Construction of DNA damage response gene pprI functiondeficient and function-complementary mutants in Deinococcus radiodurans

PprI, a DNA damage response factor from the extraordinary radioresistant bacterium Deinococcus radiodurans, plays a central regulatory role in multiple DNA damage repair. In this study, a fusion DNA fragment carrying kanamycin resistance gene with the D. radiodurans groEL promoter was cloned by PCR amplification and reversely inserted into the pprl locus in the genome of the wild-type strain R1. The resultingpprl-deficient strain, designated YR1,was very sensitive to ionizing radiation. Meanwhile, the recombinant DNA fragment was cloned into the shuttle vector pRADZ3, and resulted in plasmid pRADK with kanamycin resistance in D. radiodurans. The fragments containing complete pprl gene and 3‘-terminal deletion pprIΔ were cloned into plasmid pRADK. The resulted plasmids designated pRADKpprI and pRADKpprIΔ were then transformed to YR1. Results show that YR1 carrying pRADKpprI was able to fully restore the extreme radioresistance to the same level as the wild-type D. raiodurans R1, whereas YR1 pRADKpprIΔ failed to do so. Construction of DNA repair switch PprI function-deficient and function-complementary mutants in D. radiodurans is not only useful to elucidating the relationship between domains and functions of PprI protein, but also opens the door to the further studies of the biological functions of PprI protein in vivo.     

Functional analysis of the sbcD （dr1921 ） gene of the extremely radioresistant bacterium Deinococcus radiodurans

In eukaryotes, the Mre11-Rad50-Nbs1 (MRN) complex, which resides at the crossroads of DNA repair and checkpoint signaling, rapidly forms prominent foci at damage sites following double-strand break (DSB) induction. This complex carries out the initial processing of the DSB ends. Mutations in the genes that encode components of this complex result in DNA-damage hypersensitivity, genomic in- stability, telomere shortening, and aberrant meiosis. Therefore, the MR proteins are highly conserved during evolution. The bacterial orthologs of Mre11 and Rad50 are the SbcD and SbcC proteins, respec- tively. Deinococcus radiodurans, an extremely radioresistant bacterium, is able to mend hundreds of radiation-induced DSBs. The SbcD and SbcC proteins were identified as the products of the Dr1921 and Dr1922 genes. Disruption of the sbcD gene, by direct reverse-orientation insertional mutagenesis tech- nology, remarkably increases the cells’ sensitivity to various types of DNA damaging agents, such as ionizing radiation, ultraviolet irradiation, hydrogen peroxide, and mitomycin C. We also provide evidence that the drSbcD protein plays an important role in both growth and DNA repair in this organ- ism, especially in repair of DSBs generated after cellular exposure to 6000 Gy of IR. These results demonstrate that the drSbcD protein plays an important role in DSBs repair in D. radiodurans.     

dr1127： A novel gene of Deinococcus radiodurans responsible for oxidative stress

The functional analysis of dr1127,a novel gene in Deinococcus radiodurans was performed in this pa-per. The dr1127 gene was found occasionally in our microarray and 2-DE gel experiments. Mutation of the dr1127 gene decreased the γ-radiation and H2O2 resistance of D. radiodurans,and weakened the scavenging abilities of cell extracts for free radicals (superoxide anion,hydrogen peroxide,and hy-droxyl radical). Further oxidative damage assays demonstrated that the purified DR1127 protein of D. radiodurans could bind to double stranded DNA in vitro and protect DNA from oxidative damage in this way. These results suggest that the dr1127 gene is an important gene that can maintain γ-radiation and oxidative resistance in D. radiodurans and may take part in the oxidative stress process.     

Engineering <Emphasis Type="Italic">Deinococcus radiodurans</Emphasis> into biosensor to monitor radioactivity and genotoxicity in environment

Based on a genetically modified radioresistant bacteria Deinococcus radiodurans, we constructed a real time whole cell biosensor to monitor radioactivity and genotoxicity in highly radioactive environment. The enhanced green fluorescence protein （eGFP） was fused to the promoter of the crucial DNA damage-inducible recA gene from D. radiodurans, and the consequent DNA fragment （PrecA-egfp） carried by plasmid was introduced into D. radiodurans R1 strain to obtain the biosensor strain DRG300. This engineered strain can express eGFP protein and generate fluorescence in induction of the recA gene promoter. Based on the correlation between fluorescence intensity and protein expression level in live D. radiodurans cells, we discovered that the fluorescence induction of strain DRG300 responds in a remarkable dose-dependent manner when treated with DNA damage sources such as gamma radiation and mitomycin C. It is encouraging to find the widely detective range and high sensitivity of this reconstructed strain comparing with other whole cell biosensors in former reports. These results suggest that the strain DRG300 is a potential whole cell biosensor to construct a detective system to monitor the biological hazards of radioactive and toxic pollutants in environment in real time.     

Compression behavior and equation of state of Ni77P23 amorphous alloy

The compression behavior of Ni77P23 amorphous alloy is investigated at room temperature in a diamond-anvil cell instrument using insitu high pressure energy dispersive X-ray diffraction with a syn- chrotron radiation source. The equation of state is determined by fitting the experimental data accord- ing to Birch-Murnaghan equation: -ΔV/V0=0.08606P-3.2×10-4P2 5.7×10-6P3. It is found that the structure of Ni77P23 amorphous alloy is stable under pressures up to 30.5 GPa.     

The composition and distribution of metal clusters in the MoFe protein from a nifZ deletion strain （DJ 194） of Azotobacter vinelandii

Through the anaerobic chromatography on the columns of DEAE 52, Q-Sepharose and Sephacryl S-200, a nitrogenase MoFe protein (△nifZ Av1) was obtained from a nifZ deleted mutant of Azotobacter vinelandii (stain DJ194).The results of Western blotting after anoxic native electrophoresis and SDS-PAGE showed that △nifZ Av1 was similar to wild type MoFe protein (OP Av1) at the electrophoretic mobility, molecular weight and subunit composition. Furthermore, △nifZ Avl was also similar to OP Av1 at the molybdenum content, EPR signal (g≈4.3, 3.65 and 2.01), and the molar extinction coefficient (△ε) of circular dichroism (CD)at 660 nm region. All of these indicated that, besides having the same α2β2 composition as OP Av1, the △nifZ Av1 also contained equal amount of reductive FeMoco in the spin state of S=3/2 to OP Av1. However, the iron content and substrate (C2H2, H^ and N2)-reduction activity of △nifZ Av1 were 74% and 46%-50% of those of OP Av1, respectively. Furthermore, the △ε at around 450 nm, which reflects P-cluster in Av1, was obviously lower than that of OP Av1. It suggested that the difference between △nifZ Avl and OP Av1 resulted from P-cluster rather than FeMoco, and from the half number of P-cluster in △nifZ Av1, but the composition or redoxstate of P-cluster in △nifZ Av1 were not changed. Thus it could propose that △nifZ Av1 is composed of two different αβsubunit pairs. One is a FeMoco-and P-cluster-containing pair, and the other is a P-cluster-deficient but FeMoco-containing pair. Since the deletion of nifZ gene leads to the deficiency of only one of two P-clusters in a α2β2 tetramer, the assembly of P-cluster may not simply depend on one gene product, and so a possible mechanism of NifZ is supposed here.     

The DNA concentration effect on DNA radiation damage induced by <Superscript>7</Superscript>Li ions and γ rays

To evaluate the influence of the DNA concentration in the aqueous solution on DNA radiation damage, the plasmid DNA in the presence or absence of Mannitol （scavenger of free radical OH.） was irradiated by ^7Li ions and γ rays at various DNA concentrations. Gel electrophoresis analysis revealed that the DNA damage of single and double strand breaks induced by irradiation became more severe at lower DNA concentration. In the condition of γ-ray irradiation, most of double strand breaks （DSB） damage was neutralized and less associated with DNA concentration in the presence of mannitol. However, under ^7Li irradiation, DSB damage could not be cleared by mannitol but was gradually aggravated with decreasing DNA concentrations. These findings imply that under low-LET irradiation, most of the DSB damage is generated by free radical OH·diffusion, and thus may be counteracted by scavengers, while at higher-LET irradiation, quite a fraction of DSB induction is caused by direct ionizing energy deposition of heavy ions, which cannot be eliminated. This work also indicates that the proportion between free radical damage and direct ionizing damage is s constant which is independent of DNA concentration when the DNA concentration is under a certain value （50ng/μL）. Our study sheds light on the un- derlying mechanisms in the DNA radiation damage process.     

Improving DNA damage repair ability ofE. coli to UV-radiosensitivity by DNA-mediated gene transfer of low energy ion beam

Using calcium chloride method of transfer gene as control, a new technique of transferring gene by low energy ion beam has been applied to the study of improving DNA damage repair ability ofE. coli to UV-radiosensitivity. The genome DNA pieces ofDeinococcus radiodurans, as “foreign” genetic materials, were introduced into the UV-radiosensitive strains ofE. coli by implantation of 20 keV Ar⁺ at doses ranging from 1 × 10¹⁵ to 2 × 10¹⁵ ions/cm². Results show that the transfected strains present higher UV-radioresistance than that of un-transfected ones and start ones. The survival rate of transfected strains and their unscheduled DNA synthesis (UDS) ability is increased, indicating that the transfer gene is a success.     

Variation of soil Δδ13C values in Xifeng loess-paleosol sequence and its paleoenvironmental implication

The carbon isotopic compositions of soil organic matter （SOM） and total carbonate （TC） in Xifeng Ioess-paleosol sequence have been studied. The δ^13CsoM values vary from -23.8‰ to -20.2‰, which are higher in interglacial than in glacial stages. Contrary to δ^13Csoi values, the δ^13CTc values vary from -8.5‰ to -3.6‰ and are lower in interglacial than in glacial stages. The differences （△δ^13C） between the δ^13CsoM and δ^13CTc values vary from 14.1‰ to 19.4‰. Our results from the Xifeng loesspaleosol sequence indicate that the △δ^13C values represent the ratio of primary carbonate （PC） to secondary carbonate （SC）. The △δ^13C values were high in the loess stages, and the maximal PC-to-SC ratio can reach 6：4. But in the paleosol stages, the △δ^13C values were low, with a small proportion of PC. The △δ^13C values in Ioess-paleosol sequence also indicate the contributions of the dust to the loess sediment in the Chinese Loess Plateau because the dust contains the PC.     

On the Pólya conjecture and the weak Weyl-Berry conjecture

The Pólya conjecture and its connection with the weak Weyl-Berry conjecture are studied Specifically let Ω⊆R ⁿ (n≥1) be a non-empty bounded open set with boundary ∂Ω. LetN ₀(λ, −Δ,Ω) be the Dirichlet counting function and φ(λ) the associated Weyl term. If the interior Minkowski dimension of ∂Ω is δ∈[n−1,n], then under certain realisable conditions we prove that for λ sufficiently large the Pólya conjecture φ(λ) −N ₀(λ,−Δ,Ω)≥0 is true. Under related conditions we also prove thatϕ(λ)−N ₀(λ,−Δ, Ω)≈λ^5/2, as λ→+∞. That is, the Weak Weyl-Berry conjecture is true. Similar results are obtained for the Neumann counting function. Partially supported by the National Natural Science Foundation of China and the Royal Society of London Chen Hua: born in March 8, 1956, Professor     

The Total Irredundance Numbers on Graphs

This paper discusses the total irredundance relations between the graph G and its clone-contraction graph H, that is, let H be the clone-contraction graph of G and v1,v2,...,vk be all contraction vertices ofH. IfS is a maximal total irredundant set of H such that A = S ∩ {V1,V2,…,Vk} contains as few vertices as possible, then S＇= S-A is the maximal total irredundant set of G. Furthermore, we obtain the bound of the total irredundance A（G） number： irt ≤△（G）/2△（G）＋1 n, which n is the order of graph G, and △（G） is maximum degree in G.     

Functional Imaging of Breast Tissue and Clinical Application

0Introduction Near infrared(NIR)techniquesarebasedonsensitive,quantitativemeasurementsoffunctionalcontrastbe tweenhealthyanddiseasedtissue.Recently,researchersshowgreatinterestinmeasuringfunctionalpropertiesofbreasttis suesuchashemoglobinconcentrationoroxygensaturationbyNIRspectroscopyandNIRimaging[1,2].NIRlightcanpene trateseveralcentimetersintotissuebeforeitisattenuatedbe lowdetection.ThemainintrinsicmechanismsofNIRlightat tenuationintissuearethescatteringduetoindexofrefractionvariatio…     

Pairing problem of generators in Kac-Moody algebras

The following result is proved: in any Kac Moody algebra g(A) , (ⅰ) given any non central element h in the Cartan subalgebra  , or (ⅱ) given any real root vector x β, β∈Δ re . There exists y∈g(A) such that the subalgebra generated by y and h or y and x β contains the derived algebra g′(A) of g(A) .     

Studies on the reaction between Laccase ando-methoxyphenol by microcalorimetry

The reaction between Laccase ando-methoxyphenol have been studied by LKB-2107 batch microcalorimetry system. Thermodynamic parameters Δ _r H _m , ΔG _o , ΔG _T ^≠ and kinetic parameters (K _m ,k ₂) have been determined. The process of the reaction has been analyzed from changes in energy by using the transition state theory. Two methods for enhancing catalytic power of Laccase are proposed. The results shown that formation of an enzyme-substrate complex is “anticatalytic”. The enter and sole source of catalytic power is the stabilization of transition state; reactant-state interactions are by nature inhibitory and only waste catalytic power. Supported by the National Natural Science Foundation of China Wang Tianzhi: born in 1968. Ph D     

Removing the impact of wind direction on remote sensing of sea surface salinity

Sea surface salinity (SSS) plays an important role in the sub-polar area, where intrusions with low salinity influence the deep thermohaline circulation and the meridional heal transport. SSS fields and their seasonal and inter-annual variability are thus…     

Changes in the transthylakoid proton gradient are caused by the movement of phycobilisomes in the cyanobacterium <Emphasis Type="Italic">Synechocystis</Emphasis> sp. strain PCC 6803

Phycobilisomes (PBSs) are the main accessory light-harvesting complexes in cyanobacteria and their movement between photosystems (PSs) affects cyclic and respiratory electron transport. However, it remains unclear whether the movement of PBSs between PSs also affects the transthylakoid proton gradient (ΔpH). We investigated the effect of PBS movement on ΔpH levels in a unicellular cyanobacterium Synechocystis sp. strain PCC 6803, using glycinebetaine to immobilize and couple PBSs to photosystem II (PSII) or photosystem I (PSI) by applying under far-red or green light, respectively. The immobilization of PBSs at PSII inhibited decreases in ΔpH, as reflected by the slow phase of millisecond-delayed light emission (ms-DLE) that occurs during the movement of PBSs from PSII to PSI. By contrast, the immobilization of PBSs at PSI inhibited the increase in ΔpH that occurs when PBSs move from PSI to PSII. Comparison of the changes in ΔpH and electron transport caused by the movement of PBSs between PSs indicated that the changes in ΔpH were most likely caused by respiratory electron transport. This will further improve our understanding of the physiological role of PBS movement in cyanobacteria.     

Latitudinal shift and tilt of the ring current during magnetic storms

The equatorial ring current (ERC) theory suggested that the distribution of global disturbed horizontal geomagnetic field only depends on the cosine of station’s latitude. However, we always observe a larger ΔH at higher latitude stations than lower ones, implying that the ERC could tilt or/and shift with respect to the equatorial plane during intense storms. In this paper, we analyze 11 intense magnetic storms from 2000 to 2004, and introduce two configurational factors to characterize the topology of storm time ring current. The results show that ERC has occasionally deviated off equatorial plane with both tilt angle δ_t≈13°―25° and latitude shift δ_s≈0°―21.8°. The ground disturbed field distribution should be improved as ΔH_k = ΔH_maxcos(φ_k-δ ), which agree well with the geomagnetic observations.     

The short C-terminal hydrophilic domain of NhaH Na^＋/H^＋ antiporter from Halobacillus dabanensis with roles in resistance to salt and in pH sensing

The Na^＋/H^＋ antiporter plays key roles in maintaining low cytoplasmic NaNa^＋ level and pH homeostasis, while little is known about the Carboxyl-terminal hydrophilic tails of prokaryotic antiporters. In our previous study, the first Na^＋/H^＋ antiporter gene nhaH from moderate halophiles was cloned from Halobacillus dabanensis D-8 by functional complementation. A topological model suggested that only nine amino acid residues （^395PLIKKLGMI403） existed in the hydrophilic C-terminal domain of NhaH. The C-terminal truncated mutant of NhaH was constructed by PCR strategy and designated as nhaH△C. Salt tolerance experiment demonstrated that the deletion of hydrophilic C-terminal nine amino acid residues significantly inhibited the complementation ability of E. coil KNabc, in which three main Na^＋/H^＋ antiporters nhaA, nhaB and chaA were deleted. Everted membrane vesicles prepared from E. coil KNabc/nhaHAC decreased both Na^＋/H^＋ and Li^＋/H^＋ exchange activities of NhaH, and also resulted in an acidic shift of its pH profile for Na^＋, indicating a critical role of the short C-terminal domain of NhaH antiporter in alkali cation binding/translocation and pH sensing.     

The biomarkers of 2,6,10,15, 19-pentamethylicosenes and their carbon isotopic composition in the sediments from the Gulf of Mexico

The methanogenic archaea and sulfate reduction bacteria are flourishing in the sediments associated with gas venting and gas hydrate settings on the sea floor, where the anaerobic oxidation of methane (AOM) me-diated by these bacteria is the dominant path…     

Effect of internal deletion of Myosin II on growth and development ofDictyostelium discoideum

Four deletion mutantDictyostelium myosin II heavy chain genes, MyΔ824-941 (Δ1/ 3S2), MyΔ934-1454 (ΔS2), MyΔ934-1194 (ΔS2-1) and MyΔ1 157–1454 (ΔS2-2), were transformed by standard electfoporation into mhcA-cells (T-null), a mutantDictyostelium cell devoid of endogenous myosin II heavy chain gene. The growth, development and formation of fruiting bodies of cells expressing those mutant myosin II s under suspension culture were investigated by comparison with the wild type cell. The results indicate that internal deletion of myosin II affeds the growth and development ofDictyastelium. Furthermore, the longer the length of deletion, the more serious the defect in phenotype.