DNA replication stress: Causes,resolution and disease

DNA replication is a fundamental process of the cell that ensures accurate duplication of the genetic information and subsequent transfer to daughter cells. Various pertubations, originating from endogenous or exogenous sources, can interfere with proper progression and completion of the replication process, thus threatening genome integrity. Coordinated regulation of replication and the DNA damage response is therefore fundamental to counteract these challenges and ensure accurate synthesis of the genetic material under conditions of replication stress. In this review, we summarize the main sources of replication stress and the DNA damage signaling pathways that are activated in order to preserve genome integrity during DNA replication. We also discuss the association of replication stress and DNA damage in human disease and future perspectives in the field.     

The primary structure of Pseudomonas cytochrome c peroxidase

The primary structure of Pseudomonas cytochrome c peroxidase is presented. The intact protein was fragmented with cyanogen bromide into five fragments; partial cleavage was observed at a Met-His bond of the protein. The primary structure was established partly by automatic Edman degradations, partly by manual sequencing of peptides obtained with trypsin, thermolysin, chymotrypsin, pepsin, subtilisin and Staphylococcus aureus V8 endopeptidase. The order of the cyanogen bromide fragments was further confirmed by overlapping peptides obtained by specific cleavage of the whole protein. Pseudomonas cytochrome c peroxidase consists of 302 amino acid residues giving a calculated M_r of 33 690.     

Structural model studies for the high-valent intermediate Q of methane monooxygenase from broken-symmetry density functional calculations

Mössbauer isomer shift parameters have been obtained for both density functional theory (DFT) OPBE and OLYP functionals by linear regressions between the measured isomer shifts and calculated electron densities at Fe nuclei for a number of Fe^2+,2.5+ and Fe^{2.5+,3+,3.5+,4+} complexes grouped separately. The calculated isomer shifts and quadrupole splittings on the sample Fe complexes from OPBE and OLYP functionals are similar to those of PW91 calculations [J. Comput. Chem. 27 (2006) 1292], however the fit parameters from the linear regressions differ between PW91 and OPBE, OLYP. Four models for the active site structure of intermediate Q of the hydroxylase component of soluble methane monooxygenase (MMOH) have been studied, using three DFT functionals OPBE, OLYP, and PW91, incorporated with broken-symmetry methodology and the conductor-like screening (COSMO) solvation model. The calculated properties, including optimized geometries, electronic energies, pK_a’s, Fe net spin populations, and Mössbauer isomer shifts and quadrupole splittings, have been reported and compared with available experimental values. The high-spin antiferromagnetically (AF) coupled Fe⁴⁺ sites are correctly predicted by OPBE and OLYP methods for all active site models. PW91 potential overestimates the Fe-ligand covalencies for some of the models because of spin crossover. Our calculations and data analysis support the structure (our current model II shown in Fig. 8) proposed by Friesner and Lippard’s group [J. Am. Chem. Soc. 123 (2001) 3836-3837], which contains an Fe⁴⁺(μ-O)₂Fe⁴⁺ center, one axial water which also H-bonds to both side chains of Glu243 and Glu114, and one bidentate carboxylate group from the side chain of Glu144, which is likely to represent the active site of MMOH-Q. A new model structure (model IV shown in Fig. 9), which has a terminal hydroxo and a protonated His147 which is dissociated from a nearby Fe, is more asymmetric in its Fe(μ-O)₂Fe diamond core, and is another very good candidate for intermediate Q.     

Landscape structure influences tree density patterns in fragmented woodlands in semi‐arid eastern Australia

Landscape and local‐scale influences are important drivers of plant community structure. However, their relative contribution and the degree to which they interact remain unclear. We quantified the extent to which landscape structure, within‐patch habitat and their confounding effects determine post‐clearing tree densities and composition in agricultural landscapes in eastern subtropical Australia. Landscape structure (incorporating habitat fragmentation and loss) and within‐patch (site) features were quantified for 60 remnant patches of Eucalyptus populnea (Myrtaceae) woodland. Tree density and species for three ecological maturity classes (regeneration, early maturity, late maturity) and local site features were assessed in one 100 × 10 m plot per patch. All but one landscape characteristic was determined within a 1.3‐km radius of plots; Euclidean nearest neighbour distance was measured inside a 5‐km radius. Variation in tree density and composition for each maturity class was partitioned into independent landscape, independent site and joint effects of landscape and site features using redundancy analysis. Independent site effects explained more variation in regeneration density and composition than pure landscape effects; significant predictors were the proportion of early and late maturity trees at a site, rainfall and the associated interaction. Conversely, landscape structure explained greater variation in early and late maturity tree density and composition than site predictors. Area of remnant native vegetation within a landscape and patch characteristics (area, shape, edge contrast) were significant predictors of early maturity tree density. However, 31% of the explained variation in early mature tree differences represented confounding influences of landscape and local variables. We suggest that within‐patch characteristics are important in influencing semi‐arid woodland tree regeneration. However, independent and confounding effects of landscape structure resulting from previous vegetation clearing may have exerted a greater historical influence on older cohorts and should be accounted for when examining woodland dynamics across a broader range of environments.     

Regeneration and inhibition of proton pumping activity of bacteriorhodopsin blue membrane by cationic amine anesthetics

Bacteriorhodopsin (bR) is the prototype of an integral membrane protein with seven membrane-spanning α-helices and serves as a model of the G-protein-coupled drug receptors. This study is aimed at reaching a greater understanding of the role of amine local anesthetic cations on the proton transport in the bR protein, and furthermore, the functional role of “the cation” in the proton pumping mechanism. The effect of the amine anesthetic cations on the proton pump in the bR blue membrane was compared with those by divalent (Ca²⁺, Mg²⁺ and Mn²⁺) and monovalent metal cations (Li⁺, Na⁺, K⁺ and Cs⁺), which are essential for the correct functioning of the proton pumping of the bR protein. The results suggest that the interacting site of the divalent cation to the bR membrane may differ from that of the monovalent metal cation. The electric current profile of the bR blue membrane in the presence of the amine anesthetic cations was biphasic, involving the generation and inhibition of the proton pumping activity in a concentration-dependent manner. The extent of the regeneration of the proton pump by the additives increased in the order of monovalent metal cation<monovalent amine anesthetic cation<divalent metal cation. We found that organic cations such as the amine anesthetics can also regenerate the proton pump in the bR protein. The inhibition of proton transport in the bR protein by the anesthetic cations was elucidated using the wild type, the E204Q and the D96N mutated bRs. The hydrophobic interaction of the amine anesthetics with the bR protein plays an important part in inhibiting the bR proton pump.     

黄龙山林区封育油松种群动态研究

对不同封育年限油松种群的高度、径级、年龄结构进行分析,运用马尔可夫转移矩阵预测种群密度动态。结果表明,不同封育年限种群的生长发育明显不同。种群个体高度以2～4m和6～8。占优势,胸径以0～2cm的小径阶和〉20cm的大径阶占较大比重;在各封育系列中,种群的密度中、大级出现几率较大。在封育45年前,种群的年龄结构中大树所占比例最大;45年后,小树和幼苗则较多,但种群弱小个体大量死亡。本区天然林保护年限最短应确定为45年。     

Evaluation of lamb performance and costs in motherless rearing of German Grey Heath sheep under field conditions using automatic feeding systems

Performance and costs were evaluated for 205 German Grey Heath lambs reared artificially under field conditions using three feeding and three weaning regimes. In phase 1 (pre-weaning), ad libitum feeding of milk replacer (MR) on an automatic lamb feeder was compared to restricted MR feeding on the modified calf feeder “Stand alone II^®” or with buckets. In phase 2 (after weaning at 12 kg body weight), abrupt weaning was compared to gradual weaning systems with reduction of the amount or the concentration of MR over 2 weeks. In phase 1, average daily gains (ADG) were highest in the Ad libitum group (0.262 ± 0.032 kg/d), followed by Bucket and Stand alone with 0.227 ± 0.036 kg/d and 0.209 ± 0.023 kg/d, respectively (P < 0.05). In contrast, feed conversion rate was best for group Stand alone (1.17:1), compared to 1.20:1 in group Bucket and 1.23:1 in group Ad libitum. In phase 2, abrupt weaning caused weight loss in lambs from the Ad libitum and Bucket groups, but not in group Stand alone. No post-weaning growth check was seen in lambs weaned gradually. In general, differences in body weight (BW) between groups were low, and on d 70, no significant difference was seen between feeding groups, whereas lambs weaned abruptly still had lower BW than lambs weaned by reducing the amount of MR (P < 0.05). Total lamb mortality was 3.9%, with no accumulation of infectious disease in any of the feeding or weaning groups. Economic analysis revealed that total costs in phase 1 were highest for group Stand alone (57–70 € per lamb), followed by the Bucket group (63 € per lamb). Although MR consumption was higher in the Ad libitum group, this method was the most cost-efficient (50–54 € per lamb), because purchase price of the feeder and labor costs were relatively low. In phase 2, gradual weaning systems added costs of 4.70–9.50 € per lamb. Our findings indicate that the most efficient way to rear 60–120 lambs artificially under field conditions is by using an automatic ad libitum lamb feeder followed by abrupt weaning.     

DNA Fingerprinting of Paecilomyces Strains of Potential use for the Biological Control of Pests

Summary Telomeric fingerprinting was found to be highly differentiating for Paecilomyces fumosoroseus and Paecilomyces lilacinus isolates in comparison to intron splice site PCR and is therefore a good method for quality control of future products based on these fungi. Although the telomeric restriction length polymorphisms correctly divided the isolates into their appropriate species, further correlation with host range or geographical origin of the isolates was not found. In this respect, intron splice site PCR was more informative taxonomically. The chromosome numbers inferred from telomeric fingerprints were seven chromosomes for P. lilacinus and between six and nine chromosomes for P. fumosoroseus.     

Effects of microsite conditions on seedling establishment on the foreland of Coleman Glacier,Washington

Abstract. Questions: How do physical microsite conditions of microsites affect germination and seedling survival in different successional stages? Do different species germinate in similar microsites in a given successional stage? Location: Coleman Glacier foreland, Mount Baker, Washington State, USA. Methods: Two methods were used to characterize safe sites. 1. Grids of 300 10 cm × 10 cm plots were located in four different age classes on the foreland. 2.105 pairs of plots, with and without seedlings of Abies amabilis, were located in each age class. For each plot we identified all seedlings and all individuals < 1 m tall. Microsite characteristics such as topography and presence of rocks or woody debris were noted for each plot. Microsite characteristics were compared between plots with and without each species. In addition we examined the effect of distance from seed sources on the presence of Alnus viridis seeds and seedlings in a newly disturbed area. Results: In early successional sites, seedlings of several species were positively associated with depressions and presence of rocks, and negatively associated with ridges. Patterns were generally consistent among species. In later succession, seedlings were not significantly associated with any microsite characteristics. For Alnus viridis, seed density decreased with distance from seed sources but seedling density did not. Conclusions: Because of harsh conditions in early succession, physical microsites are important, and most species have similar microsite requirements. In later succession, physical microsites characteristics are not as important and are more variable. Microsites appear to be more important than seed rain in controlling the distribution of Alnus viridis in early succession.