Nontarget DNA binding shapes the dynamic landscape for enzymatic recognition of DNA damage

The DNA repair enzyme human uracil DNA glycosylase (UNG) scans short stretches of genomic DNA and captures rare uracil bases as they transiently emerge from the DNA duplex via spontaneous base pair breathing motions. The process of DNA scanning requires that the enzyme transiently loosen its grip on DNA to allow stochastic movement along the DNA contour, while engaging extrahelical bases requires motions on a more rapid timescale. Here, we use NMR dynamic measurements to show that free UNG has no intrinsic dynamic properties in the millisecond to microsecond and subnanosecond time regimes, and that the act of binding to nontarget DNA reshapes the dynamic landscape to allow productive millisecond motions for scanning and damage recognition. These results suggest that DNA structure and the spontaneous dynamics of base pairs may drive the evolution of a protein sequence that is tuned to respond to this dynamic regime

CNF1 Improves Astrocytic Ability to Support Neuronal Growth and Differentiation In vitro

Modulation of cerebral Rho GTPases activity in mice brain by intracerebral administration of Cytotoxic Necrotizing Factor 1 (CNF1) leads to enhanced neurotransmission and synaptic plasticity and improves learning and memory. To gain more insight into the interactions between CNF1 and neuronal cells, we used primary neuronal and astrocytic cultures from rat embryonic brain to study CNF1 effects on neuronal differentiation, focusing on dendritic tree growth and synapse formation, which are strictly modulated by Rho GTPases. CNF1 profoundly remodeled the cytoskeleton of hippocampal and cortical neurons, which showed philopodia-like, actin-positive projections, thickened and poorly branched dendrites, and a decrease in synapse number. CNF1 removal, however, restored dendritic tree development and synapse formation, suggesting that the toxin can reversibly block neuronal differentiation. On differentiated neurons, CNF1 had a similar effacing effect on synapses. Therefore, a direct interaction with CNF1 is apparently deleterious for neurons. Since astrocytes play a pivotal role in neuronal differentiation and synaptic regulation, we wondered if the beneficial in vivo effect could be mediated by astrocytes. Primary astrocytes from embryonic cortex were treated with CNF1 for 48 hours and used as a substrate for growing hippocampal neurons. Such neurons showed an increased development of neurites, in respect to age-matched controls, with a wider dendritic tree and a richer content in synapses. In CNF1-exposed astrocytes, the production of interleukin 1β, known to reduce dendrite development and complexity in neuronal cultures, was decreased. These results demonstrate that astrocytes, under the influence of CNF1, increase their supporting activity on neuronal growth and differentiation, possibly related to the diminished levels of interleukin 1β. These observations suggest that the enhanced synaptic plasticity and improved learning and memory described in CNF1-injected mice are probably mediated by astrocytes

Cell-specific deletion of C1qa identifies microglia as the dominant source of C1q in mouse brain

BACKGROUND: The complement cascade not only provides protection from infection but can also mediate destructive inflammation. Complement is also involved in elimination of neuronal synapses which is essential for proper development, but can be detrimental during aging and disease. C1q, required for several of these complement-mediated activities, is present in the neuropil, microglia, and a subset of interneurons in the brain. METHODS: To identify the source(s) of C1q in the brain, the C1qa gene was selectively inactivated in the microglia or Thy-1(+) neurons in both wild type mice and a mouse model of Alzheimer’s disease (AD), and C1q synthesis assessed by immunohistochemistry, QPCR, and western blot analysis. RESULTS: While C1q expression in the brain was unaffected after inactivation of C1qa in Thy-1(+) neurons, the brains of C1qa (FL/FL) :Cx3cr1 (CreERT2) mice in which C1qa was ablated in microglia were devoid of C1q with the exception of limited C1q in subsets of interneurons. Surprisingly, this loss of C1q occurred even in the absence of tamoxifen by 1 month of age, demonstrating that Cre activity is tamoxifen-independent in microglia in Cx3cr1 (CreERT2/WganJ) mice. C1q expression in C1qa (FL/FL) : Cx3cr1 (CreERT2/WganJ) mice continued to decline and remained almost completely absent through aging and in AD model mice. No difference in C1q was detected in the liver or kidney from C1qa (FL/FL) : Cx3cr1 (CreERT2/WganJ) mice relative to controls, and C1qa (FL/FL) : Cx3cr1 (CreERT2/WganJ) mice had minimal, if any, reduction in plasma C1q. CONCLUSIONS: Thus, microglia, but not neurons or peripheral sources, are the dominant source of C1q in the brain. While demonstrating that the Cx3cr1 (CreERT2/WganJ) deleter cannot be used for adult-induced deletion of genes in microglia, the model described here enables further investigation of physiological roles of C1q in the brain and identification of therapeutic targets for the selective control of complement-mediated activities contributing to neurodegenerative disorders. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s12974-017-0814-9) contains supplementary material, which is available to authorized users

Criteria for evaluation of disease extent by 123I-metaiodobenzylguanidine scans in neuroblastoma: a report for the International Neuroblastoma Risk Group (INRG) Task Force

BackgroundNeuroblastoma is an embryonic tumour of the sympathetic nervous system, metastatic in half of the patients at diagnosis, with a high preponderance of osteomedullary disease, making accurate evaluation of metastatic sites and response to therapy challenging. Metaiodobenzylguanidine (mIBG), taken into cells via the norepinephrine transporter, provides a sensitive and specific method of assessing tumour in both soft tissue and bone sites. The goal of this report was to develop consensus guidelines for the use of mIBG scans in staging, response assessment and surveillance in neuroblastoma.MethodsThe International Neuroblastoma Risk Group (INRG) Task Force, including a multidisciplinary group in paediatric oncology of North and South America, Europe, Oceania and Asia, formed a subcommittee on metastatic disease evaluation, including expert nuclear medicine physicians and oncologists, who developed these guidelines based on their experience and the medical literature, with approval by the larger INRG Task Force.ResultsGuidelines for patient preparation, radiotracer administration, techniques of scanning including timing, energy, specific views, and use of single photon emission computed tomography are included. Optimal timing of scans in relation to therapy and for surveillance is reviewed. Validated semi-quantitative scoring methods in current use are reviewed, with recommendations for use in prognosis and response evaluation.ConclusionsMetaiodobenzylguanidine scans are the most sensitive and specific method of staging and response evaluation in neuroblastoma, particularly when used with a semi-quantitative scoring method. Use of the optimal techniques for mIBG in staging and response, including a semi-quantitative score, is essential for evaluation of the efficacy of new therapy

Complement in the pathogenesis of Alzheimer's disease

The emergence of complement as an important player in normal brain development and pathological remodelling has come as a major surprise to most scientists working in neuroscience and almost all those working in complement. That a system, evolved to protect the host against infection, should have these unanticipated roles has forced a rethink about what complement might be doing in the brain in health and disease, where it is coming from, and whether we can, or indeed should, manipulate complement in the brain to improve function or restore homeostasis. Complement has been implicated in diverse neurological and neuropsychiatric diseases well reviewed elsewhere, from depression through epilepsy to demyelination and dementia, in most complement drives inflammation to exacerbate the disease. Here, I will focus on just one disease, the most common cause of dementia, Alzheimer’s disease. I will briefly review the current understanding of what complement does in the normal brain, noting, in particular, the many gaps in understanding, then describe how complement may influence the genesis and progression of pathology in Alzheimer’s disease. Finally, I will discuss the problems and pitfalls of therapeutic inhibition of complement in the Alzheimer brain

Membrane-mediated interactions

Interactions mediated by the cell membrane between inclusions, such as membrane proteins or antimicrobial peptides, play important roles in their biological activity. They also constitute a fascinating challenge for physicists, since they test the boundaries of our understanding of self-assembled lipid membranes, which are remarkable examples of two-dimensional complex fluids. Inclusions can couple to various degrees of freedom of the membrane, resulting in different types of interactions. In this chapter, we review the membrane-mediated interactions that arise from direct constraints imposed by inclusions on the shape of the membrane. These effects are generic and do not depend on specific chemical interactions. Hence, they can be studied using coarse-grained soft matter descriptions. We deal with long-range membrane-mediated interactions due to the constraints imposed by inclusions on membrane curvature and on its fluctuations. We also discuss the shorter-range interactions that arise from the constraints on membrane thickness imposed by inclusions presenting a hydrophobic mismatch with the membrane.Comment: 38 pages, 10 figures, pre-submission version. In: Bassereau P., Sens P. (eds) Physics of Biological Membranes. Springer, Cha

The role of inflammation in epilepsy.

Epilepsy is the third most common chronic brain disorder, and is characterized by an enduring predisposition to generate seizures. Despite progress in pharmacological and surgical treatments of epilepsy, relatively little is known about the processes leading to the generation of individual seizures, and about the mechanisms whereby a healthy brain is rendered epileptic. These gaps in our knowledge hamper the development of better preventive treatments and cures for the approximately 30% of epilepsy cases that prove resistant to current therapies. Here, we focus on the rapidly growing body of evidence that supports the involvement of inflammatory mediators-released by brain cells and peripheral immune cells-in both the origin of individual seizures and the epileptogenic process. We first describe aspects of brain inflammation and immunity, before exploring the evidence from clinical and experimental studies for a relationship between inflammation and epilepsy. Subsequently, we discuss how seizures cause inflammation, and whether such inflammation, in turn, influences the occurrence and severity of seizures, and seizure-related neuronal death. Further insight into the complex role of inflammation in the generation and exacerbation of epilepsy should yield new molecular targets for the design of antiepileptic drugs, which might not only inhibit the symptoms of this disorder, but also prevent or abrogate disease pathogenesis

β-Amyloid precursor protein (APP) and APP-RNA are rapidly affected by glutamate in cultured neurons - Selective increase of mRNAs encoding a kunitz protease inhibitor domain

Alternative splicing of β-amyloid precursor protein (APP) RNA generates APP isoforms with or without a Kunitz protease inhibitor (KPI) domain. Previously, we showed that KPI (+) APP RNA, but not KPI (-) APP RNA, is upregulated in response to experimental lesions in which neurotoxicity is dependent on NMDA receptor activation and in Alzheimer's disease hippocampus. Recent studies by Mucke et al. (1995) showed that neuronal expression of human KPI (+) APP, but not KPI (-) APP, in transgenic mice is neuroprotective against experimental lesions. In this study we examined the direct effects of the excitotoxic amino acid Glu on alternatively spliced APP RNAs and the corresponding protein isoforms in cultured rat cortical neurons. Glu treatment rapidly induced (4.5 h) KPI (+) APP RNA but not KPI (-) APP RNA. Induction of KPI (+) RNA preceded Glu-induced neuronal cell death and was partially blocked by an NMDA-receptor antagonist. In contrast to the RNA, cellular levels of KPI (+) APP were not changed by 4.5 h of Glu treatment. Instead, the cellular full-length form of the protein KPI (-) APP was reduced by ∼50% after 2 h of Glu treatment and remained depleted after 24 h of treatment. Cellular levels of KPI (+) forms of amyloid precursor-like protein 2 (APLP2) were not changed by Glu treatment. Our data are consistent with the hypothesis that sustained NMDA-receptor activation can regulate alternative splicing of the APP pre-mRNA in neurons. © 1996 Humana Press Inc.link_to_subscribed_fulltex