CDR1as regulated by hnRNPM maintains stemness of periodontal ligament stem cells via miR-7/KLF4

CDR1as is a well-identified circular RNA with regulatory roles in a variety of physiological processes. However, the effects of CDR1as on stemness of periodontal ligament stem cells (PDLSCs) and the underlying mechanisms remain unclear. In this study, we detect CDR1as in human PDLSCs, and subsequently demonstrate that CDR1as maintains PDLSC stemness. Knockdown of CDR1as decreases the expression levels of stemness-related genes and impairs the cell's multi-differentiation and cell migration abilities, while overexpression of CDR1as increases the expression levels of stemness-related genes and enhances these abilities. Furthermore, our results indicate that the RNA-binding protein hnRNPM directly interacts with CDR1as and regulates its expression in PDLSCs. In addition, we show that CDR1as promotes the expression of stemness-related genes in PDLSCs by inhibiting miR-7-mediated suppression of KLF4 expression. Collectively, our results demonstrate that CDR1as participates in the molecular circuitry that regulates PDLSC stemness.     

Stage-specific secretion of HMGB1 in cartilage regulates endochondral ossification

High mobility group box 1 protein (HMGB1) is a chromatin protein that has a dual function as a nuclear factor and as an extracellular factor. Extracellular HMGB1 released by damaged cells acts as a chemoattractant, as well as a proinflammatory cytokine, suggesting that HMGB1 is tightly connected to the process of tissue organization. However, the role of HMGB1 in bone and cartilage that undergo remodeling during embryogenesis, tissue repair, and disease is largely unknown. We show here that the stage-specific secretion of HMGB1 in cartilage regulates endochondral ossification. We analyzed the skeletal development of Hmgb1(-/-) mice during embryogenesis and found that endochondral ossification is significantly impaired due to the delay of cartilage invasion by osteoclasts, osteoblasts, and blood vessels. Immunohistochemical analysis revealed that HMGB1 protein accumulated in the cytosol of hypertrophic chondrocytes at growth plates, and its extracellular release from the chondrocytes was verified by organ culture. Furthermore, we demonstrated that the chondrocyte-secreted HMGB1 functions as a chemoattractant for osteoclasts and osteoblasts, as well as for endothelial cells, further supporting the conclusion that Hmgb1(-/-) mice are defective in cell invasion. Collectively, these findings suggest that HMGB1 released from differentiating chondrocytes acts, at least in part, as a regulator of endochondral ossification during osteogenesis.     

Golgins and GRASPs: Holding the Golgi together

The GRASP and golgin families of proteins have emerged as key components of the Golgi apparatus, with major roles in both the structural organisation of this organelle and the trafficking that occurs there. Both types of protein participate in membrane tethering events that occur upstream of membrane fusion as well as contributing to the structural scaffold that defines Golgi architecture, referred to as the Golgi matrix. The importance of these proteins is highlighted by their targeting in mitosis, apoptosis, and pathogenic infections that cause dramatic structural and functional reorganisation of the Golgi apparatus. In this review we will discuss our current understanding of GRASP and golgin function, highlighting some of the common themes that have emerged as well as describing previously unsuspected roles for these proteins in various cellular processes.     

DNA as a Phosphate Storage Polymer and the Alternative Advantages of Polyploidy for Growth or Survival

Haloferax volcanii uses extracellular DNA as a source for carbon, nitrogen, and phosphorous. However, it can also grow to a limited extend in the absence of added phosphorous, indicating that it contains an intracellular phosphate storage molecule. As Hfx. volcanii is polyploid, it was investigated whether DNA might be used as storage polymer, in addition to its role as genetic material. It could be verified that during phosphate starvation cells multiply by distributing as well as by degrading their chromosomes. In contrast, the number of ribosomes stayed constant, revealing that ribosomes are distributed to descendant cells, but not degraded. These results suggest that the phosphate of phosphate-containing biomolecules (other than DNA and RNA) originates from that stored in DNA, not in rRNA. Adding phosphate to chromosome depleted cells rapidly restores polyploidy. Quantification of desiccation survival of cells with different ploidy levels showed that under phosphate starvation Hfx. volcanii diminishes genetic advantages of polyploidy in favor of cell multiplication. The consequences of the usage of genomic DNA as phosphate storage polymer are discussed as well as the hypothesis that DNA might have initially evolved in evolution as a storage polymer, and the various genetic benefits evolved later.     

Brain death and brain life: rethinking the connection

The connection between brain life and brain death is neither as simple nor as defensible as it might at first appear. The problem rests with the two dominant competing definitions of death:...the loss of that which is necessary for the organism to continue to function as a whole;....the loss of that which is essentially significant to the nature of the organism... If death is understood as the loss of that which is necessary for the continued functioning of the organism as whole, then the apparent symmetry breaks down. If...death could be understood as the loss of that which is essentially significant to the nature of the organism....consciousness, then the symmetry would hold. However, that definition of death is indefensible. Therefore...statements about the status of anencephalic infants and early human embryos based upon a connection between brain death and brain life are unfounded.     

Frozen embryos and the obligation to adopt

Rob Lovering has developed an interesting new critique of views that regard embryos as equally valuable as other human beings: the moral argument for frozen human embryo adoption. The argument is aimed at those who believe that the death of a frozen embryo is a very bad thing, and Lovering concludes that some who hold this view ought to prevent one of these deaths by adopting and gestating a frozen embryo. Contra Lovering, we show that there are far more effective strategies for preserving the lives of frozen embryos than adoption. Moreover, we point out that those who regard the deaths of frozen embryos as a very bad thing will generally regard the deaths of all embryos as a very bad thing, whether they are discarded embryos, aborted embryos or embryos that spontaneously abort. This entails that these other embryos must be taken into account when considering moral obligations, as well as other human lives at risk from preventable causes.     

The effect of ejaculate mass on female reproductive output in the european swallowtail butterfly,Papilio machaon (L.) (Lepidoptera: Papilionidae)

Previously, we showed that virgin males of Papilio machaondeliver ejaculates that are twice as big as any ejaculates they transfer at later matings. Here, we investigate the consequences of these two size classes of ejaculates on female reproductive output and demonstrate that females that received one small ejaculate laid as many eggs, fertilized the same proportion of eggs, and lived as long as those that had received one big ejaculate. However, females that received big ejaculates laid heavier eggs, but only between the twelfth and the fifteenth days of egg-laying. We conclude that male-derived nutrients appear to have a limited effect on female reproductive output in P. machaonand that the large size of ejaculates delivered by male butterflies are determined primarily by selection on males to produce longer refractory periods in females.     

Biodiversity, biopiracy and benefits: what allegations of biopiracy tell us about intellectual property

This paper examines the concept of biopiracy, which initially emerged to challenge various aspects of the regime for intellectual property rights (IPR) in living organisms, as well as related aspects pertaining to the ownership and apportioning of benefits from 'genetic resources' derived from the world's biodiversity. This paper proposes that we take the allegation of biopiracy seriously due to the impact it has as an intervention which indexes a number of different, yet interrelated, problematizations of biodiversity, biotechnology and IPR. Using the neem tree case as an example, it describes activists' use of the term as one that involves a deliberate simplification of science and IPR. Additionally, it argues that in so doing, biopiracy is positioned as a touchstone that mobilizes actors and problems, and ultimately generates 'solutions' to the very challenges it creates. The paper will also encourage a view of biopiracy claims that does not always treat them simply as claims of theft, or as a misallocation of benefits, but rather as claims that are designed to raise broader questions about the IPR system itself. It concludes by advocating that, in order to properly understand how to address biopiracy, we must be prepared to move beyond the current narrow readings to develop a more complete picture of the term's influence in challenging how, and by whom, the decisions about what is natural and what is invented come to be made.     

Functional imaging of microdomains in cell membranes

The presence of microdomains or rafts within cell membranes is a topic of intense study and debate. The role of these structures in cell physiology, however, is also not yet fully understood with many outstanding problems. This problem is partly based on the small size of raft structures that presents significant problems to their in vivo study, i.e., within live cell membranes. But the structure and dynamics as well as the factors that control the assembly and disassembly of rafts are also of major interest. In this review we outline some of the problems that the study of rafts in cell membranes present as well as describing some views of what are considered the generalised functions of membrane rafts. We point to the possibility that there may be several different ‘types’ of membrane raft in cell membranes and consider the factors that affect raft assembly and disassembly, particularly, as some researchers suggest that the lifetimes of rafts in cell membranes may be sub-second. We attempt to review some of the methods that offer the ability to interrogate rafts directly as well as describing factors that appear to affect their functionality. The former include both near-field and far-field optical approaches as well as scanning probe techniques. Some of the advantages and disadvantages of these techniques are outlined. Finally, we describe our own views of raft functionality and properties, particularly, concerning the membrane dipole potential, and describe briefly some of the imaging strategies we have developed for their study.     

Metaphors of glaucoma

In a phenomenological study of glaucoma as an illness, 31 patients were interviewed. The goal of the study was an explication of the meanings of the experience of glaucoma, in a twofold sense: (1) the face, the role, or the self of the patient that responds to the illness; (2) the visage that the illness presents to the patient. This twofold explication was achieved by generating a classification of the interviews in terms of the dominant metaphor that emerged in each interview. The following basic metaphors are presented through case material: glaucoma as an accompaniment of aging, as blindness, as pressure, as an abstraction (and a weapon), and as fate. Variations on these themes demonstrate the necessity, in such work and for enhanced communication with patients, of paying attention to both the face that the illness presents and to the self that responds.     

单菌落PCR法直接快速鉴定重组克隆

利用单菌落PCR法直接筛选含有GFP、LTB-ST外源基因的重组克隆,阳性克隆可以扩增出目的条带,和质粒PCR扩增的结果一致。同时,单菌落PCR法也可应用于重组质粒转化后的农杆菌的筛选,单菌落PCR法的扩增结果和农杆菌液扩增的结果一致。结果表明,单菌落PCR法是一个有效简便的鉴定重组阳性克隆的方法。     

Lessons from Cryptococcal Laccase: From Environmental Saprophyte to Pathogen

Cryptococcus is an opportunistic pathogen that lives in the environment as a free-living yeast and inflicts disease in humans, primarily in immunocompromised patients such as organ-transplant recipients and people with HIV/AIDS. A key factor allowing emergence of this fungal pathogen is a copper-containing laccase enzyme that facilitates nutrient foraging as a saprophyte and offers protection against environmental dangers such as free-living amoebae and mammalian macrophages during infection. The promiscuous substrate specificity of laccase facilitates the degradation of hard plant lignin polymers as well as the synthesis of lignin-like pigments from plant flavonoids and melanin pigments from dopamine. The enzyme also possesses an iron oxidase activity that prevents Fenton product formation in macrophages and another activity that allows synthesis of immune-modulatory prostaglandins that fool the host, shutting off an effective immune response. This review provides a brief overview of key points in laccase function and its role in virulence, as well as regulation and trafficking of the enzyme during the interconversion between saprophyte and pathogen, yielding insights into pathogenesis from this adaptable pathogen.     

Beta-alanine but not taurine can function as an organic osmolyte in preimplantation mouse embryos cultured from fertilized eggs

Early preimplantation mouse embryos are susceptible to the detrimental effects of increased osmolarity and, paradoxically, their in vitro development is significantly compromised by osmolarities near that of oviductal fluid. In vitro development can be restored, however, by several compounds that are accumulated by 1-cell embryos to act as organic osmolytes, providing intracellular osmotic support and cell volume regulation. Taurine, a substrate of the beta-amino acid transporter that functions as an organic osmolyte transporter in other cells, had been proposed to function as an organic osmolyte in mouse embryos. Here, however, we found that taurine is neither able to provide protection for in vitro embryo development against increased osmolarity nor is it accumulated to higher intracellular levels as osmolarity is increased, indicating that it cannot function as an organic osmolyte in early preimplantation embryos. In contrast, beta-alanine, the other major substrate of the beta-amino acid transporter, both protects against increased osmolarity and is accumulated to somewhat higher levels as osmolarity is increased, indicating that it is able to function as an organic osmolyte in embryos. However, we also found that beta-alanine is displaced from embryos by glycine-the most effective organic osmolyte in embryos previously identified-and beta-alanine does not increase protection above that afforded by glycine at concentrations near those in vivo. Thus, the beta-amino acid transporter is likely present in early preimplantation embryos to supply beta-amino acids such as taurine for purposes other than to serve as organic osmolytes.     

Oscillations in continuous culture populations of Streptococcus pneumoniae: population dynamics and the evolution of clonal suicide

Agents that kill or induce suicide in the organisms that produce them or other individuals of the same genotype are intriguing puzzles for ecologists and evolutionary biologists. When those organisms are pathogenic bacteria, these suicidal toxins have the added appeal as candidates for the development of narrow spectrum antibiotics to kill the pathogens that produce them. We show that when clinical as well as laboratory strains of Streptococcus pneumoniae are maintained in continuous culture (chemostats), their densities oscillate by as much as five orders of magnitude with an apparently constant period. This dynamic, which is unanticipated for single clones of bacteria in chemostats, can be attributed to population-wide die-offs and recoveries. Using a combination of mathematical models and experiments with S. pneumoniae, we present evidence that these die-offs can be attributed to the autocatalytic production of a toxin that lyses or induces autolysis in members of the clone that produces it. This toxin, which our evidence indicates is a protein, appears to be novel; S. pneumoniae genetic constructs knocked out for lytA and other genes coding for known candidates for this agent oscillate in chemostat culture. Since this toxin lyses different strains of S. pneumoniae as well as other closely related species of Streptococcus, we propose that its ecological role is as an allelopathic agent. Using a mathematical model, we explore the conditions under which toxins that kill members of the same clone that produces them can prevent established populations from invasion by different strains of the same or other species. We postulate that the production of the toxin observed here as well as other bacteria-produced toxins that kill members of the same genotype, ‘clonal suicide’, evolved and are maintained to prevent colonization of established populations by different strains of the same and closely related species.     

Locked into security,keyed into modernity: The selection of burglaries as source of risk in Greece

Consensus among Greeks claims that the rates of burglaries and thefts have been increasing during the last ten years and that this constitutes a major social problem for which immigrants, and, especially Albanians, are responsible. This article theorizes that the selection of burglaries as source of risk deploys the understanding that the flip side of the increase in property crime is increased affluence and that both are part of modernization. The selection of houses as targets of aggression enables people, especially men, to construe themselves as protectors of houses objectifying the longstanding ideals of household management and as members of a modern society in which security is a matter of individual consumer choice. The portrayal of immigrants as destitute thieves reinforces the construct of the Greek nation state as a successful albeit vulnerable household writ-large.     

Computational simulation of the statistical properties of unfolded proteins

A simple Monte Carlo method was used to generate ensembles of simulated polypeptide conformations that are restricted only by steric repulsion. The models used for these simulations were based on the sequences of four real proteins, ranging in size from 26 to 268 amino acid residues, and included all non-hydrogen atoms. Two sets of calculations were performed, one that included only intra-residue steric repulsion terms and those between adjacent residues, and one that included repulsion terms between all possible atom pairs, so as to explicitly account for the excluded volume effect. Excluded volume was found to increase the average radius of gyration of the chains by 20-40%, with the expansion factor increasing with chain length. Contrary to recent suggestions, however, the excluded volume effect did not greatly restrict the distribution of dihedral angles or favor native-like topologies. The average dimensions of the ensembles calculated with excluded volume were consistent with those measured experimentally for unfolded proteins of similar sizes under denaturing conditions, without introducing any adjustable scaling factor. The simulations also reproduced experimentally determined effective concentrations for the formation of disulfide bonds in reduced and unfolded proteins. The statistically generated ensembles included significant numbers of conformations that were nearly as compact as the corresponding native proteins, as well as many that were as accessible to solvent as a fully extended chain. On the other hand, conformations with as much buried surface area as the native proteins were very rare, as were highly extended conformations. These results suggest that the overall properties of unfolded proteins can be usefully described by a random coil model and that an unfolded polypeptide can undergo significant collapse while losing only a relatively small fraction of its conformational entropy.