Global effects of land use on local terrestrial biodiversity.

Human activities, especially conversion and degradation of habitats, are causing global biodiversity declines. How local ecological assemblages are responding is less clear--a concern given their importance for many ecosystem functions and services. We analysed a terrestrial assemblage database of unprecedented geographic and taxonomic coverage to quantify local biodiversity responses to land use and related changes. Here we show that in the worst-affected habitats, these pressures reduce within-sample species richness by an average of 76.5%, total abundance by 39.5% and rarefaction-based richness by 40.3%. We estimate that, globally, these pressures have already slightly reduced average within-sample richness (by 13.6%), total abundance (10.7%) and rarefaction-based richness (8.1%), with changes showing marked spatial variation. Rapid further losses are predicted under a business-as-usual land-use scenario; within-sample richness is projected to fall by a further 3.4% globally by 2100, with losses concentrated in biodiverse but economically poor countries. Strong mitigation can deliver much more positive biodiversity changes (up to a 1.9% average increase) that are less strongly related to countries' socioeconomic status.

Viviparity does not affect the numbers and sizes of reptile offspring.

Viviparity (live-bearing) has independently evolved from oviparity (egg-laying) in more than 100 lineages of squamates (lizards and snakes). We might expect consequent shifts in selective forces to affect per-brood reproductive investment (RI = total mass of offspring relative to maternal mass) and in the way in which that output is partitioned (number vs. size of offspring per brood). Based on the assumption that newly born offspring are heavier than eggs, we predicted that live-bearing must entail either increased RI or a reduction in offspring size and/or fecundity. However, our phylogenetically controlled analysis of data on 1,259 squamate species revealed no significant differences in mean offspring size, clutch size or RI between oviparous and viviparous squamates. We attribute this paradoxical result to (1) strong selection on offspring sizes, unaffected by parity mode, (2) the lack of a larval stage in amniotes, favouring large eggs even in the ancestral oviparous mode and (3) the ability of viviparous females to decrease the mass of uterine embryos by reducing extra-embryonic water stores. Our analysis shows that squamate eggs (when laid) weigh about the same as the hatchlings that emerge from them (despite a many-fold increase in embryo mass during incubation). Most of the egg mass is due to components (such as water stores and the eggshell) not required for oviductal incubation. That repackaging enables live-born offspring to be accommodated within the mother's body without increasing total litter mass. The consequent stasis in reproductive burden during the evolutionary transition from oviparity to viviparity may have facilitated frequent shifts in parity modes.

Spatial-numerical association of response code effect as a window to mental representation of magnitude in long-term memory among Hebrew-speaking children.

The well-known spatial-numerical association of response code (SNARC) effect supports the idea that the mental number line (MNL) is organized from left to right in participants writing from left to right. In Arabic speakers writing from right to left, the direction of the SNARC effect is reversed. Until recently, no consistent numerical-spatial associations were reported in Hebrew speakers, who write letters from right to left and write numbers from left to right. However, a left-to-right SNARC effect was recently demonstrated in adult Hebrew readers by reducing the markedness association of response code (MARC) effect, which masks the SNARC effect. Adult Hebrew speakers (especially university students) are skilled English readers and writers, supporting the claim that the direction of reading is not the sole factor contributing to direction of the emergence of the left-to-right organization of the MNL. Thus, to understand the effect of reading habits on the SNARC effect, here we demonstrate the SNARC in young Hebrew-speaking children who read Hebrew letters from right to left, read numbers from left to right, and had little experience in writing English letters from left to right. Our findings, therefore, are innovative in providing supporting evidence for the claim that the direction of reading is not the sole factor contributing to direction of the emergence of the left-to-right organization of the MNL.

TAT-MTS-MCM fusion proteins reduce MMA levels and improve mitochondrial activity and liver function in MCM-deficient cells.

Methylmalonic aciduria (MMA) is a disorder of organic acid metabolism resulting from a functional defect of the mitochondrial enzyme, methylmalonyl-CoA mutase (MCM). The main treatments for MMA patients are dietary restriction of propiogenic amino acids and carnitine supplementation. Liver or combined liver/kidney transplantation has been used to treat those with the most severe clinical manifestations. Thus, therapies are necessary to help improve quality of life and prevent liver, renal and neurological complications. Previously, we successfully used the TAT-MTS-Protein approach for replacing a number of mitochondrial-mutated proteins. In this targeted system, TAT, an 11 a.a peptide, which rapidly and efficiently can cross biological membranes, is fused to a mitochondrial targeting sequence (MTS), followed by the mitochondrial mature protein which sends the protein into the mitochondria. In the mitochondria, the TAT-MTS is cleaved off and the native protein integrates into its natural complexes and is fully functional. In this study, we used heterologous MTSs of human, nuclear-encoded mitochondrial proteins, to target the human MCM protein into the mitochondria. All fusion proteins reached the mitochondria and successfully underwent processing. Treatment of MMA patient fibroblasts with these fusion proteins restored mitochondrial activity such as ATP production, mitochondrial membrane potential and oxygen consumption, indicating the importance of mitochondrial function in this disease. Treatment with the fusion proteins enhanced cell viability and most importantly reduced MMA levels. Treatment also enhanced albumin and urea secretion in a CRISPR/Cas9-engineered HepG2 MUT (-/-) liver cell line. Therefore, we suggest using this TAT-MTS-Protein approach for the treatment of MMA.

Frataxin-deficient neurons and mice models of Friedreich ataxia are improved by TAT-MTScs-FXN treatment.

Friedreich ataxia (FA) is a rare disease caused by deficiency of frataxin, a mitochondrial protein. As there is no cure available for this disease, many strategies have been developed to reduce the deleterious effects of such deficiency. One of these approaches is based on delivering frataxin to the tissues by coupling the protein to trans-activator of transcription (TAT) peptides, which enables cell membranes crossing. In this study, we tested the efficiency of TAT-MTScs-FXN fusion protein to decrease neurodegeneration markers on frataxin-depleted neurons obtained from dorsal root ganglia (DRG), one of the most affected tissues. In mice models of the disease, we tested the ability of TAT-MTScs-FXN to penetrate the mitochondria and its effect on lifespan. In DRG neurons, treatment with TAT-MTScs-FXN increased cell survival, decreased neurite degeneration and reduced apoptotic markers, such as α-fodrin cleavage and caspase 9 activation. Also, we show that heat-shock protein 60 (HSP60), a molecular chaperone targeted to mitochondria, suffered an impaired processing in frataxin-deficient neurons that was relieved by TAT-MTScs-FXN addition. In mice models of the disease, administration of TAT-MTScs-FXN was able to reach muscle mitochondria, restore the activity of the succinate dehydrogenase and produce a significant lifespan increase. These results support the use of TAT-MTScs-FXN as a treatment for Friedreich ataxia.

Development of the Mental Number Line Representation of Numbers 0-10 and Its Relationship to Mental Arithmetic.

The internal representation of numbers on the mental number line (MNL) was demonstrated by performing the computerized version of the number-to-position (CNP) task on a touchscreen while restricting response time. We found that the estimation pattern is best fit by a sigmoid function, further denoted as the "sigmoidal model". Two developmental leaps occurring during elementary school were recognized: (1) the division of the number line into two segments and (2) consistent use of different anchor points on the number line-the left endpoint in first grade, the right endpoint in second grade, and finally the midpoint in third grade. Additionally, when examining the differences between the breakpoints, we found that first graders demonstrated a breakpoint close to 6, which linearly decreased over the years until stabilizing close to 5. The relation between the ability to place individual numbers on a number line and performance of mental arithmetic showed that the consistent use of anchor points correlated significantly with faster responses in mental arithmetic.

Davunetide (NAP) as a preventative treatment for central nervous system complications in a diabetes rat model.

AIMS: Central nervous system complications including cognitive impairment are an early manifestation of diabetes mellitus, also evident in animal models. NAP (generic name, davunetide), a neuroprotective peptide was tested here for its ability to prevent diabetes-related brain pathologies in the streptozotocin injected diabetes rat model. METHODS: Diabetes was induced by an intraperitoneal streptozotocin injection (55 mg/kg). Intranasal NAP or vehicle was administered daily starting on the day following streptozotocin injection. Cognitive assessment was performed 12 weeks after diabetes induction, using the Morris water maze paradigm. Brain structural integrity was assessed on the 15th week of diabetes by magnetic resonance T2 scan. Characterization of cellular populations, apoptosis and synaptic density was performed 16 weeks after diabetes induction, using immunohistochemical markers and quantified in the prefrontal cortex, the cerebral cortex and the hippocampus of both hemispheres. RESULTS: Impaired spatial memory of the diabetic rats was observed in the water maze by attenuated learning curve and worsened performance in the probe memory test. NAP treatment significantly improved both measurements. T2 magnetic resonance imaging revealed atrophy in the prefrontal cortex of the diabetes rat group, which was prevented by NAP treatment. Immunohistochemical analysis showed that NAP treatment protected against major loss of the synaptic marker synaptophysin and astrocytic apoptosis, resulting from streptozotocin treatment. CONCLUSIONS: Our results show for the first time protective effects for NAP (davuentide) in a diabetes rat model at the behavioral and structural levels against one of the most severe complications of diabetes.

Crystal structure of a novel viral protease with a serine/lysine catalytic dyad mechanism.

The blotched snakehead virus (BSNV), an aquatic birnavirus, encodes a polyprotein (NH2-pVP2-X-VP4-VP3-COOH) that is processed through the proteolytic activity of its own protease (VP4) to liberate itself and the viral proteins pVP2, X and VP3. The protein pVP2 is further processed by VP4 to give rise to the capsid protein VP2 and four structural peptides. We report here the crystal structure of a VP4 protease from BSNV, which displays a catalytic serine/lysine dyad in its active site. This is the first crystal structure of a birnavirus protease and the first crystal structure of a viral protease that utilizes a lysine general base in its catalytic mechanism. The topology of the VP4 substrate binding site is consistent with the enzymes substrate specificity and a nucleophilic attack from the si-face of the substrates scissile bond. Despite low levels of sequence identity, VP4 shows similarities in its active site to other characterized Ser/Lys proteases such as signal peptidase, LexA protease and Lon protease. Together, the structure of VP4 provides insights into the mechanism of a recently characterized clan of serine proteases that utilize a lysine general base and reveals the structure of potential targets for antiviral therapy, especially for other related and economically important viruses, such as infectious bursal disease virus in poultry and infectious pancreatic necrosis virus in aquaculture.

The global distribution of tetrapods reveals a need for targeted reptile conservation.

The distributions of amphibians, birds and mammals have underpinned global and local conservation priorities, and have been fundamental to our understanding of the determinants of global biodiversity. In contrast, the global distributions of reptiles, representing a third of terrestrial vertebrate diversity, have been unavailable. This prevented the incorporation of reptiles into conservation planning and biased our understanding of the underlying processes governing global vertebrate biodiversity. Here, we present and analyse the global distribution of 10,064 reptile species (99% of extant terrestrial species). We show that richness patterns of the other three tetrapod classes are good spatial surrogates for species richness of all reptiles combined and of snakes, but characterize diversity patterns of lizards and turtles poorly. Hotspots of total and endemic lizard richness overlap very little with those of other taxa. Moreover, existing protected areas, sites of biodiversity significance and global conservation schemes represent birds and mammals better than reptiles. We show that additional conservation actions are needed to effectively protect reptiles, particularly lizards and turtles. Adding reptile knowledge to a global complementarity conservation priority scheme identifies many locations that consequently become important. Notably, investing resources in some of the world's arid, grassland and savannah habitats might be necessary to represent all terrestrial vertebrates efficiently.

Publisher Correction: The global distribution of tetrapods reveals a need for targeted reptile conservation.

In this Article originally published, owing to a technical error, the author 'Laurent Chirio' was mistakenly designated as a corresponding author in the HTML version, the PDF was correct. This error has now been corrected in the HTML version. Further, in Supplementary Table 3, the authors misspelt the surname of 'Danny Meirte'; this file has now been replaced.

Expression, purification and crystallization of a birnavirus-encoded protease, VP4, from blotched snakehead virus (BSNV).

Blotched snakehead virus (BSNV) is a member of the Birnaviridae family that requires a virally encoded protease known as VP4 in order to process its polyprotein into viral capsid protein precursors (pVP2 and VP3). VP4 belongs to a family of serine proteases that utilize a serine/lysine catalytic dyad mechanism. A mutant construct of VP4 with a short C-terminal truncation was overexpressed in Escherichia coli and purified to homogeneity for crystallization. Using the sitting-drop vapour-diffusion method at room temperature, protein crystals with two distinct morphologies were observed. Cubic crystals grown in PEG 2000 MME and magnesium acetate at pH 8.5 belong to space group I23, with unit-cell parameters a = b = c = 143.8 angstroms. Trigonal crystals grown in ammonium sulfate and glycerol at pH 8.5 belong to space group P321/P312, with unit-cell parameters a = b = 158.2, c = 126.4 angstroms.

Purification, crystallization and preliminary X-ray analysis of truncated and mutant forms of VP4 protease from infectious pancreatic necrosis virus.

In viruses belonging to the Birnaviridae family, virus protein 4 (VP4) is the viral protease responsible for the proteolytic maturation of the polyprotein encoding the major capsid proteins (VP2 and VP3). Infectious pancreatic necrosis virus (IPNV), the prototype of the aquabirnavirus genus, is the causative agent of a contagious disease in fish which has a large economic impact on aquaculture. IPNV VP4 is a 226-residue (24.0 kDa) serine protease that utilizes a Ser/Lys catalytic dyad mechanism (Ser633 and Lys674). Several truncated and mutant forms of VP4 were expressed in a recombinant expression system, purified and screened for crystallization. Two different crystal forms diffract beyond 2.4 A resolution. A triclinic crystal derived from one mutant construct has unit-cell parameters a = 41.7, b = 69.6, c = 191.6 A, alpha = 93.0, beta = 95.1, gamma = 97.7 degrees. A hexagonal crystal with space group P6(1)22/P6(5)22 derived from another mutant construct has unit-cell parameters a = 77.4, b = 77.4, c = 136.9 A.

A general approach for the use of oligonucleotide effectors to regulate the catalysis of RNA-cleaving ribozymes and DNAzymes.

A general approach is described for controlling the RNA-cleaving activity of nucleic acid enzymes (ribozymes and DNAzymes) via the use of oligonucleotide effectors (regulators). In contrast to the previously developed approaches of allosteric and facilitator-mediated regulation of such enzymes, this approach, called 'expansive' regulation, requires that the regulator bind simultaneously to both enzyme and substrate to form a branched three-way complex. Such three-way enzyme-substrate-regulator complexes are catalytically competent relative to the structurally unstable enzyme-substrate complexes. Using the 8-17 and bipartite DNAzymes and the hammerhead ribozyme as model systems, 20- to 30-fold rate enhancements were achieved in the presence of regulators of engineered variants of the above three enzymes, even under unoptimized conditions. Broadly, using this approach ribozyme and DNAzyme variants that are amenable to regulation by oligonucleotide effectors can be designed even in the absence of any knowledge of the folded structure of the relevant ribozyme or DNAzyme. Expansive regulation therefore represents a new and potentially useful technology for both the regulation of nucleic acid enzymes and the detection of specific RNA transcripts.

Erratum: Sex determination, longevity, and the birth and death of reptilian species.

[This corrects the article DOI: 10.1002/ece3.2277.].

Sex determination, longevity, and the birth and death of reptilian species.

Vertebrate sex-determining mechanisms (SDMs) are triggered by the genotype (GSD), by temperature (TSD), or occasionally, by both. The causes and consequences of SDM diversity remain enigmatic. Theory predicts SDM effects on species diversification, and life-span effects on SDM evolutionary turnover. Yet, evidence is conflicting in clades with labile SDMs, such as reptiles. Here, we investigate whether SDM is associated with diversification in turtles and lizards, and whether alterative factors, such as lifespan's effect on transition rates, could explain the relative prevalence of SDMs in turtles and lizards (including and excluding snakes). We assembled a comprehensive dataset of SDM states for squamates and turtles and leveraged large phylogenies for these two groups. We found no evidence that SDMs affect turtle, squamate, or lizard diversification. However, SDM transition rates differ between groups. In lizards TSD-to-GSD surpass GSD-to-TSD transitions, explaining the predominance of GSD lizards in nature. SDM transitions are fewer in turtles and the rates are similar to each other (TSD-to-GSD equals GSD-to-TSD), which, coupled with TSD ancestry, could explain TSD's predominance in turtles. These contrasting patterns can be explained by differences in life history. Namely, our data support the notion that in general, shorter lizard lifespan renders TSD detrimental favoring GSD evolution in squamates, whereas turtle longevity permits TSD retention. Thus, based on the macro-evolutionary evidence we uncovered, we hypothesize that turtles and lizards followed different evolutionary trajectories with respect to SDM, likely mediated by differences in lifespan. Combined, our findings revealed a complex evolutionary interplay between SDMs and life histories that warrants further research that should make use of expanded datasets on unexamined taxa to enable more conclusive analyses.

D-SAL and NAP: Two Peptides Sharing a SIP Domain.

NAPVSIPQ (NAP) and all D-amino acid SALLRSIPA (D-SAL) are neuroprotective peptides derived from activity-dependent neuroprotective protein (ADNP) and activity-dependent neurotrophic factor (ADNF), respectively. Both proteins were shown to protect against cognitive impairment, using different animal models and to increase neuronal survival following exposure to neurotoxins. NAP was extensively tested and found to increase microtubule stability, protect axonal transport, and inhibit apoptosis. Here, we aimed to further evaluate and correlate effects at the behavioral level, in a rat model of diabetes. Diabetes is primarily a metabolic disorder which presents secondary neurological manifestations. Diabetes induces peripheral nervous system damage which is translated into impaired sensory perception and is termed diabetic neuropathy. Diabetes-related central nervous system damage causes cognitive decline. The behavioral study aimed to evaluate the effect of NAP and D-SAL on peripheral neuropathy and cognitive decline. Peripheral neuropathy was tested by measuring the response to a thermal stimulus, and cognitive ability was measured by a social memory test and a spatial memory test using long- and short-term dependent tasks and a reference memory task. Results indicated an immediate sensory neuropathy in the diabetic model, which was prevented by both peptides and a later neuropathic phase, prevented only by NAP treatment. Cognitive tests revealed impaired performance in both social and spatial memory tests in the diabetes model. Each of the peptides improved different aspects of cognitive behavior, with NAP being more potent than D-SAL. Mechanistically, both NAP and SAL contain a SIP (SxIP) domain that has been shown to interact with microtubule end-binding proteins (EBs). Specifically, we have previously shown a direct interaction of NAP with EB1 and EB3; we have further shown an interaction of the NAP-derived 4 amino acid SKIP peptide with EB3, stimulating axonal transport. Interestingly, the all D-amino acid peptide, D-SKIP, only partially mimicked SKIP activity. Our current results implicate D-SAL activity with potentially reduced potency compared to NAP, partially mimicking the SKIP/D-SKIP results and placing the SIP (SxIP) motif as a central focus for microtubule-based neuroprotection.

Coral snakes predict the evolution of mimicry across New World snakes.

Batesian mimicry, in which harmless species (mimics) deter predators by deceitfully imitating the warning signals of noxious species (models), generates striking cases of phenotypic convergence that are classic examples of evolution by natural selection. However, mimicry of venomous coral snakes has remained controversial because of unresolved conflict between the predictions of mimicry theory and empirical patterns in the distribution and abundance of snakes. Here we integrate distributional, phenotypic and phylogenetic data across all New World snake species to demonstrate that shifts to mimetic coloration in nonvenomous snakes are highly correlated with coral snakes in both space and time, providing overwhelming support for Batesian mimicry. We also find that bidirectional transitions between mimetic and cryptic coloration are unexpectedly frequent over both long- and short-time scales, challenging traditional views of mimicry as a stable evolutionary 'end point' and suggesting that insect and snake mimicry may have different evolutionary dynamics.

NAP alpha-aminoisobutyric acid (IsoNAP).

We set out to identify NAP (davunetide) analogs, providing neuroprotection and reducing tau pathology, specifically addressing protection against protein misfolding. NAP (NAPVSIPQ, intranasal formulation AL-108) is a drug candidate that (1) had a statistically significant impact on two measures, namely digit span and delayed-match-to-sample, tests of verbal recall and visual working memory, respectively, in patient population of mild cognitive impairment [preceding Alzheimer's disease (AD)] and (2) protected functional activities of daily living in schizophrenia patients. Previous preclinical studies have shown that stabilization of NAP by replacement of all L-amino acids by D-amino acids resulted in an active peptide, D-NAP. Other NAP mimetics are now explored. A new NAP analog was designed that included replacement of the proline residues by alpha-aminoisobutyric acid to enhance ß-sheet breaker characteristics, thereby reducing protein misfolding. Three lines of investigations were chosen: (1) protection against the AD-associated amyloid ß (1-42), Aß1-42, peptide toxicity in cell cultures; (2) inhibition of AD-associated tau aggregation in vitro; and (3) cognitive protection in a mouse model of deficiencies of the NAP parent protein, activity-dependent neuroprotective protein (ADNP), exhibiting tau pathology and neurodegeneration. NAP alpha-aminoisobutyric acid (IsoNAP) protected neurons against AD-associated Aß1-42-toxicity, inhibited the aggregation of the tau-derived peptide VQIVYK (important for the aggregation of tau into paired helical filaments, which form the tangles found in AD and related disorders), and protected cognitive functions in a model of ADNP deficiency. With AD being the major tauopathy, novel NAP derivatives that reduce tauopathy and provide neuroprotection as well as cognitive protection are of scientific and clinical interest.

Tau and caspase 3 as targets for neuroprotection.

The peptide drug candidate NAP (davunetide) has demonstrated protective effects in various in vivo and in vitro models of neurodegeneration. NAP was shown to reduce tau hyperphosphorylation as well as to prevent caspase-3 activation and cytochrome-3 release from mitochondria, both characteristic of apoptotic cell death. Recent studies suggest that caspases may play a role in tau pathology. The purpose of this study was to evaluate the effect of NAP on tau hyperphosphorylation and caspase activity in the same biological system. Our experimental setup used primary neuronal cultures subjected to oxygen-glucose deprivation (OGD), with and without NAP or caspase inhibitor. Cell viability was assessed by measuring mitochondrial activity (MTS assay), and immunoblots were used for analyzing protein level. It was shown that apoptosis was responsible for all cell death occurring following ischemia, and NAP treatment showed a concentration-dependent protection from cell death. Ischemia caused an increase in the levels of active caspase-3 and hyperphosphorylated tau, both of which were prevented by either NAP or caspase-inhibitor treatment. Our data suggest that, in this model system, caspase activation may be an upstream event to tau hyperphosphorylation, although additional studies will be required to fully elucidate the cascade of events.