Development of experimental autoimmune encephalomyelitis (EAE) in mice requires vitamin D and the vitamin D receptor

The development of experimental autoimmune encephalomyelitis (EAE), a model of multiple sclerosis, has been studied in mice that were (i) vitamin D-deficient, (ii) minus the vitamin D receptor, (iii) minus a vitamin D 25-hydroxylase, and (iv) minus the vitamin D 25-hydroxyvitamin D-1α-hydroxylase. EAE development was markedly suppressed in mice lacking the vitamin D receptor and partially suppressed in vitamin D-insufficient mice. However, the absence of either of the two key hydroxylases (i.e., 25-hydroxylase and 1α-hydroxylase) neither inhibits nor enhances the development of EAE. These results indicate that vitamin D and the vitamin D receptor are required for the development of EAE. The results also suggest that 1,25-dihydroxyvitamin D(3) may not play a role in this autoimmune response.     

Assistive listening devices drive neuroplasticity in children with dyslexia

Children with dyslexia often exhibit increased variability in sensory and cognitive aspects of hearing relative to typically developing peers. Assistive listening devices (classroom FM systems) may reduce auditory processing variability by enhancing acoustic clarity and attention. We assessed the impact of classroom FM system use for 1 year on auditory neurophysiology and reading skills in children with dyslexia. FM system use reduced the variability of subcortical responses to sound, and this improvement was linked to concomitant increases in reading and phonological awareness. Moreover, response consistency before FM system use predicted gains in phonological awareness. A matched control group of children with dyslexia attending the same schools who did not use the FM system did not show these effects. Assistive listening devices can improve the neural representation of speech and impact reading-related skills by enhancing acoustic clarity and attention, reducing variability in auditory processing.Children with dyslexia, reading impairment not caused by deficits in ability or opportunity (1), often have difficulties with orienting and maintaining attention (2, 3). Although the ability to direct attention is still developing during the elementary school years (4), dyslexics have poorer task-dependent attentional shifting in both auditory and visual modalities than their typically developing peers even into adulthood (2, 3). These deficits may impact and be impacted by heightened variability in sensory processes, such as inconsistent representations of speech by the auditory nervous system, and could contribute to documented impairments in auditory processing (5–7) and difficulty with meaningfully disambiguating speech sounds (8). Children with dyslexia can exhibit abnormal subcortical processing of speech, particularly in response to acoustic elements crucial for differentiating speech sounds (9–11). Deficient auditory sensory representation and unsuccessful disambiguation of speech likely contribute to the well-documented impairments in phonological awareness and phonological memory seen in children with dyslexia (12–14), with auditory processing skills in prereaders predicting later language skill (15, 16). Because the auditory system integrates both sensory and cognitive facets of hearing, we suggest that through repeated, impaired interaction with sound, children with dyslexia can develop abnormal sensory representations of speech as well as abnormal cognitive skills for the interpretation of speech. For example, a child who repeatedly misperceives the sounds “cat” as “bat” or “pat” fails to make a robust sound-to-meaning connection between those sounds and their referent. However, because of this same integrative nature of the auditory system, deficient function can be improved with auditory training.Auditory perception and neurophysiology can be altered with auditory training (17–23). These changes can be traced directly to cross-cortical and descending cortical influence on neural receptivity in animal models and are driven by the behavioral importance of sounds (18, 24). In humans, attention and working memory are important components of training-related changes (25) and may serve to direct descending cortical influence on auditory sensory function. Computer-based perceptual games, musical training, and language learning can provide effective training for children with developmental learning disorders, such as dyslexia, because they actively engage attention to sound. Classroom assistive listening devices, which can be worn throughout the school day, can also improve auditory processing by engendering enhancements in attention, as reported by both teachers and students (26–28). Assistive listening devices (i.e., classroom FM systems) also result in neurophysiologic enhancements in response to attended vs. ignored sounds (29). Such systems increase the signal-to-noise ratio of the speaker of interest (e.g., the teacher) (30) and create a more stable acoustic input by reducing the impact of background noise on the most vulnerable portion of speech sounds (31). These acoustic enhancements, along with accompanying improvements in auditory attention, lead to boosts in academic achievement, literacy, and phonological awareness, with the greatest benefits seen for children with learning impairments (32–34).What are the biological mechanisms by which classroom FM system use improves auditory attention and phonological awareness in children with dyslexia? How might these benefits translate to the neural representation of speech? Here, we investigated the impact of classroom FM system use on auditory brainstem encoding of stop consonants, which can be deficient in children with dyslexia. Auditory brainstem function is stable from test to retest in the absence of intervention (35, 36), but can be altered by short-term auditory training (19, 20, 22), lifelong experience such as musical training and language experience (37, 38), and directed attention (39). Here we assessed auditory brainstem responses and reading performance in children with dyslexia before and after classroom FM system use for one academic year. We hypothesized that enhanced neural consistency would accompany improvement in reading skills in children wearing the FM systems but not in a control group of dyslexic children in the same classrooms who did not wear assistive listening devices. We further hypothesized that neural consistency would improve pervasively throughout the recording session and not simply offset neural fatigue.     

Heart wall myofibers are arranged in minimal surfaces to optimize organ function

Heart wall myofibers wind as helices around the ventricles, strengthening them in a manner analogous to the reinforcement of concrete cylindrical columns by spiral steel cables [Richart FE, et al. (1929) Univ of Illinois, Eng Exp Stn Bull 190]. A multitude of such fibers, arranged smoothly and regularly, contract and relax as an integrated functional unit as the heart beats. To orchestrate this motion, fiber tangling must be avoided and pumping should be efficient. Current models of myofiber orientation across the heart wall suggest groupings into sheets or bands, but the precise geometry of bundles of myofibers is unknown. Here we show that this arrangement takes the form of a special minimal surface, the generalized helicoid [Blair DE, Vanstone JR (1978) Minimal Submanifolds and Geodesics 13-16], closing the gap between individual myofibers and their collective wall structure. The model holds across species, with a smooth variation in its three curvature parameters within the myocardial wall providing tight fits to diffusion magnetic resonance images from the rat, the dog, and the human. Mathematically it explains how myofibers are bundled in the heart wall while economizing fiber length and optimizing ventricular ejection volume as they contract. The generalized helicoid provides a unique foundation for analyzing the fibrous composite of the heart wall and should therefore find applications in heart tissue engineering and in the study of heart muscle diseases.     

Identification of two functionally redundant RNA elements in the coding sequence of poliovirus using computer-generated design

Genomes of RNA viruses contain multiple functional RNA elements required for translation or RNA replication. We use unique approaches to identify functional RNA elements in the coding sequence of poliovirus (PV), a plus strand RNA virus. The general method is to recode large segments of the genome using synonymous codons, such that protein sequences, codon use, and codon pair bias are conserved but the nucleic acid sequence is changed. Such recoding does not affect the growth of PV unless it destroys the sequence/structure of a functional RNA element. Using genetic analyses and a method called "signal location search," we detected two unique functionally redundant RNA elements (α and β), each about 75 nt long and separated by 150 nt, in the 3'-terminal coding sequence of RNA polymerase, 3D(pol). The presence of wild type (WT) α or β was sufficient for the optimal growth of PV, but the alteration of both segments in the same virus yielded very low titers and tiny plaques. The nucleotide sequences and predicted RNA structures of α and β have no apparent resemblance to each other. In α, we narrowed down the functional domain to a 48-nt-long, highly conserved segment. The primary determinant of function in β is a stable and highly conserved hairpin. Reporter constructs showed that the α- and β-segments are required for RNA replication. Recoding offers a unique and effective method to search for unknown functional RNA elements in coding sequences of RNA viruses, particularly if the signals are redundant in function.     

Critical function for the Ras-GTPase activating protein RASA3 in vertebrate erythropoiesis and megakaryopoiesis

Phenotype-driven approaches to gene discovery using inbred mice have been instrumental in identifying genetic determinants of inherited blood dyscrasias. The recessive mutant scat (severe combined anemia and thrombocytopenia) alternates between crisis and remission episodes, indicating an aberrant regulatory feedback mechanism common to erythrocyte and platelet formation. Here, we identify a missense mutation (G125V) in the scat Rasa3 gene, encoding a Ras GTPase activating protein (RasGAP), and elucidate the mechanism producing crisis episodes. The mutation causes mislocalization of RASA3 to the cytosol in scat red cells where it is inactive, leading to increased GTP-bound Ras. Erythropoiesis is severely blocked in scat crisis mice, and ~94% succumb during the second crisis (~30 d of age) from catastrophic hematopoietic failure in the spleen and bone marrow. Megakaryopoiesis is also defective during crisis. Notably, the scat phenotype is recapitulated in zebrafish when rasa3 is silenced. These results highlight a critical, conserved, and nonredundant role for RASA3 in vertebrate hematopoiesis.     

Functional analysis of receptor tyrosine kinase mutations in lung cancer identifies oncogenic extracellular domain mutations of ERBB2

We assessed somatic alleles of six receptor tyrosine kinase genes mutated in lung adenocarcinoma for oncogenic activity. Five of these genes failed to score in transformation assays; however, novel recurring extracellular domain mutations of the receptor tyrosine kinase gene ERBB2 were potently oncogenic. These ERBB2 extracellular domain mutants were activated by two distinct mechanisms, characterized by elevated C-terminal tail phosphorylation or by covalent dimerization mediated by intermolecular disulfide bond formation. These distinct mechanisms of receptor activation converged upon tyrosine phosphorylation of cellular proteins, impacting cell motility. Survival of Ba/F3 cells transformed to IL-3 independence by the ERBB2 extracellular domain mutants was abrogated by treatment with small-molecule inhibitors of ERBB2, raising the possibility that patients harboring such mutations could benefit from ERBB2-directed therapy.     

From the Cover: Science in support of the Deepwater Horizon response

This introduction to the Special Feature presents the context for science during the Deepwater Horizon oil spill response, summarizes how scientific knowledge was integrated across disciplines and statutory responsibilities, identifies areas where scientific information was accurate and where it was not, and considers lessons learned and recommendations for future research and response. Scientific information was integrated within and across federal and state agencies, with input from nongovernmental scientists, across a diverse portfolio of needs—stopping the flow of oil, estimating the amount of oil, capturing and recovering the oil, tracking and forecasting surface oil, protecting coastal and oceanic wildlife and habitat, managing fisheries, and protecting the safety of seafood. Disciplines involved included atmospheric, oceanographic, biogeochemical, ecological, health, biological, and chemical sciences, physics, geology, and mechanical and chemical engineering. Platforms ranged from satellites and planes to ships, buoys, gliders, and remotely operated vehicles to laboratories and computer simulations. The unprecedented response effort depended directly on intense and extensive scientific and engineering data, information, and advice. Many valuable lessons were learned that should be applied to future events.     

INAUGURAL ARTICLE by a Recently Elected Academy Member:Nonsteroidal anti-inflammatory drug sensitizes Mycobacterium tuberculosis to endogenous and exogenous antimicrobials

Existing drugs are slow to eradicate Mycobacterium tuberculosis (Mtb) in patients and have failed to control tuberculosis globally. One reason may be that host conditions impair Mtb’s replication, reducing its sensitivity to most antiinfectives. We devised a high-throughput screen for compounds that kill Mtb when its replication has been halted by reactive nitrogen intermediates (RNIs), acid, hypoxia, and a fatty acid carbon source. At concentrations routinely achieved in human blood, oxyphenbutazone (OPB), an inexpensive anti-inflammatory drug, was selectively mycobactericidal to nonreplicating (NR) Mtb. Its cidal activity depended on mild acid and was augmented by RNIs and fatty acid. Acid and RNIs fostered OPB’s 4-hydroxylation. The resultant 4-butyl-4-hydroxy-1-(4-hydroxyphenyl)-2-phenylpyrazolidine-3,5-dione (4-OH-OPB) killed both replicating and NR Mtb, including Mtb resistant to standard drugs. 4-OH-OPB depleted flavins and formed covalent adducts with N-acetyl-cysteine and mycothiol. 4-OH-OPB killed Mtb synergistically with oxidants and several antituberculosis drugs. Thus, conditions that block Mtb’s replication modify OPB and enhance its cidal action. Modified OPB kills both replicating and NR Mtb and sensitizes both to host-derived and medicinal antimycobacterial agents.Some bacterial infections can be cured with a single dose of an antibiotic, and most others can be cured with administration of one drug over several days or weeks. In contrast, routine treatment of drug-sensitive tuberculosis (TB) takes 2 mo of therapy with four drugs and an additional 4 mo with two drugs to reduce the 2-y relapse rate below 5%. The difficulty of completing prolonged treatment is a major reason for emergence of drug resistance. When the infecting strain is resistant to isoniazid and rifampin, the two drugs recommended for all 6 mo of treatment, cure often requires 2 y of daily administration of toxic, expensive, second-line agents that are often unavailable at the point of care. When the causative strain is additionally resistant to a quinolone and an aminoglycoside, the resultant “extensively drug-resistant” TB was fatal to 80% of patients in a leading center (1), although complex multidrug regimens have achieved higher cure rates in populations not previously exposed to the additional drugs (2). In addition to sharing air with someone with TB, leading risk factors for contracting the disease are malnutrition, HIV infection, diabetes, and exposure to smoke from cigarettes or cooking fires (3). These epidemiological challenges exacerbate problems of inadequate diagnostic technology and limited access to drug susceptibility testing and to drugs. Control of the pandemic is not in sight (3).It is widely hypothesized that treatment of TB is protracted because nonreplicating (NR) subpopulations of bacilli are phenotypically tolerant to drugs that were selected for activity against replicating (R) Mycobacterium tuberculosis (Mtb) (4). Mtb can occupy diverse microenvironments in the host. Evidence from auxotrophs, analyses of gene expression, and direct and indirect biochemical measurements in vivo or ex vivo in experimental animals and people suggest that such environments expose Mtb to acid, hypoxia, reactive nitrogen intermediates (RNIs), oxidative stress, carbohydrate deficiency, and metal starvation or intoxication, and require Mtb to metabolize fatty acids or cholesterol (5–17). In vitro, many of the same conditions can make Mtb relatively refractory to killing by the standard agents, except for pyrazinamide, which is only effective at a low pH.Thus, there is a need for a high-throughput screen (HTS) for compounds that kill Mtb when the Mtb has been rendered NR by a combination of physiologically relevant host-imposed conditions. We were encouraged to devise such a screen by recent discoveries of a class of compounds that kill Mtb only when it is NR (18), an antibiotic in clinical use for other infections that kills NR Mtb better than R Mtb (19), and a compound that kills NR and R Mtb equally well (20). Unfortunately, only one of those compounds is an approved drug, and even if it were of proven utility in TB, its price would preclude its use by most of those who need it. We decided to screen other existing drugs that are not regarded as antiinfectives for those that kill NR Mtb. Here, we report finding such a drug in an HTS that combined four host-imposed conditions, some of which converted the drug into a form active on both R and NR Mtb.