Targeted Insertion of the mPing Transposable Element

Class II DNA Transposable Elements (TEs) are moved from one location to another in the genome by the action of transposase proteins that bind to repeat sequences at the ends of the elements. Although the location TE insertion is mostly random, the addition of DNA binding domains to the transposase proteins has allowed for targeted insertion of some elements. In this study, the Gal4 binding domain was added to the transposase proteins, ORF1 and TPase, which mobilize the mPing element from rice. The Gal4:TPase construct was capable of increasing the number of mPing insertions into the Gal2 and Gal4 promoter sequences in yeast. While this confirms that mPing insertion preference can be manipulated, the target specificity is relatively low. Thus, the CRISPR/Cas9 system was tested for its ability to generate targeted insertion of mPing. A dCas9:TPase fusion protein had a low transposition rate suggesting that the addition of this large protein disrupts TPase function. Unfortunately, the use of a MS2 binding domain to localize the TPase to the MS2 hairpin containing gRNA failed to produce targeted insertion. Thus, our results suggest that the addition of small DNA binding domain to the N-terminal of TPase is the best strategy for targeted insertion of mPing

Structure and Mechanism of a Metal-Sensing Regulatory RNA

SummaryOrganisms maintain the correct balance of intracellular metals primarily through metal-sensing proteins that control transport and storage of the target ion(s). Here, we reveal the basis of metal sensing and genetic control by a metalloregulatory RNA. Our data demonstrate that a previously uncharacterized orphan riboswitch, renamed the “M-box,” is a divalent metal-sensing RNA involved in Mg2+ homeostasis. A combination of genetic, biochemical, and biophysical techniques demonstrate that Mg2+ induces a compacted tertiary architecture for M-box RNAs that regulates the accessibility of nucleotides involved in genetic control. Molecular details are provided by crystallographic structure determination of a Mg2+-bound M-box RNA. Given the distribution of this RNA element, it may constitute a common mode for bacterial metal ion regulation, and its discovery suggests the possibility of additional RNA-based metal sensors in modern and primordial organisms

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Mammal communities are larger and more diverse in moderately developed areas

Developed areas are thought to have low species diversity, low animal abundance, few native predators, and thus low resilience and ecological function. Working with citizen scientist volunteers to survey mammals at 1427 sites across two development gradients (wild-rural-exurban- suburban-urban) and four plot types (large forests, small forest fragments, open areas and residential yards) in the eastern US, we show that developed areas actually had significantly higher or statistically similar mammalian occupancy, relative abundance, richness and diversity compared to wild areas. However, although some animals can thrive in suburbia, conservation of wild areas and preservation of green space within cities are needed to protect sensitive species and to give all species the chance to adapt and persist in the Anthropocene. DOI: https://doi.org/10.7554/eLife.38012.00

WHO consultation on group B Streptococcus vaccine development: Report from a meeting held on 27-28 April 2016.

Globally, group B Streptococcus (GBS) remains a leading cause of sepsis and meningitis in infants in the first 90days of life. Intrapartum antibiotic prophylaxis (IAP) for women at increased risk of transmitting GBS to their newborns has been effective in reducing part, but not all, of the GBS disease burden in many high income countries (HICs). In low- and middle-income countries (LMICs), IAP use is low. Immunization of pregnant women with a GBS vaccine represents an alternative strategy to protecting newborns and young infants, through transplacental antibody transfer and potentially by reducing new vaginal colonization. This vaccination strategy was first suggested in the 1970s and several potential GBS vaccines have completed phase I/II clinical trials. During the 2015 WHO Product Development for Vaccines Advisory Committee meeting, GBS was identified as a high priority for the development of a vaccine for maternal immunization because of the major public health burden posed by GBS in LMICs, and the high technical feasibility for successful development. Following this meeting, the first WHO technical consultation on GBS vaccines was held on the 27th and 28th of April 2016, to consider development pathways for such vaccines, focused on their potential role in reducing newborn and young infant deaths and possibly stillbirths in LMICs. Discussion topics included: (1) pathophysiology of disease; (2) current gaps in the knowledge of global disease burden and serotype distribution; (3) vaccine candidates under development; (4) design considerations for phase III trials; and (5) pathways to licensure, policy recommendations and use. Efforts to address gaps identified in each of these areas are needed to establish the public health need for, the development and deployment of, efficacious GBS vaccines. In particular, more work is required to understand the global disease burden of GBS-associated stillbirths, and to develop quality-assured standardized antibody assays to identify correlates of protection

Transcriptomic analysis of field-droughted sorghum from seedling to maturity reveals biotic and metabolic responses.

Drought is the most important environmental stress limiting crop yields. The C4 cereal sorghum [Sorghum bicolor (L.) Moench] is a critical food, forage, and emerging bioenergy crop that is notably drought-tolerant. We conducted a large-scale field experiment, imposing preflowering and postflowering drought stress on 2 genotypes of sorghum across a tightly resolved time series, from plant emergence to postanthesis, resulting in a dataset of nearly 400 transcriptomes. We observed a fast and global transcriptomic response in leaf and root tissues with clear temporal patterns, including modulation of well-known drought pathways. We also identified genotypic differences in core photosynthesis and reactive oxygen species scavenging pathways, highlighting possible mechanisms of drought tolerance and of the delayed senescence, characteristic of the stay-green phenotype. Finally, we discovered a large-scale depletion in the expression of genes critical to arbuscular mycorrhizal (AM) symbiosis, with a corresponding drop in AM fungal mass in the plants' roots

The metabolome regulates the epigenetic landscape during naive-to-primed human embryonic stem cell transition.

For nearly a century developmental biologists have recognized that cells from embryos can differ in their potential to differentiate into distinct cell types. Recently, it has been recognized that embryonic stem cells derived from both mice and humans exhibit two stable yet epigenetically distinct states of pluripotency: naive and primed. We now show that nicotinamide N-methyltransferase (NNMT) and the metabolic state regulate pluripotency in human embryonic stem cells (hESCs).  Specifically, in naive hESCs, NNMT and its enzymatic product 1-methylnicotinamide are highly upregulated, and NNMT is required for low S-adenosyl methionine (SAM) levels and the H3K27me3 repressive state. NNMT consumes SAM in naive cells, making it unavailable for histone methylation that represses Wnt and activates the HIF pathway in primed hESCs. These data support the hypothesis that the metabolome regulates the epigenetic landscape of the earliest steps in human development

Characteristics associated with quality of life among people with drug-resistant epilepsy

Quality of Life (QoL) is the preferred outcome in non-pharmacological trials, but there is little UK population evidence of QoL in epilepsy. In advance of evaluating an epilepsy self-management course we aimed to describe, among UK participants, what clinical and psycho-social characteristics are associated with QoL. We recruited 404 adults attending specialist clinics, with at least two seizures in the prior year and measured their self-reported seizure frequency, co-morbidity, psychological distress, social characteristics, including self-mastery and stigma, and epilepsy-specific QoL (QOLIE-31-P). Mean age was 42 years, 54% were female, and 75% white. Median time since diagnosis was 18 years, and 69% experienced ≥10 seizures in the prior year. Nearly half (46%) reported additional medical or psychiatric conditions, 54% reported current anxiety and 28% reported current depression symptoms at borderline or case level, with 63% reporting felt stigma. While a maximum QOLIE-31-P score is 100, participants’ mean score was 66, with a wide range (25–99). In order of large to small magnitude: depression, low self-mastery, anxiety, felt stigma, a history of medical and psychiatric comorbidity, low self-reported medication adherence, and greater seizure frequency were associated with low QOLIE-31-P scores. Despite specialist care, UK people with epilepsy and persistent seizures experience low QoL. If QoL is the main outcome in epilepsy trials, developing and evaluating ways to reduce psychological and social disadvantage are likely to be of primary importance. Educational courses may not change QoL, but be one component supporting self-management for people with long-term conditions, like epilepsy