Search

Rationale: We previously identified somatic activating mutations in the KRAS ( Kirsten rat sarcoma viral oncogene homologue ) gene in the endothelium of the majority of human sporadic brain arteriovenous malformations; a disorder characterized by direct connections between arteries and veins. However, whether this genetic abnormality alone is sufficient for lesion formation, as well as how active KRAS signaling contributes to arteriovenous malformations, remains unknown. Objective: To establish the first in vivo models of somatic KRAS gain of function in the endothelium in both mice and zebrafish to directly observe the phenotypic consequences of constitutive KRAS activity at a cellular level in vivo, and to test potential therapeutic interventions for arteriovenous malformations. Methods and Results: Using both postnatal and adult mice, as well as embryonic zebrafish, we demonstrate that endothelial-specific gain of function mutations in Kras (G12D or G12V) are sufficient to induce brain arteriovenous malformations. Active KRAS signaling leads to altered endothelial cell morphogenesis and increased cell size, ectopic sprouting, expanded vessel lumen diameter, and direct connections between arteries and veins. Furthermore, we show that these lesions are not associated with altered endothelial growth dynamics or a lack of proper arteriovenous identity but instead seem to feature exuberant angiogenic signaling. Finally, we demonstrate that KRAS-dependent arteriovenous malformations in zebrafish are refractory to inhibition of the downstream effector PI3K but instead require active MEK (mitogen-activated protein kinase kinase 1) signaling. Conclusions: We demonstrate that active KRAS expression in the endothelium is sufficient for brain arteriovenous malformations, even in the setting of uninjured adult vasculature. Furthermore, the finding that KRAS-dependent lesions are reversible in zebrafish suggests that MEK inhibition may represent a promising therapeutic treatment for arteriovenous malformation patients. Graphical Abstract: A graphical abstract is available for this article.

Rett Syndrome (RTT) is a neurodevelopmental disorder caused by loss of function of the transcriptional regulator Methyl-CpG-Binding Protein 2 (MeCP2). In addition to the characteristic loss of hand function and spoken language after the first year of life, people with RTT also have a variety of physiological and autonomic abnormalities including disrupted breathing rhythms characterized by bouts of hyperventilation and an increased frequency of apnea. These breathing abnormalities, that likely involve alterations in both the circuitry underlying respiratory pace making and those underlying breathing response to environmental stimuli, may underlie the sudden unexpected death seen in a significant fraction of people with RTT. In fact, mice lacking MeCP2 function exhibit abnormal breathing rate response to acute hypoxia and maintain a persistently elevated breathing rate rather than showing typical hypoxic ventilatory decline that can be observed among their wild-type littermates. Using genetic and pharmacological tools to better understand the course of this abnormal hypoxic breathing rate response and the neurons driving it, we learned that the abnormal hypoxic breathing response is acquired as the animals mature, and that MeCP2 function is required within excitatory, inhibitory, and modulatory populations for a normal hypoxic breathing rate response. Furthermore, mice lacking MeCP2 exhibit decreased hypoxia-induced neuronal activity within the nucleus tractus solitarius of the dorsal medulla. Overall, these data provide insight into the neurons driving the circuit dysfunction that leads to breathing abnormalities upon loss of MeCP2. The discovery that combined dysfunction across multiple neuronal populations contributes to breathing dysfunction may provide insight into sudden unexpected death in RTT.

There is a growing awareness of the ecological and biogeochemical importance of fungi in coastal marine systems. While highly diverse fungi have been discovered in these marine systems, still, little is known about their seasonality and associated drivers in coastal waters. Here, we examined fungal communities over 3 years of weekly sampling at a dynamic, temperate coastal site (Pivers Island Coastal Observatory [PICO], Beaufort, NC, USA). Fungal 18S rRNA gene abundance, operational taxonomic unit (OTU) richness, and Shannon's diversity index values exhibited prominent seasonality. Fungal 18S rRNA gene copies peaked in abundance during the summer and fall, with positive correlations with chlorophyll a, SiO(4), and oxygen saturation. Diversity (measured using internal transcribed spacer [ITS] libraries) was highest during winter and lowest during summer; it was linked to temperature, pH, chlorophyll a, insolation, salinity, and dissolved inorganic carbon (DIC). Fungal communities derived from ITS libraries were dominated throughout the year by Ascomycota, with contributions from Basidiomycota, Chytridiomycota, and Mucoromycotina, and their seasonal patterns linked to water temperature, light, and the carbonate system. Network analysis revealed that while cooccurrence and exclusion existed within fungus networks, exclusion dominated the fungus-and-phytoplankton network, in contrast with reported pathogenic and nutritional interactions between marine phytoplankton and fungi. Compared with the seasonality of bacterial communities in the same samples, the timing, extent, and associated environmental variables for fungi community are unique. These results highlight the fungal seasonal dynamics in coastal water and improve our understanding of the ecology of planktonic fungi. IMPORTANCE Coastal fungal dynamics were long assumed to be due to terrestrial inputs; here, a high-resolution time series reveals strong, repeating annual patterns linked to in situ environmental conditions, arguing for a resident coastal fungal community shaped by environmental factors. These seasonal patterns do, however, differ from those observed in the bacterioplankton at the same site; e.g., fungal diversity peaks in winter, whereas bacterial diversity maxima occur in the spring and fall. While the dynamics of these communities are linked to water temperature and insolation, fungi are also influenced by the carbonate system (pH and DIC). As both fungi and heterotrophic bacteria are thought to be key organic-material metabolizers, differences in their environmental drivers may offer clues as to which group dominates secondary production at this dynamic site. Overall, this study suggests the unique ecological roles of mycoplankton and their potentially broad niche complementarities to other microbial groups in the coastal ocean.

Marine microbes exhibit seasonal cycles in community composition, yet the key drivers of these patterns and microbial population fidelity to specific environmental conditions remain to be determined. To begin addressing these questions, we characterized microbial dynamics weekly for 3 years at a temperate, coastal site with dramatic environmental seasonality. This high-resolution time series reveals that changes in microbial community composition are not continuous; over the duration of the time series, the community instead resolves into distinct summer and winter profiles with rapid spring and fall transitions between these states. Here, we show that these community shifts involve switching between closely related strains that exhibit either summer or winter preferences. Moreover, taxa repeat this process annually in both this and another temperate coastal time series, suggesting that this phenomenon may be widespread in marine ecosystems. To address potential biogeochemical impacts of these community changes, PICRUSt-based metagenomes predict seasonality in transporters, photosynthetic proteins, peptidases and carbohydrate metabolic pathways in spite of closely related summer- and winter-associated taxa. Thus, even small temperature shifts, such as those predicted by climate change models, could affect both the structure and function of marine ecosystems.

Neurobehavioral disorders comprised of neurodegenerative, neurodevelopmental, and psychiatric disorders together represent leading causes of morbidity and mortality. Despite significant academic research and industry efforts to elucidate the disease mechanisms operative in these disorders and to develop mechanism-based therapies, our understanding remains incomplete and our access to tractable therapeutic interventions severely limited. The magnitude of these short-comings can be measured by the growing list of disappointing clinical trials based on initially promising compounds identified in genetic animal models. This review and commentary will explore why this may be so, focusing on the central role that genetic models of neurobehavioral disorders have come to occupy in current efforts to identify disease mechanisms and therapies. In particular, we will highlight the unique pitfalls and challenges that have hampered success in these models as compared to genetic models of non-neurological diseases as well as to symptom-based models of the early 20th century that led to the discovery of all major classes of psychoactive pharmaceutical compounds still used today. Using examples from specific genetic rodent models of human neurobehavioral disorders, we will highlight issues of reproducibility, construct validity, and translational relevance in the hopes that these examples will be instructive toward greater success in future endeavors. Lastly, we will champion a two-pronged approach toward identifying novel therapies for neurobehavioral disorders that makes greater use of the historically more successful symptom-based approaches in addition to more mechanism-based approaches.

Breathing disturbances are a major challenge in Rett Syndrome (RTT). These disturbances are more pronounced during wakefulness; but irregular breathing occurs also during sleep. During the day patients can exhibit alternating bouts of hypoventilation and irregular hyperventilation. But there is significant individual variability in severity, onset, duration and type of breathing disturbances. Research in mouse models of RTT suggests that different areas in the ventrolateral medulla and pons give rise to different aspects of this breathing disorder. Pre-clinical experiments in mouse models that target different neuromodulatory and neurotransmitter receptors and MeCP2 function within glia cells can partly reverse breathing abnormalities. The success in animal models raises optimism that one day it will be possible to control or potentially cure the devastating symptoms also in human patients with RTT.

Rett Syndrome (RTT) is a neurodevelopmental disorder characterized by loss of acquired skills during development, autonomic dysfunction, and an increased risk for premature lethality. Clinical experience identified a subset of individuals with RTT that present with urological dysfunction including individuals with frequent urinary tract infections, kidney stones, and urine retention requiring frequent catheterization for bladder voiding. To determine if urologic dysfunction is a feature of RTT, we queried the Rett Syndrome Natural History Study, a repository of clinical data from over 1000 individuals with RTT and found multiple instances of urological dysfunction. We then evaluated urological function in a mouse model of RTT and found an abnormal pattern of micturition. Both male and female mice possessing Mecp2 mutations show a decrease in urine output per micturition event. Furthermore, we identified signs of kidney failure secondary to urethral obstruction. Although genetic strain background significantly affects both survival and penetrance of the urethral obstruction phenotype, survival and penetrance of urethral obstruction do not directly correlate. We have identified an additional phenotype caused by loss of MeCP2, urological dysfunction. Furthermore, we urge caution in the interpretation of survival data as an endpoint in preclinical studies, especially where causes of mortality are poorly characterized.

Sudden unexpected death occurs in one quarter of deaths in Rett Syndrome (RTT), a neurodevelopmental disorder caused by mutations in Methyl-CpG-binding protein 2 (MECP2). People with RTT show a variety of autonomic nervous system (ANS) abnormalities and mouse models show similar problems including QTc interval prolongation and hypothermia. To explore the role of cardiac problems in sudden death in RTT, we characterized cardiac rhythm in mice lacking Mecp2 function. Male and female mutant mice exhibited spontaneous cardiac rhythm abnormalities including bradycardic events, sinus pauses, atrioventricular block, premature ventricular contractions, non-sustained ventricular arrhythmias, and increased heart rate variability. Death was associated with spontaneous cardiac arrhythmias and complete conduction block. Atropine treatment reduced cardiac arrhythmias in mutant mice, implicating overactive parasympathetic tone. To explore the role of MeCP2 within the parasympathetic neurons, we selectively removed MeCP2 function from cholinergic neurons (MeCP2 ChAT KO), which recapitulated the cardiac rhythm abnormalities, hypothermia, and early death seen in RTT male mice. Conversely, restoring MeCP2 only in cholinergic neurons rescued these phenotypes. Thus, MeCP2 in cholinergic neurons is necessary and sufficient for autonomic cardiac control, thermoregulation, and survival, and targeting the overactive parasympathetic system may be a useful therapeutic strategy to prevent sudden unexpected death in RTT.

textabstractAutosomal dominant polycystic kidney disease (ADPKD) is a common genetic disorder that frequently results in renal fallure due to progressive cyst development. The major locus, PKD1, maps to 16p13.3. We identified a chromosome translocation associated with ADPKD that disrupts a gene (PBP) encoding a 14 kb transcript in the PKD1 candidate region. Further mutations of the PBP gene were found in PKD1 patients, two deletions (one a de novo event) and a splicing defect, confirming that PBP is the PKD1 gene. This gene is located adjacent to the TSC2 locus in a genomic region that is reiterated more proximally on 16p. The duplicate area encodes three transcripts substantially homologous to the PKD1 transcript. Partial sequence analysis of the PKD1 transcript shows that it encodes a novel protein whose function is at present unknown.

Mouse models of the transcriptional modulator Methyl-CpG-Binding Protein 2 (MeCP2) have advanced our understanding of Rett syndrome (RTT). RTT is a 'prototypical' neurodevelopmental disorder with many clinical features overlapping with other intellectual and developmental disabilities (IDD). Therapeutic interventions for RTT may therefore have broader applications. However, the reliance on the laboratory mouse to identify viable therapies for the human condition may present challenges in translating findings from the bench to the clinic. In addition, the need to identify outcome measures in well-chosen animal models is critical for preclinical trials. Here, we report that a novel Mecp2 rat model displays high face validity for modelling psychomotor regression of a learned skill, a deficit that has not been shown in Mecp2 mice. Juvenile play, a behavioural feature that is uniquely present in rats and not mice, is also impaired in female Mecp2 rats. Finally, we demonstrate that evaluating the molecular consequences of the loss of MeCP2 in both mouse and rat may result in higher predictive validity with respect to transcriptional changes in the human RTT brain. These data underscore the similarities and differences caused by the loss of MeCP2 among divergent rodent species which may have important implications for the treatment of individuals with disease-causing MECP2 mutations. Taken together, these findings demonstrate that the Mecp2 rat model is a complementary tool with unique features for the study of RTT and highlight the potential benefit of cross-species analyses in identifying potential disease-relevant preclinical outcome measures.

The phosphatidylinositol-3-kinase/Akt/mammalian target of rapamycin (PI3K/Akt/mTOR) signaling pathway is activated in more than88% of glioblastomas (GBM). New drugs targeting this pathway are currently in clinical trials. However, noninvasive assessment of treatment response remains challenging. By using magnetic resonance spectroscopy (MRS), PI3K/Akt/mTOR pathway inhibition was monitored in 3 GBM cell lines (GS-2, GBM8, and GBM6; each with a distinct pathway activating mutation) through the measurement of 2 mechanistically linked MR biomarkers: phosphocholine (PC) and hyperpolarized lactate.(31)P MRS studies showed that treatment with the PI3K inhibitor LY294002 induced significant decreases in PC to 34 %± 9% of control in GS-2 cells, 48% ± 5% in GBM8, and 45% ± 4% in GBM6. The mTOR inhibitor everolimus also induced a significant decrease in PC to 62% ± 14%, 57% ± 1%, and 58% ± 1% in GS-2, GBM8, and GBM6 cells, respectively. Using hyperpolarized (13)C MRS, we demonstrated that hyperpolarized lactate levels were significantly decreased following PI3K/Akt/mTOR pathway inhibition in all 3 cell lines to 51% ± 10%, 62% ± 3%, and 58% ± 2% of control with LY294002 and 72% ± 3%, 61% ± 2%, and 66% ± 3% of control with everolimus in GS-2, GBM8, and GBM6 cells, respectively. These effects were mediated by decreases in the activity and expression of choline kinase α and lactate dehydrogenase, which respectively control PC and lactate production downstream of HIF-1. Treatment with the DNA damaging agent temozolomide did not have an effect on either biomarker in any cell line. This study highlights the potential of PC and hyperpolarized lactate as noninvasive MR biomarkers of response to targeted inhibitors in GBM.

Numerous mechanism-based anticancer drugs that target the phosphatidylinositol-3-kinase (PI3K) signaling pathway are in clinical trials. However, assessment of response using traditional imaging methods remains a challenge, as it is often associated with tumor stasis. Here we show for the first time the efficacy of hyperpolarized 13C magnetic resonance spectroscopy (MRS) in detecting the effect of PI3K signaling inhibition by monitoring hyperpolarized [1-13C]-lactate levels produced from hyperpolarized [1-13C]-pyruvate through lactate dehydrogenase (LDH) activity. Studies were performed on GS-2 glioblastoma and MDA-MB-231 breast adenocarcinoma cells. Following inhibition of signaling, hyperpolarized lactate dropped significantly to 42±12% and 76±5% in GS-2 cells treated with LY294002 and everolimus respectively and to 71±15% in MDA-MB-231 cells treated with LY294002. This correlated with a drop in LDH activity to 48±4%, 63±4% and 69±12%, and was associated with a drop in LDHA mRNA levels and LDHA and HIF-1α protein levels. Taken together the data demonstrate that hyperpolarized 13C MRS of pyruvate can be used to monitor the drop in LDHA activity and expression following reduced HIF-1α levels resulting from PI3K signal inhibition. In line with these findings, in vivo studies of GS-2 subcutaneous xenografts treated with everolimus resulted in inhibition in tumor growth, which was associated with a drop in the hyperpolarized lactate-to-pyruvate ratio. In contrast an increase in the ratio was detected in controls. This works demonstrates the potential of hyperpolarized 13C MRS for noninvasive imaging of drug target modulation by treatments that modulate PI3K signaling and HIF-1α levels.

arising from N. C. Derecki et al. , 105–109 (2012); doi:10.1038/nature10907 Rett syndrome is a severe neurodevelopmental disorder caused by mutations in the X chromosomal gene MECP2 (ref. 1), and its treatment so far is symptomatic. Mecp2 disruption in mice phenocopies major features of the syndrome2 that can be reversed after Mecp2 re-expression3. Recently, Derecki et al.4 reported that transplantation of wild-type bone marrow into lethally irradiated Mecp2-null (Mecp2tm1.1Jae/y) mice prevented neurological decline and early death by restoring microglial phagocytic activity against apoptotic targets4, and clinical trials of bone marrow transplantation (BMT) for patients with Rett syndrome have thus been initiated5. We aimed to replicate and extend the BMT experiments in three different Rett syndrome mouse models, but found that despite robust microglial engraftment, BMT from wild-type donors did not prevent early death or ameliorate neurological deficits. Furthermore, early and specific Mecp2 genetic expression in microglia did not rescue Mecp2-deficient mice.

When bacterial cells are stressed by a change in the environment, they respond by changing the activity of enzymes at both the transcriptional and post-transcriptional levels. The UVA component (400–315 nm) of solar radiation reaching the Earth’s surface is one of the most common stresses encountered by bacteria in their environment. Bacteria have evolved various antioxidant defense systems to increase survival when subjected to the deleterious effects of UVA irradiation. Recently, UVA-induced cytotoxicity and oxidative damage have been shown to be dependent on radiation intensity and dose distribution, not just total energy dose. We now report that when Escherichia coli is subjected to continuous sublethal, low-fluence UVA irradiation (7.4 W/m2) while growing to stationary phase, it responds by changing the activity levels of hydroperoxidases (HPI, HPII), glutathione reductase and manganese superoxide dismutase. This leads to an attenuation of the growth-delay response and an increase resistance to lethal UVA irradiation. When E. coli is given a UVA dose of 135 kJ/m2 delivered at a fluence rate of 50 W/m2, extensive protein oxidation occurs which may contribute to the inhibition of key cellular enzymes, leading to cellular dysfunction, DNA damage and eventually death. Changes in antioxidant enzymes induced by low-fluence UVA irradiation do not confer greater protection from protein oxidation after a challenge dose of UVA irradiation delivered at a fluence rate of 50 W/m2.

One quarter of deaths in Rett Syndrome (RTT), an X-linked neurodevelopmental disorder, are sudden and unexpected. RTT is associated with prolonged QTc interval (LQT), and LQT-associated cardiac arrhythmias are a potential cause of unexpected death. Standard of care for LQT in RTT is treatment with β-adrenergic antagonists; however, recent work indicates that acute treatment of mice with RTT with a β-antagonist, propranolol, did not prevent lethal arrhythmias. In contrast, acute treatment with a sodium channel blocker, phenytoin, prevented arrhythmias. Chronic dosing of propranolol may be required for efficacy; therefore, we tested the efficacy of chronic treatment with either propranolol or phenytoin on RTT mice. Phenytoin completely abolished arrhythmias, whereas propranolol showed no benefit. Surprisingly, phenytoin also normalized weight and activity, but worsened breathing patterns. To explore the role of sodium channel blockers on QT in people with RTT, we performed a retrospective analysis of QT status before and after sodium channel blocker antiepileptic therapies. Individuals with RTT and LQT significantly improved their QT interval status after being started on sodium channel blocker antiepileptic therapies. Thus, sodium channel blockers should be considered for the clinical management of LQT in individuals with RTT.

Rett syndrome (RTT), an X-linked postnatal disorder, results from mutations in Methyl CpG-binding protein 2 (MECP2). Survival and breathing in Mecp2(NULL/Y) animals are improved by an N-terminal tripeptide of insulin-like growth factor I (IGF-I) treatment. We determined that Mecp2(NULL/Y) animals also have a metabolic syndrome and investigated whether IGF-I treatment might improve this phenotype. Mecp2(NULL/Y) mice were treated with a full-length IGF-I modified with the addition of polyethylene glycol (PEG-IGF-I), which improves pharmacological properties. Low-dose PEG-IGF-I treatment slightly improved lifespan and heart rate in Mecp2(NULL/Y) mice; however, high-dose PEG-IGF-I decreased lifespan. To determine whether insulinotropic off-target effects of PEG-IGF-I caused the detrimental effect, we treated Mecp2(NULL/Y) mice with insulin, which also decreased lifespan. Thus, the clinical benefit of IGF-I treatment in RTT may critically depend on the dose used, and caution should be taken when initiating clinical trials with these compounds because the beneficial therapeutic window is narrow.

Rett syndrome (RTT) is an X-linked neurological disorder caused by mutations in the gene encoding the transcriptional modulator methyl-CpG-binding protein 2 (MeCP2). Typical RTT primarily affects girls and is characterized by a brief period of apparently normal development followed by the loss of purposeful hand skills and language, the onset of anxiety, hand stereotypies, autistic features, seizures and autonomic dysfunction. Mecp2 mouse models have extensively been studied to demonstrate the functional link between MeCP2 dysfunction and RTT pathogenesis. However, the majority of studies have focused primarily on the molecular and behavioral consequences of the complete absence of MeCP2 in male mice. Studies of female Mecp2(+/-) mice have been limited because of potential phenotypic variability due to X chromosome inactivation effects. To determine whether reproducible and reliable phenotypes can be detected Mecp2(+/-) mice, we analyzed Mecp2(+/-) mice of two different F1 hybrid isogenic backgrounds and at young and old ages using several neurobehavioral and physiological assays. Here, we report a multitude of phenotypes in female Mecp2(+/-) mice, some presenting as early as 5 weeks of life. We demonstrate that Mecp2(+/-) mice recapitulate several aspects of typical RTT and show that mosaic expression of MeCP2 does not preclude the use of female mice in behavioral and molecular studies. Importantly, we uncover several behavioral abnormalities that are present in two genetic backgrounds and report on phenotypes that are unique to one background. These findings provide a framework for pre-clinical studies aimed at improving the constellation of phenotypes in a mouse model of RTT.

Rett syndrome is a neurodevelopmental disorder typically caused by mutations in methyl-CpG-binding protein 2 (MECP2) in which 26% of deaths are sudden and of unknown cause. To explore the hypothesis that these deaths may be due to cardiac dysfunction, we characterized the electrocardiograms in 379 people with Rett syndrome and found that 18.5% show prolongation of the corrected QT interval (QTc), an indication of a repolarization abnormality that can predispose to the development of an unstable fatal cardiac rhythm. Male mice lacking MeCP2 function, Mecp2(Null/Y), also have prolonged QTc and show increased susceptibility to induced ventricular tachycardia. Female heterozygous null mice, Mecp2(Null/+), show an age-dependent prolongation of QTc associated with ventricular tachycardia and cardiac-related death. Genetic deletion of MeCP2 function in only the nervous system was sufficient to cause long QTc and ventricular tachycardia, implicating neuronally mediated changes to cardiac electrical conduction as a potential cause of ventricular tachycardia in Rett syndrome. The standard therapy for prolonged QTc in Rett syndrome, β-adrenergic receptor blockers, did not prevent ventricular tachycardia in Mecp2(Null/Y) mice. To determine whether an alternative therapy would be more appropriate, we characterized cardiomyocytes from Mecp2(Null/Y) mice and found increased persistent sodium current, which was normalized when cells were treated with the sodium channel-blocking anti-seizure drug phenytoin. Treatment with phenytoin reduced both QTc and sustained ventricular tachycardia in Mecp2(Null/Y) mice. These results demonstrate that cardiac abnormalities in Rett syndrome are secondary to abnormal nervous system control, which leads to increased persistent sodium current. Our findings suggest that treatment in people with Rett syndrome would be more effective if it targeted the increased persistent sodium current to prevent lethal cardiac arrhythmias.

Rett syndrome is a neurodevelopmental disorder caused by mutations in methyl-CpG-binding protein 2 (MECP2), a transcriptional regulator. In addition to cognitive, communication, and motor problems, affected individuals have abnormalities in autonomic function and respiratory control that may contribute to premature lethality. Mice lacking Mecp2 die early and recapitulate the autonomic and respiratory phenotypes seen in humans. The association of autonomic and respiratory deficits with premature death suggests that Mecp2 is critical within autonomic and respiratory control centers for survival. To test this, we compared the autonomic and respiratory phenotypes of mice with a null allele of Mecp2 to mice with Mecp2 removed from their brainstem and spinal cord. We found that MeCP2 is necessary within the brainstem and spinal cord for normal lifespan, normal control of heart rate, and respiratory response to hypoxia. Restoration of MeCP2 in a subset of the cells in this same region is sufficient to rescue abnormal heart rate and abnormal respiratory response to hypoxia. Furthermore, restoring MeCP2 function in neural centers critical for autonomic and respiratory function alleviates the lethality associated with loss of MeCP2 function, supporting the notion of targeted therapy toward treating Rett syndrome.

Spinocerebellar ataxia type 1 (SCA1) is a neurodegenerative disease caused by the expansion of a CAG repeat encoding a polyglutamine tract in Ataxin-1 (ATXN1). Both WT and mutant ATXN1 interact with 14-3-3 proteins, and 14-3-3 overexpression stabilizes ATXN1 levels in cells and increases ATXN1 toxicity in flies. To determine whether reducing 14-3-3 levels might mitigate SCA1 pathogenesis, we bred Sca1 154Q/+ mice to mice lacking one allele of 14-3-3 ε. 14-3-3 ε haploinsufficiency rescued cerebellar pathology and motor phenotypes but, surprisingly, not weight loss, respiratory dysfunction, or premature lethality. Biochemical studies revealed that reducing 14-3-3 ε levels exerted different effects in two brain regions especially vulnerable in SCA1: Although diminishing levels of both WT and mutant ATXN1 in the cerebellum, 14-3-3 ε haploinsufficiency did not alter ATXN1 levels in the brainstem. Furthermore, 14-3-3 ε haploinsufficiency decreased the incorporation of expanded ATXN1 into its large toxic complexes in the cerebellum but not in the brainstem, and the distribution of ATXN1’s small and large native complexes differed significantly between the two regions. These data suggest that distinct pathogenic mechanisms operate in different vulnerable brain regions, adding another level of complexity to SCA1 pathogenesis.

Rett syndrome (RTT) is characterized by specific motor, cognitive, and behavioral deficits. Because several of these abnormalities occur in other disease states associated with alterations in aminergic neurotransmitters, we investigated the contribution of such alterations to RTT pathogenesis. We found that both individuals with RTT and Mecp2 -null mice have lower-than-normal levels of aminergic metabolites and content. Deleting Mecp2 from either TH-positive dopaminergic and noradrenergic neurons or PET1-positive serotonergic neurons in mice decreased corresponding neurotransmitter concentration and specific phenotypes, likely through MeCP2 regulation of rate-limiting enzymes involved in aminergic neurotransmitter production. These data support a cell-autonomous, MeCP2-dependent mechanism for the regulation of aminergic neurotransmitter synthesis contributing to unique behavioral phenotypes.

During the course of a day human skin is exposed to solar UV radiation that fluctuates in fluence rate within the UVA (290-315 nm) and UVB (315-400 nm) spectrum. Variables affecting the fluence rate reaching skin cells include differences in UVA and UVB penetrating ability, presence or absence of sunscreens, atmospheric conditions, and season and geographical location where the exposure occurs. Our study determined the effect of UVA fluence rate in solar-simulated (SSR) and tanning-bed radiation (TBR) on four indicators of oxidative stress---protein oxidation, glutathione, heme oxygenase-1, and reactive oxygen species--in human dermal fibroblasts after receiving equivalent UVA and UVB doses. Our results show that the higher UVA fluence rate in TBR increases the level of all four indicators of oxidative stress. In sequential exposures when cells are exposed first to SSR, the lower UVA fluence rate in SSR induces a protective response that protects against oxidative stress following a second exposure to a higher UVA fluence rate. Our studies underscore the important role of UVA fluence rate in determining how human skin cells respond to a given dose of radiation containing both UVA and UVB radiation.

Nature 521, E1–E4 (2015); doi:10.1038/nature14444 In this Brief Communication Arising, the first name of author Sebastien Vingeau was misspelled ‘Sebastian’. In addition, the labels (‘WT→KO’ and ‘KO→WT’) of the two bottom panels in Extended Data Figure 1b were swapped. Both errors have been corrected online.

In Escherichia coli, Δfur (ferric uptake regulator) mutants are hypersensitive to various oxidative agents, including UVA radiation (400–315 nm). Studies suggest that UVA radiation mediates its biological effects on bacteria via oxidative mechanisms that lead to reactive oxygen species, including the superoxide anion radical ( O 2 - ) , hydroxyl radical (HO ), hydrogen peroxide (H2O2) and singlet oxygen (1O2). There is accumulating evidence that Fur may play an important role in the defense against UVA radiation. In addition to regulating almost all genes directly involved in iron acquisition, Fur also regulates the expression of manganese and iron superoxide dismutase (MnSOD, FeSOD), key enzymes in the defense against oxygen toxicity in E. coli. In Δfur mutants, there is a complete absence of FeSOD. Previous results suggest that the native iron chelating agent, enterobactin, which exists in increased levels in Δfur mutants, is an endogenous chromophore for UVA, releasing Fe2+ into the cytoplasm to catalyze the production of highly reactive hydroxyl radicals. We now report that the hypersensitivity of Δfur mutants to UVA irradiation is associated with reduced hydroperoxidase I (HPI) and hydroperoxidase II (HPII) activity, and is associated with a decrease in the transcription of katE and katG genes. The observed decrease in HPII activity in Δfur mutants is also associated with reduced rpoS gene transcription. This study provides additional evidence that the Fur gene product, in addition to its known regulatory effect on the expression of SOD and iron uptake mechanisms, also regulates HPI and HPII activity levels in E. coli. An H2O2-inducible antioxidant defense system leading to an increase in HPI activity, is unaltered in Δfur mutants.

Introduction 17-allyamino-17-demethoxygeldanamycin (17-AAG), a small molecule inhibitor of Hsp90, is currently in clinical trials in breast cancer. However, 17-AAG treatment often results in inhibition of tumor growth rather than shrinkage, making detection of response a challenge. Magnetic resonance spectroscopy (MRS) and spectroscopic imaging (MRSI) are noninvasive imaging methods than can be used to monitor metabolic biomarkers of drug-target modulation. This study set out to examine the MRS-detectable metabolic consequences of Hsp90 inhibition in a breast cancer model. Methods MCF-7 breast cancer cells were investigated, and MRS studies were performed both on live cells and on cell extracts. 31P and 1H MRS were used to determine total cellular metabolite concentrations and 13C MRS was used to probe the metabolism of [1,2-13C]-choline. To explain the MRS metabolic findings, microarray and RT-PCR were used to analyze gene expression, and in vitro activity assays were performed to determine changes in enzymatic activity following 17-AAG treatment. Results Treatment of MCF-7 cells with 17-AAG for 48 hours caused a significant increase in intracellular levels of choline (to 266 ± 18% of control, P = 0.05) and phosphocholine (PC; to 181 ± 10% of control, P = 0.001) associated with an increase in expression of choline transporter SLC44A1 and an elevation in the de novo synthesis of PC. We also detected an increase in intracellular levels of glycerophosphocholine (GPC; to 176 ± 38% of control, P = 0.03) associated with an increase in PLA2 expression and activity. Conclusions This study determined that in the MCF-7 breast cancer model inhibition of Hsp90 by 17-AAG results in a significant MRS-detectable increase in choline, PC and GPC, which is likely due to an increase in choline transport into the cell and phospholipase activation. 1H MRSI can be used in the clinical setting to detect levels of total choline-containing metabolite (t-Cho, composed of intracellular choline, PC and GPC). As Hsp90 inhibitors enter routine clinical use, t-Cho could thus provide an easily detectable, noninvasive metabolic biomarker of Hsp90 inhibition in breast cancer patients.