and 3.80% reduction in 2050 based on the climate‐only model. It is predicted that future climate may alter the predator–prey spatial interaction inducing a lower degree of overlap and a higher degree of mismatch between snow leopard and blue sheep niches. This suggests increased energetic costs of finding preferred prey for snow leopards – a species already facing energetic constraints due to the limited dietary resources in its alpine habitat. Our findings provide valuable information for extension of protected areas in future.     

Long-Lasting Immune Protection and Other Epidemiological Findings after Chikungunya Emergence in a Cambodian Rural Community,April 2012

The East/Central/South African genotype of Chikungunya virus with the E1-A226V mutation emerged in 2011 in Cambodia and spread in 2012. An outbreak of 190 cases was documented in Trapeang Roka, a rural village. We surveyed 425 village residents within 3–4 weeks after the outbreak, and determined the sensitivity and specificity of case definitions and factors associated with infection by CHIKV. Self-reported clinical presentation consisted mostly of fever, rash and arthralgia. The presence of all three clinical signs or symptoms was identified as the most sensitive (67%) and specific (84%) self-reported diagnostic clinical indicator compared to biological confirmation by MAC-ELISA or RT-PCR used as a reference. Having an indoor occupation was associated with lower odds of infection compared with people who remained at home (adjOR 0.32, 95%CI 0.12–0.82). In contrast with findings from outbreaks in other settings, persons aged above 40 years were less at risk of CHIKV infection, likely reflecting immune protection acquired when Chikungunya circulated in Cambodia before the Khmer Rouge regime in 1975. In view of the very particular history of Cambodia, our epidemiological data from Trapeang Roka are the first to support the persistence of CHIKV antibodies over a period of 40 years.     

Nitroglycerin, a nitric oxide generator attenuates ferric nitrilotriacetate-induced renal oxidative stress, hyperproliferative response and necrosis in ddY mice

Nitric oxide (NO) is a short lived, readily diffusible intracellular messenger molecule associated with multiple organ-specific regulatory functions. In this communication, we elucidate the effect of exogenous NO administration, using nitroglycerin (GTN), on ferric nitrilotriacetate (Fe-NTA)-induced renal oxidative stress, hyperproliferative response and necrosis in ddY mice. Fe-NTA is a known complete renal carcinogen as well as renal and hepatic tumor promoter, which act by generating oxidative stress in the tissues. GTN treatment to ddY mice prior to Fe-NTA administration resulted in a highly significant protection against Fe-NTA-induced renal oxidative stress, hyperproliferative response and necrosis. In oxidative stress protection studies, the decrease in the level of renal glutathione and antioxidant enzyme activities induced by Fe-NTA were significantly reversed by GTN pretreatment in a dose-dependent manner (12-46% recovery, P<0.05-0.001). GTN pretreatment also resulted in a dose-dependent inhibition (24-39% inhibition, P<0.05-0.001) of Fe-NTA-induced lipid peroxidation as measured by TBARS formation in renal tissues. Similarly, in hyperproliferation protection studies, GTN pretreatment showed a strong inhibition of Fe-NTA-induced renal ornithine decarboxylase (ODC) activity (51-57% inhibition, P<0.001) and [3H]thymidine incorporation (43-58% inhibition, P<0.001) into renal DNA. GTN pretreatment almost completely prevented kidney biomolecules from oxidative damage and protected the tissue against the observed histopathological alterations. From this data, it can be concluded that exogenously produced NO from GTN might scavenge reactive oxygen species (ROS) and decreases toxic metabolites of Fe-NTA and thereby inhibiting renal oxidative stress. In addition, exogenously produced NO can also inhibit Fe-NTA-induced hyperproliferative response by down-regulating the activity of ODC and the rate of [3H]thymidine incorporation into renal DNA and could be suggested as another possible clinical application for this NO-donor (GTN, traditionally used as a vasodilator) in oncological medicine.     

Identification of a type 1 peroxisomal targeting signal in a viral protein and demonstration of its targeting to the organelle

Peroxisomes are unimembrane, respiratory organelles of the cell. Transport of cellular proteins to the peroxisomal matrix requires a type 1 peroxisomal targeting signal (PTS1) which essentially constitutes a tripeptide from the consensus sequence S/T/A/G/C/N-K/R/H-L/I/V/M/A/F/Y. Although PTS-containing proteins have been identified in eukaryotes, prokaryotes, and parasites, viral proteins with such signals have not been identified so far. We report here the first instance of a virus, the rotavirus, which causes infantile diarrhea worldwide, containing a functional C-terminal PTS1 in one of its proteins (VP4). Analysis of 153 rotavirus VP4-deduced amino acid sequences identified five groups of conserved C-terminal PTS1 tripeptide sequences (SKL, CKL, GKL, CRL, and CRI), of which CRL is represented in approximately 62% of the sequences. Infection of cells by a CRL-containing representative rotavirus (SA11 strain) and confocal immunofluorescence analysis revealed colocalization of VP4 with peroxisomal markers and morphological changes of peroxisomes. Further, transient cellular expression of green fluorescent protein (GFP)-fused VP4CRL resulted in transport of VP4 to peroxisomes, whereas the chimera lacking the PTS1 signal, GFP-VP4DeltaCRL, resulted in diffuse cytoplasmic staining, suggesting a CRL-dependent targeting of the protein. The present study therefore demonstrates hitherto unreported organelle involvement, specifically of the peroxisomes, in rotaviral infections as demonstrated by using the SA11 strain of rotavirus and opens a new line of investigation toward understanding viral pathogenesis and disease mechanisms.     

Surface Plasmon Resonance and Enhanced Fluorescence Application of Single-step Synthesized Elliptical Nano Gold-embedded Antimony Glass Dichroic Nanocomposites

A novel series of elliptical gold (Au⁰) nanoparticles (18–40 nm) embedded antimony glass (K₂O-B₂O₃-Sb₂O₃) dichroic nanocomposites have been synthesized by a single-step melt-quench in-situ thermochemical reduction technique. X-ray and selected area electron diffractions manifest growth of Au⁰ nanoparticles along the (111) and (200) crystallographic planes. The transmission electron microscopic image reveals elliptical Au⁰ nanoparticles having an aspect ratio varying in the range 1.2–2.1. The dichroic behavior of the nanocomposites arises due to elliptical shape of the Au⁰ nanoparticles. These nanocomposites show strong surface plasmon resonance (SPR) band of Au nanoparticles in the range 610–681 nm and it exhibit red-shifts with increasing Au concentration. They, when co-doped with Sm₂O₃ and excited at 949 nm, exhibit about sevenfold enhancement of the upconverted red emission transition of ⁴G_5/2→⁶H_9/2 at 636 nm due to local electric field enhancement effect of Au⁰ nanoparticles induced by its SPR. These nanocomposites are the promising materials for laser, display, and various nanophotonic applications.     

Photomechanical wave-assisted molecular delivery in oral biofilms

Photomechanical waves (PW), the product of an intense light beam interaction with a target material, enhance molecular delivery across biological membranes and skin. The ability to deliver methylene blue (MB), a fluorescent probe and photosensitizer, into bacterial biofilms was demonstrated by applying PW on saliva-derived multi-species biofilms that were developed on agar surfaces in 24-well plates. PW were generated with a Q-switched Nd:YAG laser and were directed into the biofilms in the presence of 25 μg/ml MB. The biofilms were then irradiated with red light at 665 nm. After illumination, adherent bacteria were scraped and spread over the surface of blood agar plates. Survival fractions were calculated by counting bacterial colonies. Microbial analysis was performed via a colony lift method and a DNA checkerboard assay using whole genomic probes to 40 oral microorganisms. Visual analysis by confocal scanning laser microscopy demonstrated that the application of PW enhanced the penetration depth of MB in biofilms. Exposure to MB, PW and light led to a significant reduction of the mean levels of log₁₀ CFU counts compared with the group that received MB and light (P = 0.006). The DNA checkerboard assay showed some benefit from PW-assisted phototargeting in 25 biofilm microorganisms relative to phototreatment alone. Our data provide a basis for further exploration and optimization of PW parameters for complete eradication of microorganisms in oral microcosm biofilms.     

Hemiclonal reproduction slows down the speed of Muller's ratchet in the hybridogenetic frog Rana esculenta

Rare recombination in otherwise asexually reproducing organisms is known to beneficially influence the fitness in small populations. In most of the investigated organisms, asexual and rare sexual generations with recombination follow each other sequentially. Here we present a case where clonal reproduction and rare recombination occur simultaneously in the same population. The hybridogenetic water frog Rana esculenta (E), a hybrid between R. lessonae (L) and R. ridibunda (R) produces gametes that only contain the unaltered maternal R part of their genome. New generations of R. esculenta usually arise from E x L matings. Intraspecific E x E matings produce mostly inviable offspring, but in rare cases, female R. ridibunda arise from such matings which are capable of recombination. In the absence of conspecific males, these R females have to mate with E males, which results in further R females, or with L males, which produces new E lineages. This indirect mechanism reintroduces recombination into the otherwise clonally transmitted R genomes in R. esculenta populations. In this study, we show through Monte Carlo simulations that, in most cases, it is sufficient that only between 1 % and 10 % of mixed water frog populations consist of R females to prevent or significantly reduce the fixation and accumulation of deleterious mutations.     

查谟-克什米尔地区查谟后西瓦立克沉积中的亚洲象化石(英文)

描述了一件象属( Elephas) 的右上臼齿化石。标本产自查谟紧靠上西瓦立克亚群巨砾岩组( Boulder Conglomerate Formation) 之上的砂质、粉砂质泥岩夹砾石层中,化石地点位于查谟市南10 km,Kharian 村北约500 m 处。根据齿板数、齿脊频率、釉质层厚度、冠高指数、绝对大小和齿长/齿高指数等牙齿形态参数,暂时将之归为 Elephas cf. E. maximus indicus。还简短讨论了象属的地理分布和地质时代。