Genome-wide prediction of human VNTRs

Polymorphic minisatellites, also known as variable number of tandem repeats (VNTRs), are tandem repeat regions that show variation in the number of repeat units among chromosomes in a population. Currently, there are no general methods for predicting which minisatellites have a high probability of being polymorphic, given their sequence characteristics. An earlier approach has focused on potentially highly polymorphic and hypervariable minisatellites, which make up only a small fraction of all minisatellites in the human genome. We have developed a model, based on available minisatellite and VNTR sequence data, that predicts the probability that a minisatellite (unit size > or = 6 bp) identified by the computer program Tandem Repeats Finder is polymorphic (VNTR). According to the model, minisatellites with high copy number and high degree of sequence similarity are most likely to be VNTRs. This approach was used to scan the draft sequence of the human genome for VNTRs. A total of 157,549 minisatellite repeats were found, of which 29,224 are predicted to be VNTRs. Contrary to previous results, VNTRs appear to be widespread and abundant throughout the human genome, with an estimated density of 9.1 VNTRs/Mb.     

Habitat management for tanks and Tuans: Evolving approaches at Puckapunyal Military Area

Through a combination of on-ground actions, policies and plans, this military training area – set within an important Box–Ironbark Ecosystem – is being managed in a manner that provides compatibility with the conservation of biodiversity.     

Experience-independent development of the hamster circadian visual system

Experience-dependent functional plasticity is a hallmark of the primary visual system, but it is not known if analogous mechanisms govern development of the circadian visual system. Here we investigated molecular, anatomical, and behavioral consequences of complete monocular light deprivation during extended intervals of postnatal development in Syrian hamsters. Hamsters were raised in constant darkness and opaque contact lenses were applied shortly after eye opening and prior to the introduction of a light-dark cycle. In adulthood, previously-occluded eyes were challenged with visual stimuli. Whereas image-formation and motion-detection were markedly impaired by monocular occlusion, neither entrainment to a light-dark cycle, nor phase-resetting responses to shifts in the light-dark cycle were affected by prior monocular deprivation. Cholera toxin-b subunit fluorescent tract-tracing revealed that in monocularly-deprived hamsters the density of fibers projecting from the retina to the suprachiasmatic nucleus (SCN) was comparable regardless of whether such fibers originated from occluded or exposed eyes. In addition, long-term monocular deprivation did not attenuate light-induced c-Fos expression in the SCN. Thus, in contrast to the thalamocortical projections of the primary visual system, retinohypothalamic projections terminating in the SCN develop into normal adult patterns and mediate circadian responses to light largely independent of light experience during development. The data identify a categorical difference in the requirement for light input during postnatal development between circadian and non-circadian visual systems.     

Therapeutic Efficacy of Stable Analogues of Vasoactive Intestinal Peptide against Pathogens

Vasoactive intestinal peptide (VIP) is an anti-inflammatory neuropeptide recently identified as a potential antimicrobial peptide. To overcome the metabolic limitations of VIP, we modified the native peptide sequence and generated two stable synthetic analogues (VIP51 and VIP51(6–30)) with better antimicrobial profiles. Herein we investigate the effects of both VIP analogues on cell viability, membrane integrity, and ultrastructure of various bacterial strains and Leishmania species. We found that the two VIP derivatives kill various non-pathogenic and pathogenic Gram-positive and Gram-negative bacteria as well as the parasite Leishmania major through a mechanism that depends on the interaction with certain components of the microbial surface, the formation of pores, and the disruption of the surface membrane. The cytotoxicity of the VIP derivatives is specific for pathogens, because they do not affect the viability of mammalian cells. Docking simulations indicate that the chemical changes made in the analogues are critical to increase their antimicrobial activities. Consequently, we found that the native VIP is less potent as an antibacterial and fails as a leishmanicidal. Noteworthy from a therapeutic point of view is that treatment with both derivatives increases the survival and reduces bacterial load and inflammation in mice with polymicrobial sepsis. Moreover, treatment with VIP51(6–30) is very effective at reducing lesion size and parasite burden in a model of cutaneous leishmaniasis. These results indicate that the VIP analogues emerge as attractive alternatives for treating drug-resistant infectious diseases and provide key insights into a rational design of novel agents against these pathogens.     

Creating Novel Activated Factor XI Inhibitors through Fragment Based Lead Generation and Structure Aided Drug Design

Activated factor XI (FXIa) inhibitors are anticipated to combine anticoagulant and profibrinolytic effects with a low bleeding risk. This motivated a structure aided fragment based lead generation campaign to create novel FXIa inhibitor leads. A virtual screen, based on docking experiments, was performed to generate a FXIa targeted fragment library for an NMR screen that resulted in the identification of fragments binding in the FXIa S1 binding pocket. The neutral 6-chloro-3,4-dihydro-1H-quinolin-2-one and the weakly basic quinolin-2-amine structures are novel FXIa P1 fragments. The expansion of these fragments towards the FXIa prime side binding sites was aided by solving the X-ray structures of reported FXIa inhibitors that we found to bind in the S1-S1’-S2’ FXIa binding pockets. Combining the X-ray structure information from the identified S1 binding 6-chloro-3,4-dihydro-1H-quinolin-2-one fragment and the S1-S1’-S2’ binding reference compounds enabled structure guided linking and expansion work to achieve one of the most potent and selective FXIa inhibitors reported to date, compound 13, with a FXIa IC₅₀ of 1.0 nM. The hydrophilicity and large polar surface area of the potent S1-S1’-S2’ binding FXIa inhibitors compromised permeability. Initial work to expand the 6-chloro-3,4-dihydro-1H-quinolin-2-one fragment towards the prime side to yield molecules with less hydrophilicity shows promise to afford potent, selective and orally bioavailable compounds.     

A micromanipulation method to measure the mechanical properties of single tomato suspension cells

A micromanipulation method has been developed to measure the force required to burst single tomato cells (Lycopersicon esculentum vf36) taken from suspension cultures. The method works by compressing a cell between parallel surfaces whilst measuring the force being imposed on the cell, and its deformation. The mean bursting force for two-week-old cells was 3.6 mN (standard error 0.1 mN), at a compression speed of 23 m s^–1. Usually force-deformation curves showed a single bursting event, but sometimes multiple bursts were observed, implying cells could reseal after failure. If cells were deformed without bursting, and then held, they showed a relaxation of the force. This was attributed to water loss, although wall relaxation was also a possibility. The half time of this relaxation was between 1–10 s. Tests on protoplasts gave bursting forces 1000 fold lower than intact cells, and cells treated with Triton to disrupt the membranes and destroy turgor collapsed with no bursting. As expected, both turgor and the presence of a wall were essential to maintaining cell strength.     

Social Jetlag: Misalignment of Biological and Social Time

Humans show large differences in the preferred timing of their sleep and activity. This so‐called “chronotype” is largely regulated by the circadian clock. Both genetic variations in clock genes and environmental influences contribute to the distribution of chronotypes in a given population, ranging from extreme early types to extreme late types with the majority falling between these extremes. Social (e.g., school and work) schedules interfere considerably with individual sleep preferences in the majority of the population. Late chronotypes show the largest differences in sleep timing between work and free days leading to a considerable sleep debt on work days, for which they compensate on free days. The discrepancy between work and free days, between social and biological time, can be described as ‘social jetlag.’ Here, we explore how sleep quality and psychological wellbeing are associated with individual chronotype and/or social jetlag. A total of 501 volunteers filled out the Munich ChronoType Questionnaire (MCTQ) as well as additional questionnaires on: (i) sleep quality (SF‐A), (ii) current psychological wellbeing (Basler Befindlichkeitsbogen), (iii) retrospective psychological wellbeing over the past week (POMS), and (iv) consumption of stimulants (e.g., caffeine, nicotine, and alcohol). Associations of chronotype, wellbeing, and stimulant consumption are strongest in teenagers and young adults up to age 25 yrs. The most striking correlation exists between chronotype and smoking, which is significantly higher in late chronotypes of all ages (except for those in retirement). We show these correlations are most probably a consequence of social jetlag, i.e., the discrepancies between social and biological timing rather than a simple association to different chronotypes. Our results strongly suggest that work (and school) schedules should be adapted to chronotype whenever possible.