PCNA-induced DNA synthesis past cis-syn and trans-syn-I thymine dimers by calf thymus DNA polymerase delta in vitro.

Calf thymus proliferating cell nuclear antigen (PCNA) promoted DNA synthesis past cis-syn and trans-syn-I cyclobutane thymine dimers by calf thymus DNA polymerase delta (Pol delta) in vitro. Templates containing site-specific cis-syn and trans-syn-I thymine dimers were prepared via a combination of solid phase synthesis with photoproduct building blocks and DNA ligation. Extension of a 15-mer primer on the UV dimer-containing templates by Pol delta produced termination and bypass products in a dNTP and PCNA dependent manner. In the absence of PCNA and at dNTP concentrations varying between 1 and 100 microM, Pol delta could not bypass the cis-syn dimer and terminated elongation one nucleotide prior to the 3'-T of the dimer. DNA synthesis past the trans-syn-I dimer was even less efficient. In the presence of PCNA, termination occurred primarily one nucleotide prior to the 3'-T of both dimers at 1 microM dNTPs but opposite the 5'-T of the dimers at 100 microM dNTPs. In addition, under the latter conditions, bypass of the dimers was observed, to the extent of about 30% of the products for the cis-syn dimer and about 15% for the trans-syn-I dimer.     

Reestablishment of self tolerance by suppressor T-cells after active Heymann's nephritis

Adoptive transfer experiments were performed to obtain evidence that the down-regulation of the autoimmune response in rats with active Heymann's nephritis (HN) is due to suppressor T cells. Late in the course of HN antigen-specific OX8+ suppressor T cells were found in the spleen, but never in the draining lymph nodes. These cells were shown to suppress the autoimmune response when transferred to naive recipients that were subsequently challenged. By mixing B cells or helper T cells from rats with HN with suppressor T cells from high-dose tolerant rats we showed that OX8+ suppressor T cells appeared to have a direct suppressive effect on autoreactive B cells. A profound suppressive effect on helper T cells appeared after 10 weeks. Possible mechanisms to account for the failure of Lewis rats to maintain self tolerance are discussed.     

Metabolic Activity and Population Dynamics of Rhizobia Introduced into Unamended and Bentonite-Amended Loamy Sand

Respiration measurements showed that the cumulative amount of CO₂ respired by rhizobia introduced into sterile bentonite-amended loamy sand was significantly higher than it was in unamended loamy sand. The maintenance respiration of rhizobial cells was not influenced by the presence of bentonite clay. Carbon was used more efficiently during growth in bentonite-amended than in unamended loamy sand. The presence of bentonite clay increased the growth rate of rhizobia introduced into sterile soil. Survival studies performed in nonsterile bentonite-amended loamy sand showed that the use of high (10¹⁰ cells per g of dry soil) rather than lower (10⁴ to 10⁷ cells per g of dry soil) inoculum densities increased the final survival levels of introduced rhizobia. In unamended loamy sand, the application of 10¹⁰ or 10⁷ cells per g of dry soil resulted in similar final survival levels. Pore shape and the continuity of the water-filled pore system were suggested to largely determine the colonization rate of protective microhabitats.     

甘南黄河流域4种典型林分土壤C、N、P化学计量特征

土壤碳（C）、氮（N）、磷（P）是参与植物光合作用和影响生态系统初级生产力的主要元素。甘南高原是黄河流域重要的生态屏障,为了解该区不同林分土壤养分状况的差异,选取该区4种典型林分：云杉林、华北落叶松林、巴山冷杉林以及岷江冷杉糙皮桦混交林为研究对象,研究土壤C、N、P化学计量特征。结果表明：（1）岷江冷杉及糙皮桦混交林土壤C、N含量最高,云杉林土壤N、P含量最低。不同林分间P含量差异显著（P<0.05）,不同土层间C、N含量差异均显著（P<0.05）。（2）云杉林土壤C ： N值显著高于其他林分,岷江冷杉及糙皮桦混交林土壤N ： P及C ： P高于其他林分。（3）海拔、土壤pH、容重与土壤含水量是影响土壤养分的重要因素。土壤C含量与N、P含量均显著相关（P<0.05）。总体来说,不同林分土壤化学计量特征具有显著差异,混交林土壤养分状况较纯林好,未来森林管理和植被建设中,可以通过选择合适的树种和提高树种多样性有效改善森林土壤质量。     

Transport of bacterial inoculants through intact cores of two different soils as affected by water percolation and the presence of wheat plants

Abstract Water flow-innduced transport of Burkholderia cepacia strain P2 and Pseudomonas fluorescens strain R2f cells through intact cores of loamy sand and silt loam field soils was measured for two percolation regimes, 0.9 and 4.4 mm h⁻¹, applied daily during 1 hour. For each strain, transport was generally similar between the two water regimes. Translocation of B. cepacia , with 4.4 mm h⁻¹, did occur initially in both soils. In the loamy sand soil, no change in the bacterial distribution occurred during the experiment (51 days). In the silt loam, B. cepacia cell numbers in the lower soil layers were significantly reduced, to levels at or below the limit of detection. Transport of P. fluorescens in both soils also occurred initially and was comparable to that of B. cepacia . Later in the experiment, P. fluorescens was not detectable in the lower soil layers of the loamy sand cores, due to a large decrease in surviving cell numbers. In the silt loam, the inoculant cell distribution did not change with time. Pre-incubation of the inoculated cores before starting percolation reduced B. cepacia inoculant transport in the loamy sand soil measured after 5 days, but not that determined after 54 days. Delayed percolation in the silt loam soil affected bacterial transport only after 54 days. The presence of growing wheat plants overall enhanced bacterial translocation as compared to that in unplanted soil cores, but only with percolating water. Percolation water from silt loam cores appeared the day after the onset of percolation and often contained inoculant bacteria. With loamy sand, percolation water appeared only 5 days after the start of percolation, and no inoculant bacteria were found. The results presented aid in predicting the fate of genetically manipulated bacteria in a field experiment.     

Saccharomyces cerevisiae replication factor C. II. Formation and activity of complexes with the proliferating cell nuclear antigen and with DNA polymerases delta and epsilon.

Lag times in DNA synthesis by DNA polymerase delta holoenzyme were due to ATP-mediated formation of an initiation complex on the primed DNA by the polymerase with the proliferating cell nuclear antigen (PCNA) and replication factor C (RF-C). Lag time analysis showed that high affinity binding of RF-C to the primer terminus required PCNA and that this complex was recognized by the polymerase. The formation of stable complexes was investigated through their isolation by Bio-Gel A-5m filtration. A stable complex of RF-C and PCNA on primed single-stranded mp18 DNA was isolated when these factors were preincubated with the DNA and with ATP, or, less efficiently with ATP gamma S. These and additional experiments suggest that ATP binding promotes the formation of a labile complex of RF-C with PCNA at the primer terminus, whereas its hydrolysis is required to form a stable complex. Subsequently, DNA polymerase delta binds to either complex in a replication competent fashion without further energy requirement. DNA polymerase epsilon did not associate stably with RF-C and PCNA onto the DNA, but its transient participation with these cofactors into a holoenzyme-like initiation complex was inferred from its kinetic properties and replication product analysis. The kinetics of the elongation phase at 30 degrees, 110 nucleotides/s by DNA polymerase delta holoenzyme and 50 nucleotides/s by DNA polymerase epsilon holoenzyme, are in agreement with in vivo rates of replication fork movement in yeast. A model for the eukaryotic replication fork involving both DNA polymerase delta and epsilon is proposed.     

ATP activation of DNA polymerase III holoenzyme of Escherichia coli. I. ATP-dependent formation of an initiation complex with a primed template

ATP (or dATP) stimulates DNA synthesis by DNA polymerase III holoenzyme (holoenzyme) on the synthetic template-primer poly(dA).oligo(dT)12. Nonhydrolyzable ATP analogs and other natural (deoxy)ribonucleoside triphosphates are inactive. Because the nonhydrolyzable analog 5'-deoxyadenylylimidodiphosphate is efficiently used by holoenzyme for incorporation, the ATP (or dATP) requirement for activation of replication of natural DNA could be determined. Analysis of lag times in DNA synthesis and isolation of intermediates showed that ATP (or dATP) is required in the formation of an initiation complex between holoenzyme and primed DNA template, but not for subsequent DNA synthesis. ATP is bound to holoenzyme in the absence of DNA with a KD value of 0.8 microM; 2 to 3 molecules of ATP per molecule of holoenzyme are bound without apparent cooperativity. Binding of ATP to DNA polymerase III (holoenzyme minus beta subunit) is weak (KD greater than 5 microM) and binding to the beta subunit alone is not observed. However, holoenzyme reconstituted by mixing DNA polymerase III with beta subunit binds ATP as tightly (KD = 0.6 microM) as the original holoenzyme.     

Neighbouring group participation in the unblocking of phosphotriesters of nucleic acids.

Two examples of neighbouring group participation during the removal of protecting groups from phosphotriesters of partially or fully protected intermediates of nucleic acids are presented. The first example shows that ammonolysis of aryl groups from phosphotriesters of partially protected - 5'- hydroxy free - nucleic acids (e.g., 4b approximately to; Ar=2C1C 6H4) gives rise to the formation of unnatural nucleic acids (e.g., 7 approximately to and 8 approximately to). The second one illustrates that fluoride ion promoted hydrolysis of 2,2,2-trichloroethyl groups from phosphotriesters of fully protected nucleic acids (e.g., 18a approximately to), having t-butyldimethylsilyl groups at the 2'-positions, leads to the formation of a considerable amount of side-products (e.g., 20 approximately to and 21 approximately to).     

An antigen resembling HL-A7 on the leukocytes of vervet monkeys.

Human leukocyte typing sera of known specificities were used to test the leukocyte antigens of vervet monkeys. The results suggest that these leukocytes contained an antigen resembling the HL-A7 antigen of human leukocytes. This is similar to a previous observation with leukocytes from baboons. These findings are consistent with the suggestion that the 4a/4b complex is the precursor substance from which the other specificities have evolved.