The phosphoinositide PI(3,5)P2 mediates activation of mammalian but not plant TPC proteins: functional expression of endolysosomal channels in yeast and plant cells

Two-pore channel proteins (TPC) encode intracellular ion channels in both animals and plants. In mammalian cells, the two isoforms (TPC1 and TPC2) localize to the endo-lysosomal compartment, whereas the plant TPC1 protein is targeted to the membrane surrounding the large lytic vacuole. Although it is well established that plant TPC1 channels activate in a voltage- and calcium-dependent manner in vitro, there is still debate on their activation under physiological conditions. Likewise, the mode of animal TPC activation is heavily disputed between two camps favoring as activator either nicotinic acid adenine dinucleotide phosphate (NAADP) or the phosphoinositide PI(3,5)P₂. Here, we investigated TPC current responses to either of these second messengers by whole-vacuole patch-clamp experiments on isolated vacuoles of Arabidopsis thaliana. After expression in mesophyll protoplasts from Arabidopsis tpc1 knock-out plants, we detected the Arabidopsis TPC1-EGFP and human TPC2-EGFP fusion proteins at the membrane of the large central vacuole. Bath (cytosolic) application of either NAADP or PI(3,5)P₂ did not affect the voltage- and calcium-dependent characteristics of AtTPC1-EGFP. By contrast, PI(3,5)P₂ elicited large sodium currents in hTPC2-EGFP-containing vacuoles, while NAADP had no such effect. Analogous results were obtained when PI(3,5)P₂ was applied to hTPC2 expressed in baker’s yeast giant vacuoles. Our results underscore the fundamental differences in the mode of current activation and ion selectivity between animal and plant TPC proteins and corroborate the PI(3,5)P₂-mediated activation and Na⁺ selectivity of mammalian TPC2.     

Common variants at 12q14 and 12q24 are associated with hippocampal volume

Aging is associated with reductions in hippocampal volume that are accelerated by Alzheimer's disease and vascular risk factors. Our genome-wide association study (GWAS) of dementia-free persons (n = 9,232) identified 46 SNPs at four loci with P values of <4.0 × 10(-7). In two additional samples (n = 2,318), associations were replicated at 12q14 within MSRB3-WIF1 (discovery and replication; rs17178006; P = 5.3 × 10(-11)) and at 12q24 near HRK-FBXW8 (rs7294919; P = 2.9 × 10(-11)). Remaining associations included one SNP at 2q24 within DPP4 (rs6741949; P = 2.9 × 10(-7)) and nine SNPs at 9p33 within ASTN2 (rs7852872; P = 1.0 × 10(-7)); along with the chromosome 12 associations, these loci were also associated with hippocampal volume (P < 0.05) in a third younger, more heterogeneous sample (n = 7,794). The SNP in ASTN2 also showed suggestive association with decline in cognition in a largely independent sample (n = 1,563). These associations implicate genes related to apoptosis (HRK), development (WIF1), oxidative stress (MSR3B), ubiquitination (FBXW8) and neuronal migration (ASTN2), as well as enzymes targeted by new diabetes medications (DPP4), indicating new genetic influences on hippocampal size and possibly the risk of cognitive decline and dementia.     

Human trampling effects on regeneration and age structures of Pinus edulis and Juniperus monosperma

We examined effects of human foot traffic on age structures and densities of seedlings and saplings of Pinus edulis (Colorado pinyon) and Junisperus monosperma (one-seed juniper) in a heavily used urban park, Garden of the Gods, Colorado. Age structures show no stand-destroying disturbances, but they do contain small peaks 85-95 yr ago (minimum age), which have been interpreted as responses to heavy grazing. For Pinus edulis reverse J-shaped age structures indicate a strongly reproducing population, while flat age structures of J. monosperma show low present reproduction. Young trees showed strong preferences for establishing under existing trees and shrubs and not among herbs or on bare soil. Pinus edulis seedling density was reduced by 73% in heavily trampled areas compared to lightly trampled areas. However, there were no differences in density when only the area protected by rocks, shrubs, or trees was considered. This indicates that direct effects such as physical damage and soil erosion kill young trees, and indirect effects, such as lower seed production, do not cause the lower densities in heavily trampled areas. Pinus edulis saplings and J. monosperma seedlings and saplings showed no differences in density across trampling intensities. In heavily trampled areas of Garden of the Gods Park, recent increases in use have apparently reduced Pinus edulis seedling establishment enough that long-term regeneration is threatened. Managers of all pinyon-juniper woodlands must recognize that in areas strongly impacted by foot traffic, and also presumably by similar disturbances such as vehicle traffic, sufficient regeneration likely does not occur to replace trees. The areal extent of severely disturbed areas should be limited, and managers should seek to avoid further degradation of less damaged areas.     

Extensive characterization of sphere models established from colorectal cancer cell lines

Links between cancer and stem cells have been proposed for many years. As the cancer stem cell (CSC) theory became widely studied, new methods were developed to culture and expand cancer cells with conserved determinants of “stemness”. These cells show increased ability to grow in suspension as spheres in serum-free medium supplemented with growth factors and chemicals. The physiological relevance of this phenomenon in established cancer cell lines remains unclear. Cell lines have traditionally been used to explore tumor biology and serve as preclinical models for the screening of potential therapeutic agents. Here, we grew cell-forming spheres (CFS) from 25 established colorectal cancer cell lines. The molecular and cellular characteristics of CFS were compared to the bulk of tumor cells. CFS could be isolated from 72 % of the cell lines. Both CFS and their parental CRC cell lines were highly tumorigenic. Compared to their parental cells, they showed similar expression of putative CSC markers. The ability of CRC cells to grow as CFS was greatly enhanced by prior treatment with 5-fluorouracil. At the molecular level, CFS and parental CRC cells showed identical gene mutations and very similar genomic profiles, although microarray analysis revealed changes in CFS gene expression that were independent of DNA copy-number. We identified a CFS gene expression signature common to CFS from all CRC cell lines, which was predictive of disease relapse in CRC patients. In conclusion, CFS models derived from CRC cell lines possess interesting phenotypic features that may have clinical relevance for drug resistance and disease relapse.     

一种基于SFDD的状态防火墙规则集比对方法

状态防火墙是一种新型防火墙,而传统防火墙决策图(FDD)构造算法并不适用于状态防火墙的规则集比对.本文在FDD基础上,提出一种状态防火墙决策图(SFDD)构造算法,将规则集转化成图形化的等价的SFDD,用于状态防火墙规则集比对.理论分析和仿真实验结果表明,利用SFDD构造算法进行比对,能有效检测出规则集之间的全部不同点;当状态防火墙的状态部分规则和无状态部分规则条目数量均达到3 000时,比对过程所耗费的平均时间不超过2s.     

Review of the fatigue behavior of hard coating–ductile substrate systems

With the wide application of coating materials in aerospace and other fields, their safety under fatigue conditions in service is important.However, research on the fatigue properties of ceramic hard coatings started late, and a unified standard is yet to be established to evaluate the fatigue life of hard coating–ductile substrate systems.Studies also present different opinions on whether coatings can improve or reduce the fatigue life of substrates.In this paper, the influence of the properties of ceramic coatings on fatigue performance is reviewed, and the effects of coating on the mechanism of fatigue crack initiation in substrates are discussed, aiming to help readers understand the fatigue behavior of hard coating–ductile substrate systems.     

Global Optimization in Any Minkowski Metric: A Permutation-Translation Simulated Annealing Algorithm for Multidimensional Scaling

It is well known that considering a non-Euclidean Minkowski metric in Multidimensional Scaling, either for the distance model or for the loss function, increases the computational problem of local minima considerably. In this paper, we propose an algorithm in which both the loss function and the composition rule can be considered in any Minkowski metric, using a multivariate randomly alternating Simulated Annealing procedure with permutation and translation phases. The algorithm has been implemented in Fortran and tested over classical and simulated data matrices with sizes up to 200 objects. A study has been carried out with some of the common loss functions to determine the most suitable values for the main parameters. The experimental results confirm the theoretical expectation that Simulated Annealing is a suitable strategy to deal by itself with the optimization problems in Multidimensional Scaling, in particular for City-Block, Euclidean and Infinity metrics.