Extramammary Paget's disease

Extramammary Paget's disease is an in situ skin and mucosal carcinoma frequently associated with and probably arising in a subjacent or regionally proximate carcinoma. Microscopic spread of tumor cells almost always extends beyond clinically apparent disease. Surgical treatment requires carefully planned, systematic excision under precise histologic control. An ideal treatment method remains to be developed. Inadequate excision usually results in recurrences that can be successfully treated by reexcision. Associated invasive carcinomas occur frequently, and mortality is high in these patients.     

Cytotoxic Chromosomal Targeting by CRISPR/Cas Systems Can Reshape Bacterial Genomes and Expel or Remodel Pathogenicity Islands

In prokaryotes, clustered regularly interspaced short palindromic repeats (CRISPRs) and their associated (Cas) proteins constitute a defence system against bacteriophages and plasmids. CRISPR/Cas systems acquire short spacer sequences from foreign genetic elements and incorporate these into their CRISPR arrays, generating a memory of past invaders. Defence is provided by short non-coding RNAs that guide Cas proteins to cleave complementary nucleic acids. While most spacers are acquired from phages and plasmids, there are examples of spacers that match genes elsewhere in the host bacterial chromosome. In Pectobacterium atrosepticum the type I-F CRISPR/Cas system has acquired a self-complementary spacer that perfectly matches a protospacer target in a horizontally acquired island (HAI2) involved in plant pathogenicity. Given the paucity of experimental data about CRISPR/Cas–mediated chromosomal targeting, we examined this process by developing a tightly controlled system. Chromosomal targeting was highly toxic via targeting of DNA and resulted in growth inhibition and cellular filamentation. The toxic phenotype was avoided by mutations in the cas operon, the CRISPR repeats, the protospacer target, and protospacer-adjacent motif (PAM) beside the target. Indeed, the natural self-targeting spacer was non-toxic due to a single nucleotide mutation adjacent to the target in the PAM sequence. Furthermore, we show that chromosomal targeting can result in large-scale genomic alterations, including the remodelling or deletion of entire pre-existing pathogenicity islands. These features can be engineered for the targeted deletion of large regions of bacterial chromosomes. In conclusion, in DNA–targeting CRISPR/Cas systems, chromosomal interference is deleterious by causing DNA damage and providing a strong selective pressure for genome alterations, which may have consequences for bacterial evolution and pathogenicity.     

A pair of inhibitory neurons are required to sustain labile memory in the Drosophila mushroom body

Labile memory is thought to be held in the brain as persistent neural network activity. However, it is not known how biologically relevant memory circuits are organized and operate. Labile and persistent appetitive memory in Drosophila requires output after training from the α'β' subset of mushroom body (MB) neurons and from a pair of modulatory dorsal paired medial (DPM) neurons. DPM neurons innervate the entire MB lobe region and appear to be pre- and postsynaptic to the MB, consistent with a recurrent network model. Here we identify a role after training for synaptic output from the GABAergic anterior paired lateral (APL) neurons. Blocking synaptic output from APL neurons after training disrupts labile memory but does not affect long-term memory. APL neurons contact DPM neurons most densely in the α'β' lobes, although their processes are intertwined and contact throughout all of the lobes. Furthermore, APL contacts MB neurons in the α' lobe but makes little direct contact with those in the distal α lobe. We propose that APL neurons provide widespread inhibition to stabilize and maintain synaptic specificity of a labile memory trace in a recurrent DPM and MB α'β' neuron circuit.     

Mitogenomic insights into a recently described and rarely observed killer whale morphotype

Identifying evolutionary divergent taxonomic units, e.g. species and subspecies, is important for conservation and evolutionary biology. The ‘type D’ killer whale, Orcinus orca, is a rarely observed morphotype with a pelagic, circumpolar subantarctic distribution, making dedicated research and therefore taxonomic study extremely difficult to date. In this study, we used DNA target enrichment hybridisation capture coupled to high throughput sequencing, to obtain the first DNA sequence from the only known museum specimen of this recently described morphotype. The high coverage, complete mitogenome sequence was compared to a previously published global dataset of 139 individuals, indicating that this type is highly divergent to all previously genetically sequenced killer whale forms. The estimated divergence time (390,000 years ago) from its most recent common ancestor with other extant killer whale lineages was the second oldest split within the killer whale phylogeny. This study provides the first genetic support of type D potentially being a distinct subspecies or species of killer whale, although further samples are needed to identify whether there is monophyly of mitogenome sequences and whether nuclear DNA also indicates reproductive isolation. These findings also highlight the value of natural history museum collections and new technologies to investigate the taxonomy of rare, cryptic or difficult to access species.     

The effect of level of feeding, genetic merit, body condition score and age on biological parameters of a mammary gland model

An evolutionary algorithm was applied to a mechanistic model of the mammary gland to find the parameter values that minimised the difference between predicted and actual lactation curves of milk yields in New Zealand Jersey cattle managed at different feeding levels. The effect of feeding level, genetic merit, body condition score at parturition and age on total lactation yields of milk, fat and protein, days in milk, live weight and evolutionary algorithm derived mammary gland parameters was then determined using a multiple regression model. The mechanistic model of the mammary gland was able to fit lactation curves that corresponded to actual lactation curves with a high degree of accuracy. The senescence rate of quiescent (inactive) alveoli was highest at the very low feeding level. The active alveoli population at peak lactation was highest at very low feeding levels, but lower nutritional status at this feeding level prevented high milk yields from being achieved. Genetic merit had a significant linear effect on the active alveoli population at peak and mid to late lactation, with higher values in animals, which had higher breeding values for milk yields. A type of genetic merit × feeding level scaling effect was observed for total yields of milk and fat, and total number of alveoli produced from conception until the end of lactation with the benefits of increases in genetic merit being greater at high feeding levels. A genetic merit × age scaling effect was observed for total lactation protein yields. Initial rates of differentiation of progenitor cells declined with age. Production levels of alveoli from conception to the end of lactation were lowest in 5- to 8-year-old animals; however, in these older animals, quiescent alveoli were reactivated more frequently. The active alveoli population at peak lactation and rates of active alveoli proceeding to quiescence were highest in animals of intermediate body condition scores of 4.0 to 5.0. The results illustrate the potential uses of a mechanistic model of the mammary gland to fit a lactation curve and to quantify the effects of feeding level, genetic merit, body condition score, and age on mammary gland dynamics throughout lactation.     

Dynamic structure of retinylidene ligand of rhodopsin probed by molecular simulations

Rhodopsin is currently the only available atomic-resolution template for understanding biological functions of the G protein-coupled receptor (GPCR) family. The structural basis for the phenomenal dark state stability of 11-cis-retinal bound to rhodopsin and its ultrafast photoreaction are active topics of research. In particular, the beta-ionone ring of the retinylidene inverse agonist is crucial for the activation mechanism. We analyzed a total of 23 independent, 100 ns all-atom molecular dynamics simulations of rhodopsin embedded in a lipid bilayer in the microcanonical (N,V,E) ensemble. Analysis of intramolecular fluctuations predicts hydrogen-out-of-plane (HOOP) wagging modes of retinal consistent with those found in Raman vibrational spectroscopy. We show that sampling and ergodicity of the ensemble of simulations are crucial for determining the distribution of conformers of retinal bound to rhodopsin. The polyene chain is rigidly locked into a single, twisted conformation, consistent with the function of retinal as an inverse agonist in the dark state. Most surprisingly, the beta-ionone ring is mobile within its binding pocket; interactions are non-specific and the cavity is sufficiently large to enable structural heterogeneity. We find that retinal occupies two distinct conformations in the dark state, contrary to most previous assumptions. The beta-ionone ring can rotate relative to the polyene chain, thereby populating both positively and negatively twisted 6-s-cis enantiomers. This result, while unexpected, strongly agrees with experimental solid-state (2)H NMR spectra. Correlation analysis identifies the residues most critical to controlling mobility of retinal; we find that Trp265 moves away from the ionone ring prior to any conformational transition. Our findings reinforce how molecular dynamics simulations can challenge conventional assumptions for interpreting experimental data, especially where existing models neglect conformational fluctuations.     

Percolation on fitness landscapes: effects of correlation, phenotype, and incompatibilities

We study how correlations in the random fitness assignment may affect the structure of fitness landscapes, in three classes of fitness models. The first is a phenotype space in which individuals are characterized by a large number n of continuously varying traits. In a simple model of random fitness assignment, viable phenotypes are likely to form a giant connected cluster percolating throughout the phenotype space provided the viability probability is larger than 1/2(n). The second model explicitly describes genotype-to-phenotype and phenotype-to-fitness maps, allows for neutrality at both phenotype and fitness levels, and results in a fitness landscape with tunable correlation length. Here, phenotypic neutrality and correlation between fitnesses can reduce the percolation threshold, and correlations at the point of phase transition between local and global are most conducive to the formation of the giant cluster. In the third class of models, particular combinations of alleles or values of phenotypic characters are "incompatible" in the sense that the resulting genotypes or phenotypes have zero fitness. This setting can be viewed as a generalization of the canonical Bateson-Dobzhansky-Muller model of speciation and is related to K-SAT problems, prominent in computer science. We analyze the conditions for the existence of viable genotypes, their number, as well as the structure and the number of connected clusters of viable genotypes. We show that analysis based on expected values can easily lead to wrong conclusions, especially when fitness correlations are strong. We focus on pairwise incompatibilities between diallelic loci, but we also address multiple alleles, complex incompatibilities, and continuous phenotype spaces. In the case of diallelic loci, the number of clusters is stochastically bounded and each cluster contains a very large sub-cube. Finally, we demonstrate that the discrete NK model shares some signature properties of models with high correlations.     

10-y Risks of Death and Emergency Re-admission in Adolescents Hospitalised with Violent,Drug- or Alcohol-Related,or Self-Inflicted Injury: A Population-Based Cohort Study

Background

Hospitalisation for adversity-related injury (violent, drug/alcohol-related, or self-inflicted injury) has been described as a “teachable moment”, when intervention may reduce risks of further harm. Which adolescents are likely to benefit most from intervention strongly depends on their long-term risks of harm. We compared 10-y risks of mortality and re-admission after adversity-related injury with risks after accident-related injury.

Methods and Findings

We analysed National Health Service admissions data for England (1 April 1997–31 March 2012) for 10–19 y olds with emergency admissions for adversity-related injury (violent, drug/alcohol-related, or self-inflicted injury; n = 333,009) or for accident-related injury (n = 649,818). We used Kaplan–Meier estimates and Cox regression to estimate and compare 10-y post-discharge risks of death and emergency re-admission. Among adolescents discharged after adversity-related injury, one in 137 girls and one in 64 boys died within 10 y, and 54.2% of girls and 40.5% of boys had an emergency re-admission, with rates being highest for 18–19 y olds. Risks of death were higher than in adolescents discharged after accident-related injury (girls: age-adjusted hazard ratio 1.61, 95% CI 1.43–1.82; boys: 2.13, 95% CI 1.98–2.29), as were risks of re-admission (girls: 1.76, 95% CI 1.74–1.79; boys: 1.41, 95% CI 1.39–1.43). Risks of death and re-admission were increased after all combinations of violent, drug/alcohol-related, and self-inflicted injury, but particularly after any drug/alcohol-related or self-inflicted injury (i.e., with/without violent injury), for which age-adjusted hazard ratios for death in boys ranged from 1.67 to 5.35, compared with 1.25 following violent injury alone (girls: 1.09 to 3.25, compared with 1.27). The main limitation of the study was under-recording of adversity-related injuries and misclassification of these cases as accident-related injuries. This misclassification would attenuate the relative risks of death and re-admission for adversity-related compared with accident-related injury.

Conclusions

Adolescents discharged after an admission for violent, drug/alcohol-related, or self-inflicted injury have increased risks of subsequent harm up to a decade later. Introduction of preventive strategies for reducing subsequent harm after admission should be considered for all types of adversity-related injury, particularly for older adolescents.