Are amyloid-degrading enzymes viable therapeutic targets in Alzheimer's disease?

: The amyloid cascade hypothesis of Alzheimer's disease envisages that the initial elevation of amyloid β-peptide (Aβ) levels, especially of Aβ(1-42) , is the primary trigger for the neuronal cell death specific to onset of Alzheimer's disease. There is now substantial evidence that brain amyloid levels are manipulable because of a dynamic equilibrium between their synthesis from the amyloid precursor protein and their removal by amyloid-degrading enzymes (ADEs) providing a potential therapeutic strategy. Since the initial reports over a decade ago that two zinc metallopeptidases, insulin-degrading enzyme and neprilysin (NEP), contributed to amyloid degradation in the brain, there is now an embarras de richesses in relation to this category of enzymes, which currently number almost 20. These now include serine and cysteine proteinases, as well as numerous zinc peptidases. The experimental validation for each of these enzymes, and which to target, varies enormously but up-regulation of several of them individually in mouse models of Alzheimer's disease has proved effective in amyloid and plaque clearance, as well as cognitive enhancement. The relative status of each of these enzymes will be critically evaluated. NEP and its homologues, as well as insulin-degrading enzyme, remain as principal ADEs and recently discovered mechanisms of epigenetic regulation of NEP expression potentially open new avenues in manipulation of AD-related genes, including ADEs.     

Conservation targets for viable species assemblages?

Estimates of minimum areas required for effective biodiversity conservation differ substantially. Scientific reserve design and placement procedures indicate that between 30 and 75% of any region may be required to sample biodiversity features. These estimates do not routinely incorporate measures for sampling viable populations of species or explore the area requirements of sampling viable populations of species assemblages. To determine the area requirements for sampling viable populations of a herbivore assemblage, spatially explicit abundance data from the Kruger National Park, South Africa, were analyzed. Area requirements were consistently above 50% and were unaffected by selected target population sizes. In addition, area requirements appeared to be insensitive to selection unit size (analytical grain), habitat quality, the coarseness of the land classification system used or the presence of low-density species. Thus, traditional conservation area targets of 10–15% appear inadequate for representing viable populations of a herbivore assemblage from African savanna regions. This suggests that conservation targets of at least 50% of land classification units may represent a more appropriate conservation rule of thumb, or alternatively, that the use of data independent conservation targets may need to be abandoned.     

Are hyaluronan receptors involved in three-dimensional cell migration?

Hyaluronan (HA), an unbranched polysaccharide consisting of repeated glucuronic acid/N-acetylglucosamine disaccharide units, is ubiquitously present in the extracellular matrix of many tissues (for a more comprehensive review see: Fraser et al., 1997). Increased amounts of hyaluronan are produced by solid tumors and tumor-associated fibroblasts, and tumor-induced HA is correlated with poor prognosis. HA is well known to stimulate the migration of a large variety of cell types. Stimulation of cell migration by HA has been explained by different mechanisms. HA was shown to specifically bind to cell surface receptors, and inhibition of HA-receptor function was demonstrated to decrease cell migration and tumor growth. On the other hand, HA as a large hydrophilic molecule is also known to modulate the extracellular packing of collagen and fibrin, leading to increased fiber size and porosity of extracellular substrates. Hence a modified matrix architecture might similarly account for increased locomotion of cells. In this review, we attempted to summarize the available data on HA-induced cell migration, with particular emphasis on the role of HA receptors in three-dimensional cell migration. Although the HA receptor CD44 has been shown to mediate migration of cells over two-dimensional hyaluronan-coated surfaces in vitro, there is only little evidence that HA-binding to CD44 or other HA receptors has major impact on the locomotion of cells through three-dimensional matrices in vivo. We showed recently that the promigratory effect of HA in fibrin gels is largely due to HA-mediated modulation of fibrin polymerization. By increasing the porosity of fibrin gels, HA strongly accelerates cell migration. The porosity of matrices therefore appears as an important and probably underestimated determinant of cell migration and tumor spread.     

Are species of bacteria unclassifiable?

Summary Every one of eleven different strains randomly selected from 10 different randomly selected genera have shown the same high frequency of occurrence of colony mutants as did almost all strains ofAcetobacter (previously considered outstanding in this respect). Correlation of other properties with such mutant colony forms was not specifically studied, but in 4 strains correlation was noticed, suggesting its presence in the others, as was so often found inAcetobacter. It is suggested from this, that a similar study of strains of other genera might reveal a similarly high frequency of occurrence of mutants, most so-called pure cultures being thus probably mixtures of different cells with different properties. Also the proportion of each cell-type in the culture may vary from predominance to extinction according to the biochemical and other tests applied for the purpose of the ‘characterization’ of the species for taxonomic purposes. If the classification of such varying mixtures is considered of doubtful use, then it seems to follow that ‘species’ of bacteria are virtually unclassifiable, and that even the conception of a genus should be on a broader basis than is often the case at present.     

Are formal oxidation states above one viable in cyclopentadienylcopper cyanides?

Recent experiments have led to the discovery of the thermally unstable organocopper compounds (η(3)-C(3)H(5))CuMe(2), [(η(3)-C(3)H(5))CuMe(3)](-), and CuMe (4)(-) in which the copper atom is in the +3 formal oxidation state. In a quest for more stable organocopper compounds with copper in formal oxidation states above one, the binuclear cyclopentadienylcopper cyanides Cp(2)Cu(2)(CN)(n) (Cp = η(5)-C(5)H(5); n = 1, 2, 3) have been studied using density functional theory (DFT). The lowest energy structures are found to have terminal Cp rings and bridging cyanide ligands up to a maximum of two bridges. Higher-energy Cp(2)Cu(2)(CN)(n) (n = 1, 2, 3) structures are found with bridging Cp rings. The Cp(2)Cu(2)(CN)(3) derivatives, with the copper atoms in an average +2.5 oxidation state, are clearly thermodynamically disfavored with respect to cyanogen loss. However, Cp(2)Cu(2)(CN)(2) and Cp(2)Cu(2)(CN), with the copper atoms in the average oxidation states +1.5 and +2, respectively, are predicted to have marginal viability. The prospects for the copper(II) derivative Cp(2)Cu(2)(CN)(2) contrast with that of the "simple" Cu(CN)(2), which is shown both experimentally and theoretically to be unstable with respect to cyanogen loss to give CuCN.     

Some relationships between R-factor and chromosomal β-lactamase in Gram-negative bacteria

1. The beta-lactamases specified by Klebsiella aerogenes 418 and the R-factor R-7268 have been partially purified. 2. The molecular weights of the K. aerogenes strains 418 and 373, Aerobacter cloacae 53, R-7268 and R-TEM beta-lactamases were all about 20000; that of the enzymes from Escherichia coli strains 419 and 214T was about 31000. 3. These enzymes were also compared by means of their K(m) values for benzylpenicillin and ampicillin, and their behaviour on starch-gel electrophoresis. 4. The beta-lactamases specified by the two Klebsiella strains, the Aerobacter strain, and the R-factors R-TEM and R-7268 were found to comprise a broadly similar group. However, within this group, only two enzymes seemed to be identical, namely those specified by the two R-factors. The two E. coli strains produce identical beta-lactamases which are very different from the ;Klebsiella/Aerobacter-type' enzymes.     

Are the molecular strategies that control apoptosis conserved in bacteria?

The Staphylococcus aureus cid and lrg operons have been shown to encode putative membrane proteins that are involved in the regulation of murein hydrolase activity and penicillin tolerance. Cid proteins enhance murein hydrolase activity and penicillin sensitivity, whereas Lrg proteins have an inhibitory effect on these processes. It has been proposed that the Cid and Lrg proteins function in a way analogous to bacteriophage-encoded holins and antiholins, respectively, which control the timing of bacteriophage-induced lysis. This article explores the possibility that the Cid-Lrg regulatory system controls bacterial programmed cell death using a molecular strategy that it is functionally analogous to that mediated by the eukaryotic Bcl-2 family of apoptosis regulatory proteins.     

The viable but nonculturable phenotype: a crossroads in the life-cycle of non-differentiating bacteria?

In nature, prokaryotes must face alternating periods of prosperity and adversity. Differentiating bacteria confront situations of adversity by developing resistant structures. When there is a plentiful period, they adopt a vegetative state and when the period is adverse, a resistant structure, thereby completing a cycle. Non-differentiating bacteria do not develop such morphological distinct resistant structures. It has been proposed that many of these bacteria withstand periods of adversity by adopting the viable but nonculturable phenotype (VBNC). Bacteria of this phenotype conserve detectable metabolic function but become unculturable. Is it possible that the VBNC phenotype can revert to a culturable state, and vice versa, thus establishing a life-cycle? This review presents and evaluates different hypotheses regarding this question. Moreover, it attempts to analyse and proffer answers to other questions related to this phenotype. Is this a successful phenotype which prolongs survival? Is this a strategy for the survival of individual cells, or is it a strategy for the survival of a population? Finally, is it possible that this phenotype is, in fact, an example of altruistic death?     

Are bacteria an important food source for rotifers in eutrophic lakes?

In situ grazing measurements using fluorescent particles of0.5, 2.4 and 6.3 µm diameter in eutrophic Lake Loosdrecht(The Netherlands) showed that Anuraeopsis fissa, a small rotifer,filtered the smallest, bacteria-sized particles as efficientlyor more efficiently than the larger particles. In contrast,three other rotifer species (Brachionus angularis, Filinia longisetaand Pompholyx sulcata) filtered the bacteria-sized particlesless efficiently than the larger particles. Both Keratella cochlearisand Conochilus unicornis only ingested the bacteria-sized particles.Anuraeopsis fissa had a higher uptake of fluorescent bacteria-sizedparticles than K.cochlearis, both in 1 µm filtrate oflake water and in lake water. Within both species, uptake didnot differ between juveniles and adults. When cultured on threedifferent size fractions of lake water (1, 3 and 15 µmfiltrate) in July, all rotifer species declined in numbers onthe 1 and 3 µm filtrates, while A.fissa and B.angularisincreased in numbers on the 15 µm filtrate. The high abundanceof small bacteria in the lake water could not support rotiferpopulations. It is concluded that bacteria are not a suitablefood source of high quality for A.fissa because its populationdoes not grow even though the bacterial concentration was higherthan its estimated threshold food concentration. In August,when individually cultured, the mortality was high for all species,but especially for F.longiseta. The lifespan of K.cochleariswas reduced in the 1 and 3 µm filtrates of lake water,compared with in the 15 µm filtrate. The lifespan of A.fissawas similar in all filtrates, but reproduction was reduced inthe 1 and 3 µm filtrates, as in Keratella. On the 15 µmfiltrate, their ages at first reproduction and growth ratesdid not differ. Individuals of A.fissa older than 4 days showeda higher survival in the 15 µm filtrate than in the othertwo filtrates, as did K.cochlearis throughout its life. Hence,bacteria seem to be a more important food source for youngerindividuals of A.fissa than of K.cochlearis.     

Are intracellular bacteria hallmarks for human tumors?

正There are ten major characteristics that are used to differentiate cancer from normal cells and tissue (Hanahan and Weinberg, 2011); detection of bacterial species associated with cancer cells is not included among these characteristics,and at first glance this might seem to be irrelevant. The     

Are specific nonannular cholesterol binding sites present in G-protein coupled receptors?

The G-protein coupled receptors (GPCRs) are the largest class of molecules involved in signal transduction across membranes, and represent major drug targets in all clinical areas. Membrane cholesterol has been reported to have a modulatory role in the function of a number of GPCRs. Interestingly, recently reported crystal structures of GPCRs have shown structural evidence of cholesterol binding sites. Two possible mechanisms have been previously suggested by which membrane cholesterol could influence the structure and function of GPCRs (i) through a direct/specific interaction with GPCRs, which could induce a conformational change in the receptor, or (ii) through an indirect way by altering the membrane physical properties in which the receptor is embedded or due to a combination of both. We discuss here a novel mechanism by which membrane cholesterol could affect structure and function of GPCRs and propose that cholesterol binding sites in GPCRs could represent ‘nonannular’ binding sites. Interestingly, previous work from our laboratory has demonstrated that membrane cholesterol is required for the function of the serotonin_1A receptor, which could be due to specific interaction of the receptor with cholesterol. Based on these results, we envisage that there could be specific/nonannular cholesterol binding site(s) in the serotonin_1A receptor. We have analyzed putative cholesterol binding sites from protein databases in the serotonin_1A receptor, a representative GPCR, for which we have previously demonstrated specific requirement of membrane cholesterol for receptor function. Our analysis shows that cholesterol binding sites are inherent characteristic features of serotonin_1A receptors and are conserved over evolution. Progress in deciphering molecular details of the nature of GPCR-cholesterol interaction in the membrane would lead to better insight into our overall understanding of GPCR function in health and disease, thereby enhancing our ability to design better therapeutic strategies to combat diseases related to malfunctioning of GPCRs.     

Genome-wide association mapping in bacteria?

Bacteria display many interesting phenotypes such as virulence, tissue specificity and host range, for which it would be useful to know the genetic basis. Association mapping involves identifying causal variants by showing that particular genotypes are statistically associated with a phenotypic trait in a sample of strains taken from a natural population. With the advent of high-throughput genotyping, association mapping is becoming an increasingly powerful approach. However, until recently, association studies had not been used in bacteria because of their strong population structure, which can produce false positives and/or loss of statistical power unless elucidated and taken into account in analyses. Here, we describe how association mapping could be successfully applied to bacteria and outline the necessary sampling and genotyping strategies.     

Helicobacter pylori oriC��the first bipartite origin of chromosome replication in Gram-negative bacteria

Binding of the DnaA protein to oriC leads to DNA melting within the DNA unwinding element (DUE) and initiates replication of the bacterial chromosome. Helicobacter pylori oriC was previously identified as a region localized upstream of dnaA and containing a cluster of DnaA boxes bound by DnaA protein with a high affinity. However, no unwinding within the oriC sequence has been detected. Comprehensive in silico analysis presented in this work allowed us to identify an additional region (oriC2), separated from the original one (oriC1) by the dnaA gene. DnaA specifically binds both regions, but DnaA-dependent DNA unwinding occurs only within oriC2. Surprisingly, oriC2 is bound exclusively as supercoiled DNA, which directly shows the importance of the DNA topology in DnaA-oriC interactions, similarly as previously presented only for initiator-origin interactions in Archaea and some Eukaryota. We conclude that H. pylori oriC exhibits bipartite structure, being the first such origin discovered in a Gram-negative bacterium. The H. pylori mode of initiator-oriC interactions, with the loop formation between the subcomplexes of the discontinuous origin, resembles those discovered in Bacillus subtilis chromosome and in many plasmids, which might suggest a similar way of controlling initiation of replication.     

Corticotropin-releasing factor family and its receptors: tumor therapeutic targets?

Urocortin (UCN) and corticotropin-releasing factor (CRF) are members of CRF family. Though CRF is mainly distributed in central nervous system (CNS), UCN has been reported to play biologically diverse roles in several systems such as cardiovascular, respiratory, digestive, reproductive, stress, immunologic system, etc. UCN and CRF bind to two known receptors, CRFR1 and CRFR2, to function. Both CRF receptors are distributed in CNS and periphery tissues, and their expression in cancer tissues has been reported. Now there are many documents indicating UCN/CRF play an important role in the regulation of carcinogenesis. There is also evidence indicating UCN/CRF have anticancer effects via CRFRs. This paper will review the effects of CRF family in cancers.     

Are bacteria the third partner of theAzolla-Anabaena symbiosis?

The development of the prokaryotic colony inAzolla filiculoides indicates thatAnabaena azollae is maintained through the life cycle of the fern and present in the leaves and megasporocarps. The same biological pattern is applied to the bacteria that are also present in these structures and seems to follow a development pattern identical to the cyanobacteria and probably can be considered the third partner of this symbiotic association.     

Surface receptors: Are they involved in transformation of spermatozoa of Ascaris?

Exposure of the spheroidal spermatozoa of Ascaris suum to an extract of the male accessory gland causes their transformation into ameboid cells. We have investigated the mechanism of this transformation, also termed activation, by labeling the proteins of accessory gland extracts with fluorescein isothiocyanate (FITC) or [125I], followed by qualitative localization of the sperm activating substances (SAS) and quantitative measurements of [125I]-SAS binding. Fluorescent patches of FITC-conjugated SAS were localized at the spermatozoan surface and were concentrated primarily at the posterior region. Few fluorescent patches were detectable in the region of the newly formed pseudopodia following transformation. Although spermatozoan transformation occurs within 2-5 min after exposure to SAS, the fluorescent patches became more distinct after a minimum of 8 min and reached maximum density at 15-30 min. Spermatozoa activated with [125I]-SAS became radioactively labeled in direct proportion to the amount of available [125I]-SAS until a saturation level was reached. SDS-polyacrylamide gel electrophoresis combined with autoradiography indicated that the cells bind two SAS components, of small (9,000 MW) and large (56,000 MW) sizes. These same two components were also detectable in a membrane fraction, obtained by differential centrifugation, of the spermatozoa after incubation with [125I]-SAS. binding of the two SAS components was not inhibited by preincubation of the spermatozoa with trypsin or Concanavalin A; however, the 56,000 MW component of SAS was not detectable in autoradiograms of spermatozoa incubated with periodic acid (1.6-10 mM) treated SAS. Such cells also failed to transform into ameboid spermatozoa. These results indicate that the two components of SAS that bind to the spermatozoan surface are possibly responsible for inducing the cell transformations associated with activation.