Spatial distribution and vertical variation of arsenic in Guangdong soil profiles, China.

Total of 260 soil profiles were reported to investigate the arsenic spatial distribution and vertical variation in Guangdong province. The arsenic concentration followed an approximately lognormal distribution. The arsenic geometric mean concentration of 10.4 mg/kg is higher than that of China. An upper baseline concentration of 23.4 mg/kg was estimated for surface soils. The influence of soil properties on arsenic concentration was not important. Arsenic spatial distributions presented similar patterns that high arsenic concentration mainly located in limestone, and sandshale areas, indicating that soil arsenic distribution was dependent on bedrock properties than anthropogenic inputs. Moreover, from A- to C-horizon arsenic geometric mean concentrations had an increasing tendency of 10.4, 10.7 to 11.3 mg/kg. This vertical variation may be related to the lower soil organic matter and soil degradation and erosion. Consequently, the soil arsenic export into surface and groundwaters would reach 1040 t year-1 in the study area.

[Possible mechanism of the protective effects of ginsenoside Rg1 on apoptosis in PC12 cells].

AIM: To explore the possible mechanism of the protective effect of ginsenoside Rg1 on dopamine-induced apoptosis in PC12 cells. METHODS: Flow cytometric assay was performed to quantify the apoptotic cells and detect the positive percentage of Bcl-2 and Bax proteins in PC12 cells. The expression of bcl-2 and bax mRNA was identified by reverse transcription polymerase chain reaction (RT-PCR). The CPP32 activity was measured by fluorescent spectrofluorometer. RESULTS: Dopamine-induced apoptosis in PC12 cells was inhibited by pretreatment with 10 mumol.L-1 Rg1. Simultaneously, the expression of Bcl-2 protein and mRNA were increased, and the Bax protein and mRNA were decreased. The activation of CPP32 was inhibited after pretreatment with Rg1. CONCLUSION: Rg1 protects against dopamine-induced apoptosis in PC12 cells and this effect might be attributed to its regulation of the ratio of Bcl-2 to Bax and its inhibition of the activation of CPP32.

The UL5 gene of herpes simplex virus type 1: isolation of a lacZ insertion mutant and association of the UL5 gene product with other members of the helicase-primase complex.

The UL5 gene product is required continuously during viral DNA synthesis (L. Zhu and S. K. Weller, Virology 166:366-378, 1988) and has been shown to be a component of a three protein helicase-primase complex encoded by herpes simplex virus type 1 (J. J. Crute, T. Tsurumi, L. Zhu, S. K. Weller, P.D. Olivo, M. D. Challberg, E. S. Mocarski, and I. R. Lehman, Proc. Natl. Acad. Sci. USA 86:2186-2189, 1989). The other members of the complex are viral proteins encoded by genes UL8 and UL52. In this study, we isolated a permissive cell line (L2-5) which contains the wild-type UL5 gene under the control of the strong and inducible promoter for the large subunit of herpes simplex virus type 1 ribonucleotide reductase (ICP6). An insertion mutant containing a mutation in the UL5 gene (hr99) was isolated by using the insertional mutagen ICP6::lacZ, in which the Escherichia coli lacZ gene is expressed under control of the viral ICP6 promoter. When grown on Vero cells, hr99 does not form plaques or synthesize viral DNA, although both defects are complemented efficiently on the L2-5 cells. These results confirm that the UL5 gene product is essential for viral growth and DNA replication. Furthermore, since no detectable UL5 protein is synthesized in hr99-infected cells, these cells provide a valuable control not only for the detection of the UL5 protein itself but also for the detection of protein-protein interactions with UL8 and UL52 by coimmunoprecipitation. In addition, the lacZ insertion in hr99 provides a convenient screening system for the introduction of site-specific mutations into the viral genome (L. Zhu and S. K. Weller, J. Virol. 66:469-479, 1992). Thus, hr99 is a valuable tool in the structure-function analysis of the UL5 gene.

The six conserved helicase motifs of the UL5 gene product, a component of the herpes simplex virus type 1 helicase-primase, are essential for its function.

The UL5 protein of herpes simplex virus type 1, one component of the viral helicase-primase complex, contains six sequence motifs found in all members of a superfamily of DNA and RNA helicases. Although this superfamily contains more than 20 members ranging from bacteria to mammalian cells and their viruses, the importance of these motifs has not been addressed experimentally for any one of them. In this study, we have examined the functional significance of these six motifs for the UL5 protein through the introduction of site-specific mutations resulting in single amino acid substitutions of the most highly conserved residues within each motif. A transient replication complementation assay was used to test the effect of each mutation on the function of the UL5 protein in viral DNA replication. In this assay, a mutant UL5 protein expressed from an expression clone is used to complement a replication-deficient null mutant with a mutation in the UL5 gene for the amplification of herpes simplex virus origin-containing plasmids. Eight mutations in conserved regions and three similar mutations in nonconserved regions of the UL5 gene were analyzed, and the results indicate that all six conserved motifs are essential to the function of UL5 protein in viral DNA replication; on the other hand, mutations in nonconserved regions are tolerated. These data provide the first direct evidence for the importance of these conserved regions in any member of the superfamily of DNA and RNA helicases. In addition, three motif mutations were introduced into the viral genome, and the phenotypic analyses of these mutants are consistent with results from the transient replication complementation assay. The ability of these three mutant UL5 proteins to form specific interactions with other members of the helicase-primase complex, UL8 and UL52, indicates that the functional domains required for replication activity of UL5 are separable from domains responsible for protein-protein interactions. It is anticipated that this type of structure-function analysis will lead to the identification of protein domains that contribute not only to the enzymatic activities of helicase or primase but also to protein-protein interactions within members of the complex.

Protective effect of ginsenoside Rg1 on dopamine-induced apoptosis in PC12 cells.

AIM: To explore the possible molecular mechanism of exogenous dopamine-induced apoptosis in PC12 cells and the protective effect of ginsenoside Rg1. METHODS: Flow cytometric assay was used to quantify the apoptotic cells and measure the percentage of cells with positive Bcl-2 and Bax proteins. The morphology of apoptotic cells was evaluated by transmission electron microscope. DNA fragmentation was observed by gel electrophoresis. Caspase-3 activity was determined by fluorescent spectrofluorometer and the expressive bcl-2 and bax mRNA by semiquantitative reverse transcription polymerase chain reaction (RT-PCR). RESULTS: Dopamine 0.15, 0.30, 0.45, and 0.60 mmol/L induced PC12 cell apoptosis from 1.1 % +/- 0.4 % (control) to 41 % +/- 3 %, 46.4 % +/ -2.7 %, 53 % +/ -3 %, and 64.5 % +/- 2.7 %, respectively. After treatment with dopamine 0.45 mmol/L following pretreatment with Rg1 10 micromol/L for 24 h, the percentage of apoptotic cells and caspase-3 activity decreased from 53 % +/- 3 % and 683 +/- 8 (mean fluorescence intensity, MFI) to 1.9 % +/- 0.6 % and 325 +/- 5, and the percentage of cells with positive Bcl-2 protein increased from 14.3 % +/- 1.1 % to 25.9 % +/- 1.6 %, however, the percentage of cells with positive Bax protein decreased from 48 % +/- 3 % to 35 % +/- 3 %, compared with group treated with DA 0.45 mmol/L alone. CONCLUSION: Ginsenoside Rg1 protected PC12 cells against apoptosis by inhibiting the activation of caspase-3 and regulating the ratio of Bcl-2 to Bax protein.

Herpes simplex virus 1 helicase-primase: a complex of three herpes-encoded gene products.

In an earlier report, we described a DNA helicase that is specifically induced upon infection of Vero cells with herpes simplex virus 1. We have purified this enzyme to near homogeneity and found it to consist of three polypeptides with molecular weights of 120,000, 97,000, and 70,000. Immunochemical analysis has shown these polypeptides to be the products of three of the genes UL52, UL5, and UL8 that are required for replication of a plasmid containing a herpes simplex 1 origin (oriS). In addition to helicase activity, the enzyme contains a tightly associated DNA primase. Thus, the three-subunit enzyme is a helicase-primase complex that may prime lagging-strand synthesis as it unwinds DNA at the viral replication fork.