Acid-active proteases to optimize dietary protein digestibility: a step towards sustainable nutrition.

Introduction: Historically, prioritizing abundant food production often resulted in overlooking nutrient quality and bioavailability, however, environmental concerns have now propelled sustainable nutrition and health efficacy to the forefront of global attention. In fact, increasing demand for protein is the major challenge facing the food system in the 21st century with an estimation that 70% more food is needed by 2050. This shift has spurred interest in plant-based proteins for their sustainability and health benefits, but most alternative sources of protein are poorly digestible. There are two approaches to solve digestibility: improve the digestibility of food proteins or improve the digestive capacity of consumers. Enhancing nutrient digestibility and bioavailability across diverse protein sources is crucial, with proteases presenting a promising avenue. Research, inspired by the proteases of human breast milk, has demonstrated that exogenous microbial proteases can activate within the human digestive tract and substantially increase the digestion of targeted proteins that are otherwise difficult to fully digest. Methods: Here, we introduce the use of an acid-active family of bacterial proteases (S53) to improve the digestibility and nutritional quality of a variety of protein sources, evaluated using the INFOGEST 2.0 protocol. Results: Results from in vitro digestibility indicate that the most effective protease in the S53 family substantially improves the digestibility of an array of animal and plant-derived proteins-soy, pea, chickpea, rice, casein, and whey. On average, this protease elevated protein digestibility by 115% during the gastric phase and by 15% in the intestinal phase, based on the degree of hydrolysis. Discussion: The widespread adoption of these proteases has the potential to enhance nutritional value and contribute to food security and sustainability. This approach would complement ongoing efforts to improve proteins in the food supply, increase the quality of more sustainable protein sources and aid in the nourishment of patients with clinically compromised, fragile intestines and individuals like older adults and high-performance athletes who have elevated protein needs.

Validity and Reliability of a Telehealth Physical Fitness and Functional Assessment Battery for Ambulatory Youth With and Without Mobility Disabilities: Observational Measurement Study.

BACKGROUND: Youth (age 15-24 years) with and without disability are not adequately represented enough in exercise research due to a lack of time and transportation. These barriers can be overcome by including accessible web-based assessments that eliminate the need for on-site visitations. There is no simple, low-cost, and psychometrically sound compilation of measures for physical fitness and function that can be applied to youth with and without mobility disabilities. OBJECTIVE: The first purpose was to determine the statistical level of agreement of 4 web-modified clinical assessments with how they are typically conducted in person at a laboratory (convergent validity). The second purpose was to determine the level of agreement between a novice and an expert rater (interrater reliability). The third purpose was to explore the feasibility of implementing the assessments via 2 metrics: safety and duration. METHODS: The study enrolled 19 ambulatory youth: 9 (47%) with cerebral palsy with various mobility disabilities from a children's hospital and 10 (53%) without disabilities from a university student population. Participants performed a battery of tests via videoconferencing and in person. The test condition (teleassessment and in person) order was randomized. The battery consisted of the hand grip strength test with a dynamometer, the five times sit-to-stand test (FTST), the timed up-and-go (TUG) test, and the 6-minute walk test (6MWT) either around a standard circular track (in person) or around a smaller home-modified track (teleassessment version, home-modified 6-minute walk test [HM6MWT]). Statistical analyses included descriptive data, intraclass correlation coefficients (ICCs), and Bland-Altman plots. RESULTS: The mean time to complete the in-person assessment was 16.9 (SD 4.8) minutes and the teleassessment was 21.1 (SD 5.9) minutes. No falls, injuries, or adverse events occurred. Excellent convergent validity was shown for telemeasured hand grip strength (right ICC=0.96, left ICC=0.98, P<.001) and the TUG test (ICC=0.92, P=.01). The FTST demonstrated good agreement (ICC=0.95, 95% CI 0.79-0.98; P=.01). The HM6MWT demonstrated poor absolute agreement with the 6MWT. However, further exploratory analysis revealed a strong positive correlation between the tests (r=0.83, P<.001). The interrater reliability was excellent for all tests (all ICCs>0.9, P<.05). CONCLUSIONS: This study suggests that videoconference assessments are convenient and useful measures of fitness and function among youth with and without disabilities. This paper presents operationalized teleassessment procedures that can be replicated by health professionals to produce valid and reliable measurements. This study is a first step toward developing teleassessments that can bypass the need for on-site data collection visitations for this age group. Further research is needed to identify psychometrically sound teleassessment procedures, particularly for measures of cardiorespiratory endurance or walking ability.

Altered response to Toll-like receptor 4 activation in fibromyalgia: A low-dose, human experimental endotoxemia pilot study.

In this pilot study, a human intravenous injection of low-dose endotoxin (lipopolysaccharide, LPS) model was used to test if fibromyalgia is associated with altered immune responses to Toll-like receptor 4 (TLR4) activation. Eight women with moderately-severe fibromyalgia and eight healthy women were administered LPS at 0.1 ng/kg in session one and 0.4 ng/kg in session two. Blood draws were collected hourly to characterize the immune response. The primary analytes of interest, leptin and fractalkine, were assayed via commercial radioimmunoassay and enzyme-linked immunosorbent assay kits, respectively. Exploratory analyses were performed on 20 secreted cytokine assays by multiplex cytokine panels, collected hourly. Exploratory analyses were also performed on testosterone, estrogen, and cortisol levels, collected hourly. Additionally, standard clinical complete blood counts with differential (CBC-D) were collected before LPS administration and at the end of the session. The fibromyalgia group demonstrated enhanced leptin and suppressed fractalkine responses to LPS administration. In the exploratory analyses, the fibromyalgia group showed a lower release of IFN-γ, CXCL10, IL-17A, and IL-12 and higher release of IL-15, TARC, MDC, and eotaxin than the healthy group. The results of this study suggest that fibromyalgia may involve an altered immune response to TLR4 activation.

Evidence of neuroinflammation in fibromyalgia syndrome: a [ 18 F]DPA-714 positron emission tomography study.

ABSTRACT: This observational study aimed to determine whether individuals with fibromyalgia (FM) exhibit higher levels of neuroinflammation than healthy controls (HCs), as measured with positron emission tomography using [ 18 F]DPA-714, a second-generation radioligand for the translocator protein (TSPO). Fifteen women with FM and 10 HCs underwent neuroimaging. Distribution volume (V T ) was calculated for in 28 regions of interest (ROIs) using Logan graphical analysis and compared between groups using multiple linear regressions. Group (FM vs HC) was the main predictor of interest and TSPO binding status (high- vs mixed-affinity) was added as a covariate. The FM group had higher V T in the right postcentral gyrus ( b = 0.477, P = 0.033), right occipital gray matter (GM; b = 0.438, P = 0.039), and the right temporal GM ( b = 0.466, P = 0.042). The FM group also had lower V T than HCs in the left isthmus of the cingulate gyrus ( b = -0.553, P = 0.014). In the subgroup of high-affinity binders, the FM group had higher V T in the bilateral precuneus, postcentral gyrus, parietal GM, occipital GM, and supramarginal gyrus. Group differences in the right parietal GM were associated with decreased quality of life, higher pain severity and interference, and cognitive problems. In support of our hypothesis, we found increased radioligand binding (V T ) in the FM group compared with HCs in several brain regions regardless of participants' TSPO binding status. The ROIs overlapped with prior reports of increased TSPO binding in FM. Overall, increasing evidence supports the hypothesis that FM involves microglia-mediated neuroinflammation in the brain.

A Systematic Review of Supervision Research Related to Board Certified Behavior Analysts.

Supervision is paramount within applied behavior analysis. This is particularly true in the case of Board Certified Behavior Analysts® (BCBAs®) and trainees (i.e., those accruing hours and experience to become BCBAs). Given the importance of supervision in shaping future BCBAs (and subsequently the field of applied behavior analysis), researchers have emphasized supervision practices as they relate to BCBAs and trainees. However, what type of work is occurring in this area, such as empirical intervention studies or provision of supervisory recommendations through discussion-style articles, needs to be clarified. Therefore, we conducted the first and most comprehensive systematic review of supervision research related to BCBAs and trainees. Results indicated that most of the articles and work pertaining to BCBA supervision are discussion-style articles, with several survey studies and few empirical intervention studies. We discuss implications and areas for future supervision research. Supplementary Information: The online version contains supplementary material available at 10.1007/s40617-023-00805-0.

Clinical and Analytic Accuracy of Simultaneously Acquired Hemoglobin Measurements: A Multi-Institution Cohort Study to Minimize Redundant Laboratory Usage.

OBJECTIVES: Frequent diagnostic blood sampling contributes to anemia among critically ill children. Reducing duplicative hemoglobin testing while maintaining clinical accuracy can improve patient care efficacy. The objective of this study was to determine the analytical and clinical accuracy of simultaneously acquired hemoglobin measurements with different methods. DESIGN: Retrospective cohort study. SETTING: Two U.S. children's hospitals. PATIENTS: Children (< 18 yr old) admitted to the PICU. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: We identified hemoglobin results from complete blood count (CBC) panels paired with blood gas (BG) panels and point-of-care (POC) devices. We estimated analytic accuracy by comparing hemoglobin distributions, correlation coefficients, and Bland-Altman bias. We measured clinical accuracy with error grid analysis and defined mismatch zones as low, medium, or high risk-based on deviance from unity and risk of therapeutic error. We calculated pairwise agreement to a binary decision to transfuse based on a hemoglobin value. Our cohort includes 49,004 ICU admissions from 29,926 patients, resulting in 85,757 CBC-BG hemoglobin pairs. BG hemoglobin was significantly higher (mean bias, 0.43-0.58 g/dL) than CBC hemoglobin with similar Pearson correlation ( R2 ) (0.90-0.91). POC hemoglobin was also significantly higher, but of lower magnitude (mean bias, 0.14 g/dL). Error grid analysis revealed only 78 (< 0.1%) CBC-BG hemoglobin pairs in the high-risk zone. For CBC-BG hemoglobin pairs, at a BG hemoglobin cutoff of greater than 8.0 g/dL, the "number needed to miss" a CBC hemoglobin less than 7 g/dL was 275 and 474 at each institution, respectively. CONCLUSIONS: In this pragmatic two-institution cohort of greater than 29,000 patients, we show similar clinical and analytic accuracy of CBC and BG hemoglobin. Although BG hemoglobin values are higher than CBC hemoglobin values, the small magnitude is unlikely to be clinically significant. Application of these findings may reduce duplicative testing and decrease anemia among critically ill children.

Excuse Me, I'm Speaking: Analysis of Women's Representation as Keynote and Invited Speakers in Behavior Analysis.

Researchers have recently had greater interest in publishing data on women's participation in behavior analysis. This emphasis has resulted in analyses of women's representation as authors of peer-reviewed publications, and the results have been largely encouraging. Another area to study in this line of research is serving as a keynote or invited speaker at conferences. Although there are some limited data published in this area, no study has yet examined women's participation in these roles across all U.S. state associations for behavior analysis. Thus, we analyzed all U.S. state association keynote and invited conference speakers from 2015 to 2020.

Abnormal immune system response in the brain of women with Fibromyalgia after experimental endotoxin challenge.

Background: The pathophysiology of fibromyalgia (FM) is thought to include an overactive immune system, leading to central nervous system sensitization, allodynia, and hyperalgesia. We aimed to test this theory using an experimental immune system activation procedure and neuroimaging with magnetic resonance spectroscopic imaging (MRSI). Methods: Twelve women with FM and 13 healthy women (healthy controls; HC) received 0.3 or 0.4 ng/kg endotoxin and underwent MRSI before and after the infusion. Changes in brain levels of choline (CHO), myo-inositol (MI), N-Acetylaspartate (NAA), and MRSI-derived brain temperature were compared between groups and dosage levels using mixed analyses of variance. Results: Significant group-by-time interactions in brain temperature were found in the right thalamus. Post-hoc testing revealed that brain temperature increased by 0.55 °C in the right thalamus in FM (t(10) = -3.483, p = 0.006), but not in HCs (p > 0.05). Dose-by-time interactions revealed brain temperature increases in the right insula after 0.4 ng/kg (t(12) = -4.074, p = 0.002), but not after 0.3 ng/kg (p > 0.05). Dose-by-time interactions revealed decreased CHO in the right Rolandic operculum after 0.4 ng/kg endotoxin (t(13) = 3.242, p = 0.006) but not 0.3 ng/kg. In the left paracentral lobule, CHO decreased after 0.3 ng/kg (t(9) = 2.574, p = 0.030) but not 0.4 ng/kg. Dose-by-time interactions affected MI in several brain regions. MI increased after 0.3 ng/kg in the right Rolandic operculum (t(10) = -2.374, p = 0.039), left supplementary motor area (t(9) = -2.303, p = 0.047), and left occipital lobe (t(10) = -3.757, p = 0.004), with no changes after 0.4 ng/kg (p > 0.05). Group-by time interactions revealed decreased NAA in the left Rolandic operculum in FM (t(13) = 2.664, p = 0.019), but not in HCs (p > 0.05). A dose-by-time interaction showed decreased NAA in the left paracentral lobule after 0.3 ng/kg (t(9) = 3.071, p = 0.013) but not after 0.4 ng/kg (p > 0.05). In the combined sample, there was a main effect of time whereby NAA decreased in the left anterior cingulate (F[1,21] = 4.458, p = 0.047) and right parietal lobe (F[1,21] = 5.457, p = 0.029). Conclusion: We found temperature increases and NAA decreases in FM that were not seen in HCs, suggesting that FM patients may have abnormal immune responses in the brain. The 0.3 and 0.4 ng/kg had differential effects on brain temperature and metabolites, with neither dose effecting a stronger response overall. There is insufficient evidence provided by the study to determine whether FM involves abnormal central responses to low-level immune challenges.

Molecular MR Imaging of Renal Fibrogenesis in Mice.

BACKGROUND: In most CKDs, lysyl oxidase oxidation of collagen forms allysine side chains, which then form stable crosslinks. We hypothesized that MRI with the allysine-targeted probe Gd-oxyamine (OA) could be used to measure this process and noninvasively detect renal fibrosis. METHODS: Two mouse models were used: hereditary nephritis in Col4a3-deficient mice (Alport model) and a glomerulonephritis model, nephrotoxic nephritis (NTN). MRI measured the difference in kidney relaxation rate, ΔR1, after intravenous Gd-OA administration. Renal tissue was collected for biochemical and histological analysis. RESULTS: ΔR1 was increased in the renal cortex of NTN mice and in both the cortex and the medulla of Alport mice. Ex vivo tissue analyses showed increased collagen and Gd-OA levels in fibrotic renal tissues and a high correlation between tissue collagen and ΔR1. CONCLUSIONS: Magnetic resonance imaging using Gd-OA is potentially a valuable tool for detecting and staging renal fibrogenesis.

FRETing about the details: Case studies in the use of a genetically encoded fluorescent amino acid for distance-dependent energy transfer.

Förster resonance energy transfer (FRET) is a valuable method for monitoring protein conformation and biomolecular interactions. Intrinsically fluorescent amino acids that can be genetically encoded, such as acridonylalanine (Acd), are particularly useful for FRET studies. However, quantitative interpretation of FRET data to derive distance information requires careful use of controls and consideration of photophysical effects. Here we present two case studies illustrating how Acd can be used in FRET experiments to study small molecule induced conformational changes and multicomponent biomolecular complexes.

Engineered RecA Constructs Reveal the Minimal SOS Activation Complex.

The SOS response is a bacterial DNA damage response pathway that has been heavily implicated in bacteria's ability to evolve resistance to antibiotics. Activation of the SOS response is dependent on the interaction between two bacterial proteins, RecA and LexA. RecA acts as a DNA damage sensor by forming lengthy oligomeric filaments (RecA*) along single-stranded DNA (ssDNA) in an ATP-dependent manner. RecA* can then bind to LexA, the repressor of SOS response genes, triggering LexA degradation and leading to induction of the SOS response. Formation of the RecA*-LexA complex therefore serves as the key "SOS activation signal." Given the challenges associated with studying a complex involving multiple macromolecular interactions, the essential constituents of RecA* that allow LexA cleavage are not well defined. Here, we leverage head-to-tail linked and end-capped RecA constructs as tools to define the minimal RecA* filament that can engage LexA. In contrast to previously postulated models, we found that as few as three linked RecA units are capable of ssDNA binding, LexA binding, and LexA cleavage. We further demonstrate that RecA oligomerization alone is insufficient for LexA cleavage, with an obligate requirement for ATP and ssDNA binding to form a competent SOS activation signal with the linked constructs. Our minimal system for RecA* highlights the limitations of prior models for the SOS activation signal and offers a novel tool that can inform efforts to slow acquired antibiotic resistance by targeting the SOS response.

Concurrent Exercise Training: Long-Term Changes in Body Composition and Motives for Continued Participation in Women with Obesity.

The purpose of this project was to examine the effect of a concurrent exercise program (sprint interval training and resistance exercise) on body composition in women with obesity and factors associated with continued exercise participation following the program. Twenty women (37.1 ± 7.4 y, height = 1.63 ± 0.09 m, weight = 98.22 ± 0.22 kg, BMI = 34.2 ± 2.50 kg/m2) participated in a 10-week exercise intervention consisting of a sprint interval treadmill protocol and resistance training three times a week totaling 30 sessions. Body composition was measured by dual-energy X-ray absorptiometry (iDXA) at pretest, 12 weeks, and six months post-intervention. Semi-structured interviews assessed participants' perception of the program at both 12 weeks and six months. Participants significantly reduced fat mass (p < 0.001), gynoid fat mass (p < 0.010), android fat mass (p = 0.003), and visceral fat mass (p = 0.003) at 12 weeks post-test. At six months, participants maintained their reductions in fat mass (p = 0.015), visceral fat (p = 0.040) and gynoid fat mass (p = 0.032). There were no significant main time effects in lean mass (p = 0.099) or caloric intake (p = 0.053) at 12 weeks or six months. Themes that emerged from the semi-structured interviews at 12 weeks reflected enjoyment in the training, increases in competence and knowledge, as well as apprehension of continuing training on their own. At six months, themes that emerged reflected overcoming barriers, competence regarding high-intensity training, and a lack of competence to engage in resistance training. Sprint interval training coupled with resistance training is a feasible exercise protocol for women with obesity and results in reduced fat mass over six months. Improving women's competence for training is imperative for continued participation.

Generating Efficient Methanomethylophilus alvus Pyrrolysyl-tRNA Synthetases for Structurally Diverse Non-Canonical Amino Acids.

Genetic code expansion (GCE) technologies commonly use the pyrrolysyl-tRNA synthetase (PylRS)/tRNAPyl pairs from Methanosarcina mazei (Mm) and Methanosarcina barkeri (Mb) for site-specific incorporation of non-canonical amino acids (ncAAs) into proteins. Recently a homologous PylRS/tRNAPyl pair from Candidatus Methanomethylophilus alvus Mx1201 (Ma) was developed that, lacking the N-terminal tRNA-recognition domain of most PylRSs, overcomes insolubility, instability, and proteolysis issues seen with Mb/Mm PylRSs. An open question is how to alter Ma PylRS specificity to encode specific ncAAs with high efficiency. Prior work focused on "transplanting" ncAA substrate specificity by reconstructing the same active site mutations found in functional Mm/Mb PylRSs in Ma PylRS. Here, we found that this strategy produced low-efficiency Ma PylRSs for encoding three structurally diverse ncAAs: acridonyl-alanine (Acd), 3-nitro-tyrosine, and m-methyl-tetrazinyl-phenylalanine (Tet3.0-Me). On the other hand, efficient Ma PylRS variants were generated by a conventional life/death selection process from a large library of active site mutants: for Acd encoding, one variant was highly functional in HEK293T cells at just 10 µM Acd; for nitroY encoding, two variants also encoded 3-chloro, 3-bromo-, and 3-iodo-tyrosine at high efficiency; and for Tet-3.0-Me, all variants were more functional at lower ncAA concentrations. All Ma PylRS variants identified through selection had at least two different active site residues when compared with their Mb PylRS counterparts. We conclude that Ma and Mm/Mb PylRSs are sufficiently different that "active site transplantation" yields suboptimal Ma GCE systems. This work establishes a paradigm for expanding the utility of the promising Ma PylRS/tRNAPyl GCE platform.

Fatigue and Pain Severity in Gulf War Illness Is Associated With Changes in Inflammatory Cytokines and Positive Acute Phase Proteins.

OBJECTIVE: The aim of the study is to investigate relationships between inflammatory analytes and symptoms of pain and fatigue in Gulf War illness (GWI). METHODS: In this preliminary study, 12 male veterans meeting GWI criteria provided daily blood samples and symptom ratings over 25 days. Linear mixed models were used to analyze associations between symptoms and sera concentrations of cytokines, acute phase proteins, insulin, and brain-derived neurotropic factor. RESULTS: Analyses included 277 days with both blood draws and self-reports. Days with worse fatigue severity were associated with higher C-reactive protein and serum amyloid A, and lower eotaxin 1. Muscle pain and joint pain were associated with leptin, monocyte chemoattractant protein 1, and interferon γ-induced protein. Joint pain was further associated with serum amyloid A and eotaxin 3. CONCLUSIONS: Gulf War illness involves fatigue and pain associated with inflammation. Conventional and novel anti-inflammatories should be further explored for the treatment of GWI.

Improving recording and reporting of dementia and frailty via electronic patient record by ambulance staff in a single service (IDEAS).

BACKGROUND: Dementia is common in older adults assessed by ambulance services. However, inconsistent reporting via the patient record may result in this diagnosis being overlooked by healthcare staff further down the care pathway. This can have a deleterious effect on subsequent patient care, increasing morbidity and mortality. We sought to understand how and where ambulance staff would like to record this finding on the electronic patient record (ePR). METHODS: We designed and implemented a survey of ambulance staff in a single service to understand how they identify patients with dementia, how they record dementia on the ePR and how the ePR could be improved to better capture dementia. Scoping questions on frailty were included. The survey was tested using cognitive interviewing. Analysis was conducted using descriptive statistics for closed questions and thematic analysis for open questions as appropriate. RESULTS: 131 surveys were completed; 60% of participants were paramedics and 40% were other grades of front line staff. Participants reported consulting electronic/paper sources, and individuals such as carers involved in the patients' care, to establish whether dementia had been diagnosed. Frailty assessments were prompted by social context, reduced mobility, a fall or diagnosis of dementia. Staff reported documenting dementia in 20 different areas on the ePR and 46% of participants stated a preference for a designated area to record the information. However, 15% indicated it was not necessary to record dementia or that no ePR changes were required. CONCLUSIONS: We have highlighted the variation in ambulance staff practice in recording of dementia. Alterations to the ePR are required to ensure that dementia is recorded consistently and is easily retrievable. Clearer guidance on when to assess frailty may also enhance information provision to care staff in other sectors, resulting in more appropriate clinical and social care.

Reducing ambulance conveyance for older people with and without dementia: evidence of the role of social care from a regional, year-long service evaluation using retrospective routine data.

INTRODUCTION: Older people, especially those with dementia, have a high risk of deterioration following admission to hospital. More than 60% of older people attended by South Central Ambulance Service (SCAS) clinicians are conveyed to hospital, although many conveyances may not have been due to life-threatening conditions. We aimed to understand patterns of conveyance and alternative referral pathways used following ambulance attendance to an older person. METHODS: Service evaluation, using routinely collected, anonymised electronic records. PARTICIPANTS: Electronic records of people aged ≥75 years for whom an ambulance was dispatched between April 2016 and March 2017 within the geographical boundaries of SCAS NHS Foundation Trust, who were alive on arrival of the ambulance. Conveyance rates are described according to patient and emergency-call characteristics. Logistic regression was used to produce adjusted odds ratios for conveyance. Alternative referral pathways used are described. RESULTS: Of 110,781 patients attended, 64% were conveyed to hospital. Factors associated with reduced odds of conveyance included out-of-hours calls (adjusted odds ratio (aOR) 0.82 [0.79-0.85]), living alone with a care package or with family plus care package (aOR 0.66 [0.62-0.69]; aOR 0.58 [0.54-0.62] respectively) and a record of dementia (0.91 [0.87-0.96]). Living in a nursing home was associated with an increased risk of conveyance (aOR 1.25 [1.15-1.36]). Patients with dementia with more income were significantly less likely to be conveyed than those with less income. Alternative referral services were used in 22% of non-conveyed patients, most commonly GP, out-of-hours and falls services. DISCUSSION: People aged ≥75 years have high rates of conveyance, which are influenced by factors such as out-of-hours calls, dementia and receipt of social care. Low use of alternative referral services may reflect limited availability or difficulty in access. A better understanding of how these factors influence ambulance clinician decision-making is integral to improvement of outcomes for older people.

Auditory Processing of Non-speech Stimuli by Children in Dual-Language Immersion Programs.

Perception of low-level auditory cues such as frequency modulation (FM) and rise time (RT) is crucial for development of phonemic representations, segmentation of word boundaries, and attunement to prosodic patterns in language. While learning an additional language, children may develop an increased sensitivity to these cues to extract relevant information from multiple types of linguistic input. Performance on these auditory processing tasks such as FM and RT by children learning another language is, however, unknown. Here we examine 92 English-speaking 7-8-year-olds in the U.S. and their performance in FM and RT perceptual tasks at the end of their second year in Cantonese or Spanish dual-language immersion compared to children in general English education programs. Results demonstrate that children in immersion programs have greater sensitivity to FM, but not RT, controlling for various factors. The immersion program students were also observed to have better phonological awareness performance. However, individual differences in FM sensitivity were not associated with phonological awareness, a pattern typically observed in monolinguals. These preliminary findings suggest a possible impact of formal language immersion on low-level auditory processing. Additional research is warranted to understand causal relationships and ultimate impact on language skills in multilinguals.

An improved fluorescent noncanonical amino acid for measuring conformational distributions using time-resolved transition metal ion FRET.

With the recent explosion in high-resolution protein structures, one of the next frontiers in biology is elucidating the mechanisms by which conformational rearrangements in proteins are regulated to meet the needs of cells under changing conditions. Rigorously measuring protein energetics and dynamics requires the development of new methods that can resolve structural heterogeneity and conformational distributions. We have previously developed steady-state transition metal ion fluorescence resonance energy transfer (tmFRET) approaches using a fluorescent noncanonical amino acid donor (Anap) and transition metal ion acceptor to probe conformational rearrangements in soluble and membrane proteins. Here, we show that the fluorescent noncanonical amino acid Acd has superior photophysical properties that extend its utility as a donor for tmFRET. Using maltose-binding protein (MBP) expressed in mammalian cells as a model system, we show that Acd is comparable to Anap in steady-state tmFRET experiments and that its long, single-exponential lifetime is better suited for probing conformational distributions using time-resolved FRET. These experiments reveal differences in heterogeneity in the apo and holo conformational states of MBP and produce accurate quantification of the distributions among apo and holo conformational states at subsaturating maltose concentrations. Our new approach using Acd for time-resolved tmFRET sets the stage for measuring the energetics of conformational rearrangements in soluble and membrane proteins in near-native conditions.

Discovering antiviral restriction factors and pathways using genetic screens.

Viral infections activate the powerful interferon (IFN) response that induces the expression of several hundred IFN stimulated genes (ISGs). The principal role of this extensive response is to create an unfavourable environment for virus replication and to limit spread; however, untangling the biological consequences of this large response is complicated. In addition to a seemingly high degree of redundancy, several ISGs are usually required in combination to limit infection as individual ISGs often have low to moderate antiviral activity. Furthermore, what ISG or combination of ISGs are antiviral for a given virus is usually not known. For these reasons, and since the function(s) of many ISGs remains unexplored, genome-wide approaches are well placed to investigate what aspects of this response result in an appropriate, virus-specific phenotype. This review discusses the advances screening approaches have provided for the study of host defence mechanisms, including clustered regularly interspaced short palindromic repeats/CRISPR associated protein 9 (CRISPR/Cas9), ISG expression libraries and RNA interference (RNAi) technologies.