Integration of diverse DNA substrates by a casposase can be targeted to R-loops in vitro by its fusion to Cas9

CRISPR systems build adaptive immunity against mobile genetic elements by DNA capture and integration catalysed by Cas1–Cas2 protein complexes. Recent studies suggested that CRISPR repeats and adaptation module originated from a novel type of DNA transposons called casposons. Casposons encode a Cas1 homologue called casposase that alone integrates into target molecules single and double-stranded DNA containing terminal inverted repeats (TIRs) from casposons. A recent study showed Methanosarcina mazei casposase is able to integrate random DNA oligonucleotides, followed up in this work using Acidoprofundum boonei casposase, from which we also observe promiscuous substrate integration. Here we first show that the substrate flexibility of Acidoprofundum boonei casposase extends to random integration of DNA without TIRs, including integration of a functional gene. We then used this to investigate targeting of the casposase-catalysed DNA integration reactions to specific DNA sites that would allow insertion of defined DNA payloads. Casposase–Cas9 fusions were engineered that were catalytically proficient in vitro and generated RNA-guided DNA integration products from short synthetic DNA or a gene, with or without TIRs. However, DNA integration could still occur unguided due to the competing background activity of the casposase moiety. Expression of Casposase-dCas9 in Escherichia coli cells effectively targeted chromosomal and plasmid lacZ revealed by reduced β-galactosidase activity but DNA integration was not detected. The promiscuous substrate integration properties of casposases make them potential DNA insertion tools. The Casposase–dCas9 fusion protein may serves as a prototype for development in genetic editing for DNA insertion that is independent of homology-directed DNA repair

Adaptation processes that build CRISPR immunity: creative destruction, updated

Prokaryotes can defend themselves against invading mobile genetic elements (MGEs) by acquiring immune memory against them. The memory is a DNA database located at specific chromosomal sites called CRISPRs (clustered regularly interspaced short palindromic repeats) that store fragments of MGE DNA. These are utilised to target and destroy returning MGEs, preventing re-infection. The effectiveness of CRISPR-based immune defence depends on ‘adaptation’ reactions that capture and integrate MGE DNA fragments into CRISPRs. This provides the means for immunity to be delivered against MGEs in ‘interference’ reactions. Adaptation and interference are catalysed by Cas (CRISPR-associated) proteins, aided by enzymes well known for other roles in cells. We survey the molecular biology of CRISPR adaptation, highlighting entirely new developments that may help us to understand how MGE DNA is captured. We focus on processes in Escherichia coli, punctuated with reference to other prokaryotes that illustrate how common requirements for adaptation, DNA capture and integration, can be achieved in different ways. We also comment on how CRISPR adaptation enzymes, and their antecedents, can be utilised for biotechnology

Aerosols, Hurricanes, and their Interactions : A Case Study of Hurricane Sandy

While the effects of aerosols on precipitation have been studied, their effects on more extreme precipitation events like Tropical Cyclones have only been questioned relatively recently. Because of the rarity of the intersection of significant quantities of aerosols and forming/formed tropical cyclones, as well as the possible destruction caused, most experiments about their effects take place in computer models that may not fully simulate the effects of the aerosols. Limitations in satellite sensing make it difficult to track processes and material distributions in hurricanes from afar as well. Hurricane Sandy, a devastating hurricane that formed in October of 2012, may have formed while influenced by relatively smaller but still significant amounts of dust from an African dust event. Since this quantity is different from most heavy polluting scenarios that are simulated, evaluating the extent of the presence, position, and activation of aerosols within this hurricane may give us insight into the potential influences of aerosols in tropical cyclones. MODIS, MISR, AIRS, and CALIPSO data, while having limitations, is analyzed. The extent and possible effects of additional Saharan dust aerosol loading is discussed

The Dark Matter Distribution in Abell 383: Evidence for a Shallow Density Cusp from Improved Lensing, Stellar Kinematic and X-ray Data

We extend our analyses of the dark matter (DM) distribution in relaxed clusters to the case of Abell 383, a luminous X-ray cluster at z=0.189 with a dominant central galaxy and numerous strongly-lensed features. Following our earlier papers, we combine strong and weak lensing constraints secured with Hubble Space Telescope and Subaru imaging with the radial profile of the stellar velocity dispersion of the central galaxy, essential for separating the baryonic mass distribution in the cluster core. Hydrostatic mass estimates from Chandra X-ray observations further constrain the solution. These combined datasets provide nearly continuous constraints extending from 2 kpc to 1.5 Mpc in radius, allowing stringent tests of results from recent numerical simulations. Two key improvements in our data and its analysis make this the most robust case yet for a shallow slope \beta of the DM density profile \rho_DM ~ r^-\beta on small scales. First, following deep Keck spectroscopy, we have secured the stellar velocity dispersion profile to a radius of 26 kpc for the first time in a lensing cluster. Secondly, we improve our previous analysis by adopting a triaxial DM distribution and axisymmetric dynamical models. We demonstate that in this remarkably well-constrained system, the logarithmic slope of the DM density at small radii is \beta < 1.0 (95% confidence). An improved treatment of baryonic physics is necessary, but possibly insufficient, to reconcile our observations with the recent results of high-resolution simulations.Comment: Accepted to ApJ Letter

Attaining the rate-independent limit of a rate-dependent strain gradient plasticity theory

The existence of characteristic strain rates in rate-dependent material models, corresponding to rate-independent model behavior, is studied within a back stress based rate-dependent higher order strain gradient crystal plasticity model. Such characteristic rates have recently been observed for steady-state processes, and the present study aims to demonstrate that the observations in fact unearth a more widespread phenomenon. In this work, two newly proposed back stress formulations are adopted to account for the strain gradient effects in the single slip simple shear case, and characteristic rates for a selected quantity are identified through numerical analysis. Evidently, the concept of a characteristic rate, within the rate-dependent material models, may help unlock an otherwise inaccessible parameter space

Merging fragments of classical logic

We investigate the possibility of extending the non-functionally complete logic of a collection of Boolean connectives by the addition of further Boolean connectives that make the resulting set of connectives functionally complete. More precisely, we will be interested in checking whether an axiomatization for Classical Propositional Logic may be produced by merging Hilbert-style calculi for two disjoint incomplete fragments of it. We will prove that the answer to that problem is a negative one, unless one of the components includes only top-like connectives.Comment: submitted to FroCoS 201

Motivation and needs of informal physics practitioners

Physicists engage with the public to varying degrees at different stages of their careers. However, their public engagement covers many activities, events, and audiences, making their motivations and professional development needs not well understood. As part of ongoing efforts to build and support community in the informal physics space, we conducted interviews with physicists with a range of different experiences in public engagement. We use personas methodology and self-determination theory to articulate their public engagement motivation, challenges, and needs. We present our set of three personas: the physicist who engages in informal physics for self-reflection, the physicist who wants to spark interest and understanding in physics, and the physicist who wants to provide diverse role models to younger students and inspire them to pursue a STEM career. Needs covered a range of resources including science communication training, community building among informal physics practitioners, and mechanisms to recognize, elevate and value informal physics. By bringing user-centered design methodology to a new topical area of physics education research, we expand our understanding of motivations and needs of practitioners in physics public engagement. Therefore, departments, organizations and institutions could draw upon the personas developed to consider the ways to better support physicists in their respective environment.Comment: 9 pages; preliminary portions of some of this analysis appeared in PERC 2022; submitted to PhysRevPE

The trauma film paradigm as an experimental psychopathology model of psychological trauma: intrusive memories and beyond

A better understanding of psychological trauma is fundamental to clinical psychology. Following traumatic event(s), a clinically significant number of people develop symptoms, including those of Acute Stress Disorder and/or Post Traumatic Stress Disorder. The trauma film paradigm offers an experimental psychopathology model to study both exposure and reactions to psychological trauma, including the hallmark symptom of intrusive memories. We reviewed 74 articles that have used this paradigm since the earliest review (Holmes & Bourne, 2008) until July 2014. Highlighting the different stages of trauma processing, i.e. pre-, peri- and post-trauma, the studies are divided according to manipulations before, during and after film viewing, for experimental as well as correlational designs. While the majority of studies focussed on the frequency of intrusive memories, other reactions to trauma were also modelled. We discuss the strengths and weaknesses of the trauma film paradigm as an experimental psychopathology model of trauma, consider ethical issues, and suggest future directions. By understanding the basic mechanisms underlying trauma symptom development, we can begin to translate findings from the laboratory to the clinic, test innovative science-driven interventions, and in the future reduce the debilitating effects of psychopathology following stressful and/or traumatic events

Molecular epidemiology and nasal carriage of Staphylococcus aureus and methicillin-resistant S. aureus among young children attending day care centers and kindergartens in Hong Kong

OBJECTIVES: To investigate the prevalence and molecular epidemiology of Staphylococcus aureus and methicillin-resistant S. aureus (MRSA) nasal carriage in children. METHODS: We collected nasal and nasopharyngeal swabs from 2211 children aged 2-5 years attending 79 day care centers (DCCs) and 113 kindergartens (KGs) in all 18 geographical districts in Hong Kong. RESULTS: The overall carriage rates of S. aureus and MRSA were 27.6% (95% confidence interval [CI], 24.8-28.5%) and 1.3% (95% CI, 0.8-1.8%), respectively. Molecular typing (staphylococcal cassette chromosome mec [SCCmec], sequence type [ST], clonal cluster [CC]) showed that all the 28 MRSA isolates had SCCmec IV (n = 13) or V (n = 15) including 12 isolates with community-associated-MRSA genotypes (ST59-IV/V, ST30-IV and ST88-V), 10 isolates with healthcare-associated-MRSA genotypes (ST45-IV/V, CC5-IV and ST630-V) and six isolates with novel genotypes (ST10-V and CC1-IV). Spa typing indicated that there was some within and between DCCs/KGs transmission of certain MRSA and methicillin-sensitive S. aureus strains but this was not extensive. CONCLUSION: Our findings indicate the potential for DCCs to be a reservoir for emerging MRSA genotypes and highlight the need to enhance education and infection control measures to reduce their cross-transmission in this population.postprin