Druggable protein–protein interactions – from hot spots to hot segments

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What's hot in animal biosafety?

In recent years, the emergence or re-emergence of critical issues in infectious disease and public health has presented new challenges and opportunities for laboratory animal care professionals. The re-emergence of bioterrorism as a threat activity of individuals or small groups has caused a heightened awareness of biosecurity and improved biosafety. The need for animal work involving high-risk or high-consequence pathogens and for arthropod-borne diseases has stimulated renewed interest in animal biosafety matters, particularly for work in containment. Application of these principles to animals retained in outdoor environments has been a consequence of disease eradication programs. The anticipated global eradication of wild poliovirus has prompted the promulgation of new biosafety guidelines for future laboratory and animal work. Increased concern regarding the use of biologically derived toxins and hazardous chemicals has stimulated a new categorization of facility containment based on risk assessment. Recognition that prion disease agents and other high-consequence pathogens require safe handling and thorough destruction during terminal decontamination treatment has led to the development of new biosafety guidelines and technologies. The implementation of these guidelines and technologies will promote state-of-the-art research while minimizing risk to laboratory animals, researchers, and the environment.     

Too hot to handle? Balancing increased trapability with capture mortality in hot weather pitfall trapping

Trapping at air temperatures close to, or exceeding, critical thermal maxima is important for comprehensive sampling of vertebrate assemblages and collection of sufficient data for impact assessment. However, pitfall trapping on hot days also potentially exposes trapped animals to stress or death through overheating or desiccation. We investigate causes of mortality from 14 305 captures over a 22‐year pitfall trap study in arid South Australia and compared mortality rates with maximum temperatures, solar radiation and rainfall. Overall mortality rate was 3.2% with chewing by rodents and handling accidents the most influential cause of death recorded. The highest mortality rates were experienced by the tiny skink, Lerista labialis, which was difficult to detect in traps each day and hence problematic to assess the effect of weather variables on capture mortality. For all other abundant species, high maximum temperature was only a significant explanatory variable for increased death rates of the house mouse Mus domesticus, and increased solar radiation was positively related to capture mortality for the house mouse, the frog Neobatrachus sudelli and the small skink Ctenotus schomburgkii. However, capture rates for these taxa and eight other common species would have been significantly lower if trapping did not occur on days of 40 °C or more. We conclude that trapping in hot weather is both desirable and justifiable and suggest techniques for further reducing mortality rates in pitfall studies.     

Biodiversity hotspots: hot for what?

In complex areas of international policy, such as biodiversity conservation, there is a risk that well-promoted strategies will be received by decision makers as a cure-all. The U.S.-based Conservation International is promoting biodiversity hotspots as a 'silver bullet' strategy for conserving most species for least cost. We assess the degree to which this goal is compatible with four social values that characterize the conservation movement. We find that biodiversity hotspots provide only a partial response because conservation does not treat all species as equal. We argue that explicit recognition of such values is fundamental to a structured debate contributing to the development of a common strategy for biodiversity conservation.     

Do some corals like it hot?

Global increases in sea temperatures threaten coral reef resilience because thermal stress can cause corals to bleach; that is, to lose their photosynthetic microalgal symbionts. Recent evidence suggests that some corals associate with genotypes of microalgae that resist future thermal stress, however, these genotypes might provide less energy for growth when thermal stresses are curtailed. Coral reef resilience depends on whether phenotypic and genotypic changes in host-symbiont associations can match projected increases in the frequency and severity of thermal stress, as well as on our ability to ameliorate continuing human impacts.     

Clustering of Mycobacterium avium subsp. paratuberculosis in rabbits and the environment: how hot is a hot spot?

Clustering of pathogens in the environment leads to hot spots of diseases at local, regional, national, and international levels. Scotland contains regional hot spots of Johne's disease (caused by Mycobacterium avium subsp. paratuberculosis) in rabbits, and there is increasing evidence of a link between paratuberculosis infections in rabbits and cattle. The spatial and temporal dynamics of paratuberculosis in rabbits within a hot spot region were studied with the overall aim of determining environmental patterns of infection and thus the risk of interspecies transmission to livestock. The specific aims were to determine if prevalence of paratuberculosis in rabbits varies temporally between seasons and whether the heterogeneous spatial environmental distribution of M. avium subsp. paratuberculosis on a large scale (i.e., regional hot spots) is replicated at finer resolutions within a hot spot. The overall prevalence of M. avium subsp. paratuberculosis in rabbits was 39.7%; the temporal distribution of infection in rabbits followed a cyclical pattern, with a peak in spring of 55.4% and a low in summer of 19.4%. Spatially, M. avium subsp. paratuberculosis-infected rabbits and, thus, the risk of interspecies transmission were highly clustered in the environment. However, this is mostly due to the clustered distribution of rabbits. The patterns of M. avium subsp. paratuberculosis infection in rabbits are discussed in relation to the host's socioecology and risk to livestock.     

Recombination or mutational hot spots in human mtDNA?

Awadalla, Eyre-Walker, and Maynard Smith (1999) recently argued that there might be recombination in human mitochondrial DNA (mtDNA). Their claim was based on their observation of decaying linkage disequilibrium (LD) as a function of physical distance. Their study was much criticized, and follow-up studies have failed to find any evidence for recombination. We argue that the criticisms levied, even if correct, could not possibly explain the findings of Awadalla, Eyre-Walker, and Maynard Smith (1999). Nonetheless, the test proposed by Awadalla, Eyre-Walker, and Maynard Smith (1999 ) is not robust because recombination is not the only explanation for decay of LD. We show that such a pattern can be caused by mutational hot spots as well. However, a closer look at the data suggests that the pattern observed was not caused by mutational hot spots but rather by chance. Thus, there appears to be no evidence for recombination in the mtDNA polymorphism data. In conclusion, we discuss the possibility of detecting recombination in mtDNA and the implications of its existence.     

CHO细胞基因组中稳定hot spot位点研究进展

近些年来,治疗性重组蛋白类药物是生物制药领域研究的热点。工业化生产中常用于重组蛋白表达的细胞系是中国仓鼠卵巢(Chinese hamster ovary,CHO)细胞。传统CHO细胞系的表达大多数基于随机整合的方式,这可能会使目标基因整合到异染色质区域或者不稳定的染色质区域,导致CHO细胞表达不稳定,需要多轮筛选才能获得理想的表达细胞系。最新研究表明,外源基因在CHO细胞预测/特定的基因组位点中进行特异性整合,可以使重组CHO细胞的表达保持长期一致性和稳定性。CHO细胞基因组中高效稳定的转录整合位点被称为热点(hot spot)。阐述CHO细胞基因组稳定的hot spot位点近几年的研究进展,其中包括热门的hot spot位点,以及如何研究新的hot spot位点的方法。总结如何将外源基因高效定位于预测的CHO细胞hot spot位点,实现高水平稳定的表达重组蛋白,为发现新的有效的hot spot位点,构建稳定表达CHO细胞系提供参考。     

Can a genome change its (hot)spots?

A new study by Jeffreys et al. shows that the rate of recombination in recombination hotspots in humans is not constant through time. This observation adds weight to the idea that hotspots are transient on evolutionary timescales. However, questions remain as to what controls their evolution and how these rapid changes influence broad-scale rates of recombination.     

Probing binding hot spots at protein–RNA recognition sites

We use evolutionary conservation derived from structure alignment of polypeptide sequences along with structural and physicochemical attributes of protein–RNA interfaces to probe the binding hot spots at protein–RNA recognition sites. We find that the degree of conservation varies across the RNA binding proteins; some evolve rapidly compared to others. Additionally, irrespective of the structural class of the complexes, residues at the RNA binding sites are evolutionary better conserved than those at the solvent exposed surfaces. For recognitions involving duplex RNA, residues interacting with the major groove are better conserved than those interacting with the minor groove. We identify multi-interface residues participating simultaneously in protein–protein and protein–RNA interfaces in complexes where more than one polypeptide is involved in RNA recognition, and show that they are better conserved compared to any other RNA binding residues. We find that the residues at water preservation site are better conserved than those at hydrated or at dehydrated sites. Finally, we develop a Random Forests model using structural and physicochemical attributes for predicting binding hot spots. The model accurately predicts 80% of the instances of experimental ΔΔG values in a particular class, and provides a stepping-stone towards the engineering of protein–RNA recognition sites with desired affinity.     

Isolations of α-glucosidase-producing thermophilic bacilli from hot springs of Turkey

From 42 different hot springs in six provinces belonging to distinct geographical regions of Turkey, 451 thermophilic bacilli were isolated and 67 isolates with a high amylase activity were selected to determine the α-glucosidase production capacities by using pNPG as a substrate. α-Glucosidase production capacities of the isolates varied within the range from 77.18 to 0.001 U/g. Eleven of our thermophilic bacilli produced α-glucosidase at significant levels comparable with that of the reference strains tested; thus, five strains, F84b (77.18 U/g), A333 (48.64 U/g), F84a (36.64 U/g), E134 (32.09 U/g), and A343 (10.79 U/g), were selected for further experiments. 16S rDNA sequence analysis revealed that these selected isolates all belonged to thermophilic bacilli 16S rDNA genetic group 5, four of them representing the genus Geobacillus, while strain A343 had an uncultured bacterium as the closest relative. Changes in α-glucosidase levels in the intracellular and extracellular fractions were determined during 48-h cultivation of A333, A343, F84a, F84b, E134, and the reference strain G. stearothermophilus ATCC 12980. According to α-glucosidase production type and enzyme levels in intracellular and extracellular fractions, Geobacillus spp. A333, F84a, and F84b were defined as extracellular enzyme producers, whereas the thermophilic bacterium A343 was found to be an intracellular α-glucosidase producer, similar to ATCC 12980 strain. Geobacillus sp. E134 differed in α-glucosidase production type from all tested isolates and the reference strain; it was described as a membrane-associated cell-bound enzyme producer. In this study, apart from screening a great number of new thermophilic bacilli from the hot springs of Turkey, which have not yet been thoroughly studied, five new thermostable α-1,4-glucosidase-producing bacilli that have biotechnological potential with α-glucosidases located at different cell positions were obtained. The text was submitted by the authors in English.     

Why are not all chilies hot? A trade-off limits pungency

Evolutionary biologists increasingly recognize that evolution can be constrained by trade-offs, yet our understanding of how and when such constraints are manifested and whether they restrict adaptive divergence in populations remains limited. Here, we show that spatial heterogeneity in moisture maintains a polymorphism for pungency (heat) among natural populations of wild chilies (Capsicum chacoense) because traits influencing water-use efficiency are functionally integrated with traits controlling pungency (the production of capsaicinoids). Pungent and non-pungent chilies occur along a cline in moisture that spans their native range in Bolivia, and the proportion of pungent plants in populations increases with greater moisture availability. In high moisture environments, pungency is beneficial because capsaicinoids protect the fruit from pathogenic fungi, and is not costly because pungent and non-pungent chilies grown in well-watered conditions produce equal numbers of seeds. In low moisture environments, pungency is less beneficial as the risk of fungal infection is lower, and carries a significant cost because, under drought stress, seed production in pungent chilies is reduced by 50 per cent relative to non-pungent plants grown in identical conditions. This large difference in seed production under water-stressed (WS) conditions explains the existence of populations dominated by non-pungent plants, and appears to result from a genetic correlation between pungency and stomatal density: non-pungent plants, segregating from intra-population crosses, exhibit significantly lower stomatal density (p = 0.003), thereby reducing gas exchange under WS conditions. These results demonstrate the importance of trait integration in constraining adaptive divergence among populations.     

Protein homeostasis — more than resisting a hot bath

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Colonization of sulfide‐oxygen interfaces on hot spring tufa by Thermothrix thiopara

Cream‐colored streamers of Thermothrix thiopara were found at the sulfide‐oxygen interfaces of active tufa mounds where reducing geothermal groundwaters mixed with the oxidizing atmosphere. In the Jemez hot springs, the molar ratio of sulfide to oxygen was 0.3 to 0.8 at streamer locations within the interface. This corresponded to the optimum stoichiometric proportion (0.5) necessary for sulfur metabolism. The mechanism of cell positioning at the interface was studied by shifting the interface location with a plastic cover to extend reducing conditions from the mouth of the spring to the edge of the plastic. Macroscopically visible streamers of filamentous cells became established at the new interface within a period of eight days. They could then be reestablished at the original interface by removing the cover.

Calcite crystals, pyrite crystals, and membrane enrichment vials were incubated on both sides of the interface and the kinetics of colonization determined. The preferential attachment of rod‐shaped cells to pyrite appeared to be the mechanism by which cells located themselves where pyrite occurred in situ, upstream from the interface. The formation of filamentous cells from rod‐shaped cells was induced by oxygen‐limited growth conditions. This moved the cells slightly downstream and directly within the interface.     

Endangered species diversity ‘hot spots’ in Russia and centers of endemism

Geographic distributions of all 125 endangered and threatened animals and all 70 endangered flowering plant species were compiled and mapped by administrative territory (republics, states, provinces, and districts) of the Russian Federation, based on distributional data in the official Red Data Books of the R.S.F.S.R. [Eliseev et al. (eds) (1985) Red Book of the R.S.F.S.R.: Animals. Rosleskhoz Publishing, Moscow; Golovanov et al. (eds). (1988). Red Book of the R.S.F.S.R.: Plants. Rosagprom Publishing, Moscow]. Territories were ranked using an algorithm that gives highest rank to the single territory with the greatest number of endangered species present, then iteratively assigns the next highest rank to the territory containing the greatest number of species not found in a territory of higher rank. This algorithm minimizes the number of territories necessary to include one population of each endangered species, but may designate very low rank to a territory that holds high numbers of endangered species if many of those species are found in a territory of higher rank. When different sets of species were used to determine territory rank with this algorithm the only significant correlation between rank lists was between ranks generated when all species are used and when only species endemic to single territories were used. Mountainous territories on Russia's southern borders held the greatest numbers of endangered species and were highly ranked whether or not species occurring outside of Russia were excluded from the ranking algorithm. The Maritime state, Krasnodar state, Dagestan republic, Sakhalin province and the Jewish province all are centers of endangered species diversity that together contain 50 or more of endangered species for multiple taxonomic groups; this was the Dobson et al. [(1997) Science 275: 550–553] definition of an endangered species hot spot. These are all mountainous territories on Russia's southern border. Precise territorial ranking within the mountainous southern regions of the Southern Far East, Caucasus, and Altai/Sayan regions of southern Siberia was highly influenced by species endemic to single territories.     

Are low temperature habitats hot spots of microbial evolution driven by viruses?

There is an increasing body of evidence to show that viruses are important drivers of microbial evolution and that they can store a great deal of the Earth's microbial diversity in their genomes. Examination of microbial diversity in polar regions has revealed a higher than expected diversity of viruses, bacteria and eukaryotic microbes. Further, the few available studies in polar regions reveal that viral control of microbial mortality is important in these habitats. In this opinion article, we argue that strong relationships between viruses and their hosts in a range of polar habitats could be key in explaining why polar regions are in fact hot spots of microbial diversity and evolution. Further, we argue that periodic glaciations, and particularly the Neoproterozoic low-latitude glaciation, known as 'snowball Earth', could have been periods of intense diversification in aquatic refuges.     

Metabotropic glutamate receptors in neurodegeneration/neuroprotection: Still a hot topic?

Moving from early studies, we here review the most recent evidence linking metabotropic glutamate (mGlu) receptors to processes of neurodegeneration/neuroprotection. The use of knockout mice and subtype-selective drugs has increased our knowledge of the precise role played by individual mGlu receptor subtypes in these processes. Activation of mGlu1 and mGlu5 receptors may either amplify or reduce neuronal damage depending on the context and the nature of the toxic insults. In contrast, mGlu1 and mGlu5 receptors antagonists are consistently protective in in vitro and in vivo models of neuronal death. A series of studies suggest that mGlu1 receptor antagonists or negative allosteric modulators (NAMs) are promising candidates for the treatment of ischemic brain damage, whereas mGlu5 receptor NAMs, which have been clinically developed for the treatment of Parkinson's disease (PD) and l-DOPA-induced dyskinesias, protect nigro-striatal dopaminergic neurons against 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) toxicity in mice and monkeys. Activation of glial mGlu3 receptors promotes the formation of various neurotrophic factors, such as transforming growth factor-β (TGF-β), glial-derived neurotrophic factor (GDNF), nerve growth factor (NGF), and brain-derived neurotrophic factor (BDNF). Hence, selective mGlu3 receptor agonists or positive allosteric modulators (PAMs) (not yet available) are potentially helpful in the treatment of chronic neurodegenerative disorders such as PD, Alzheimer's disease (AD), and amyotrophic lateral sclerosis. Selective mGlu2 receptor PAMs should be used with caution in AD patients because these drugs are shown to amplify β-amyloid neurotoxicity. Finally, mGlu4 receptor agonists/PAMs share with mGlu5 receptor NAMs the ability to improve motor symptoms associated with PD and attenuate nigro-striatal degeneration at the same time. No data are yet available on the role of mGlu7 and mGlu8 receptors in neurodegeneration/neuroprotection.