snakePipes: facilitating flexible, scalable and integrative epigenomic analysis.

SUMMARY: Due to the rapidly increasing scale and diversity of epigenomic data, modular and scalable analysis workflows are of wide interest. Here we present snakePipes, a workflow package for processing and downstream analysis of data from common epigenomic assays: ChIP-seq, RNA-seq, Bisulfite-seq, ATAC-seq, Hi-C and single-cell RNA-seq. snakePipes enables users to assemble variants of each workflow and to easily install and upgrade the underlying tools, via its simple command-line wrappers and yaml files. AVAILABILITY AND IMPLEMENTATION: snakePipes can be installed via conda: `conda install -c mpi-ie -c bioconda -c conda-forge snakePipes'. Source code (https://github.com/maxplanck-ie/snakepipes) and documentation (https://snakepipes.readthedocs.io/en/latest/) are available online. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Epigenetic alterations in longevity regulators, reduced life span, and exacerbated aging-related pathology in old father offspring mice.

Advanced age is not only a major risk factor for a range of disorders within an aging individual but may also enhance susceptibility for disease in the next generation. In humans, advanced paternal age has been associated with increased risk for a number of diseases. Experiments in rodent models have provided initial evidence that paternal age can influence behavioral traits in offspring animals, but the overall scope and extent of paternal age effects on health and disease across the life span remain underexplored. Here, we report that old father offspring mice showed a reduced life span and an exacerbated development of aging traits compared with young father offspring mice. Genome-wide epigenetic analyses of sperm from aging males and old father offspring tissue identified differentially methylated promoters, enriched for genes involved in the regulation of evolutionarily conserved longevity pathways. Gene expression analyses, biochemical experiments, and functional studies revealed evidence for an overactive mTORC1 signaling pathway in old father offspring mice. Pharmacological mTOR inhibition during the course of normal aging ameliorated many of the aging traits that were exacerbated in old father offspring mice. These findings raise the possibility that inherited alterations in longevity pathways contribute to intergenerational effects of aging in old father offspring mice.

Every-other-day feeding extends lifespan but fails to delay many symptoms of aging in mice.

Dietary restriction regimes extend lifespan in various animal models. Here we show that longevity in male C57BL/6J mice subjected to every-other-day feeding is associated with a delayed onset of neoplastic disease that naturally limits lifespan in these animals. We compare more than 200 phenotypes in over 20 tissues in aged animals fed with a lifelong every-other-day feeding or ad libitum access to food diet to determine whether molecular, cellular, physiological and histopathological aging features develop more slowly in every-other-day feeding mice than in controls. We also analyze the effects of every-other-day feeding on young mice on shorter-term every-other-day feeding or ad libitum to account for possible aging-independent restriction effects. Our large-scale analysis reveals overall only limited evidence for a retardation of the aging rate in every-other-day feeding mice. The data indicate that every-other-day feeding-induced longevity is sufficiently explained by delays in life-limiting neoplastic disorders and is not associated with a more general slowing of the aging process in mice.Dietary restriction can extend the life of various model organisms. Here, Xie et al. show that intermittent periods of fasting achieved through every-other-day feeding protect mice against neoplastic disease but do not broadly delay organismal aging in animals.

High-dose maternal folic acid supplementation before conception impairs reversal learning in offspring mice.

Maternal folic acid (FA) supplementation prior to and during gestation is recommended for the prevention of neural tube closure defects in the developing embryo. Prior studies, however, suggested that excessive FA supplementation during gestation can be associated with toxic effects on the developing organism. Here, we address whether maternal dietary folic acid supplementation at 40 mg/kg chow (FD), restricted to a period prior to conception, affects neurobehavioural development in the offspring generation. Detailed behavioural analyses showed reversal learning impairments in the Morris water maze in offspring derived from dams exposed to FD prior to conceiving. Furthermore, offspring of FD dams showed minor and transient gene expression differences relative to controls. Our data suggest that temporary exposure of female germ cells to FD is sufficient to cause impaired cognitive flexibility in the subsequent generation.

deepTools2: a next generation web server for deep-sequencing data analysis.

We present an update to our Galaxy-based web server for processing and visualizing deeply sequenced data. Its core tool set, deepTools, allows users to perform complete bioinformatic workflows ranging from quality controls and normalizations of aligned reads to integrative analyses, including clustering and visualization approaches. Since we first described our deepTools Galaxy server in 2014, we have implemented new solutions for many requests from the community and our users. Here, we introduce significant enhancements and new tools to further improve data visualization and interpretation. deepTools continue to be open to all users and freely available as a web service at deeptools.ie-freiburg.mpg.de The new deepTools2 suite can be easily deployed within any Galaxy framework via the toolshed repository, and we also provide source code for command line usage under Linux and Mac OS X. A public and documented API for access to deepTools functionality is also available.

Limited effects of an eIF2αS51A allele on neurological impairments in the 5xFAD mouse model of Alzheimer's disease.

Alzheimer's disease (AD) has been associated with increased phosphorylation of the translation initiation factor 2α (eIF2α) at serine 51. Increased phosphorylation of eIF2α alters translational control and may thereby have adverse effects on synaptic plasticity, learning, and memory. To analyze if increased levels of p-eIF2α indeed promote AD-related neurocognitive impairments, we crossed 5xFAD transgenic mice with an eIF2α(S51A) knock-in line that expresses the nonphosphorylatable eIF2α variant eIF2α(S51A). Behavioral assessment of the resulting mice revealed motor and cognitive deficits in 5xFAD mice that were, with the possible exception of locomotor hyperactivity, not restored by the eIF2α(S51A) allele. Telemetric intracranial EEG recordings revealed no measurable effects of the eIF2α(S51A) allele on 5xFAD-associated epileptic activity. Microarray-based transcriptome analyses showed clear transcriptional alterations in 5xFAD hippocampus that were not corrected by the eIF2α(S51A) allele. In contrast to prior studies, our immunoblot analyses did not reveal increased levels of p-eIF2α in the hippocampus of 5xFAD mice, suggesting that elevated p-eIF2α levels are not a universal feature of AD models. Collectively, our data indicate that 5xFAD-related pathologies do not necessarily require hyperphosphorylation of eIF2α to emerge; they also show that heterozygosity for the nonphosphorylatable eIF2α(S51A) allele has limited effects on 5xFAD-related disease manifestations.

The anti-diabetic drug metformin reduces BACE1 protein level by interfering with the MID1 complex.

Alzheimer's disease (AD), the most common form of dementia in the elderly, is characterized by two neuropathological hallmarks: senile plaques, which are composed of Aß peptides, and neurofibrillary tangles, which are composed of hyperphosphorylated TAU protein. Diabetic patients with dysregulated insulin signalling are at increased risk of developing AD. Further, several animal models of diabetes show increased Aß expression and hyperphosphorylated tau. As we have shown recently, the anti-diabetic drug metformin is capable of dephosphorylating tau at AD-relevant phospho-sites. Here, we investigated the effect of metformin on the main amyloidogenic enzyme BACE1 and, thus, on the production of Aß peptides, the second pathological hallmark of AD. We find similar results in cultures of primary neurons, a human cell line model of AD and in vivo in mice. We show that treatment with metformin decreases BACE1 protein expression by interfering with an mRNA-protein complex that contains the ubiquitin ligase MID1, thereby reducing BACE1 activity. Together with our previous findings these results indicate that metformin may target both pathological hallmarks of AD and may be of therapeutic value for treating and/or preventing AD.

Rapamycin extends murine lifespan but has limited effects on aging.

Aging is a major risk factor for a large number of disorders and functional impairments. Therapeutic targeting of the aging process may therefore represent an innovative strategy in the quest for novel and broadly effective treatments against age-related diseases. The recent report of lifespan extension in mice treated with the FDA-approved mTOR inhibitor rapamycin represented the first demonstration of pharmacological extension of maximal lifespan in mammals. Longevity effects of rapamycin may, however, be due to rapamycin's effects on specific life-limiting pathologies, such as cancers, and it remains unclear if this compound actually slows the rate of aging in mammals. Here, we present results from a comprehensive, large-scale assessment of a wide range of structural and functional aging phenotypes, which we performed to determine whether rapamycin slows the rate of aging in male C57BL/6J mice. While rapamycin did extend lifespan, it ameliorated few studied aging phenotypes. A subset of aging traits appeared to be rescued by rapamycin. Rapamycin, however, had similar effects on many of these traits in young animals, indicating that these effects were not due to a modulation of aging, but rather related to aging-independent drug effects. Therefore, our data largely dissociate rapamycin's longevity effects from effects on aging itself.

Adult-onset fluoxetine treatment does not improve behavioral impairments and may have adverse effects on the Ts65Dn mouse model of Down syndrome.

Down syndrome is caused by triplication of chromosome 21 and is associated with neurocognitive phenotypes ranging from severe intellectual disability to various patterns of more selective neuropsychological deficits, including memory impairments. In the Ts65Dn mouse model of Down syndrome, excessive GABAergic neurotransmission results in local over-inhibition of hippocampal circuits, which dampens hippocampal synaptic plasticity and contributes to cognitive impairments. Treatments with several GABA(A) receptor antagonists result in increased plasticity and improved memory deficits in Ts65Dn mice. These GABA(A) receptor antagonists are, however, not suitable for clinical applications. The selective serotonin reuptake inhibitor fluoxetine, in contrast, is a widely prescribed antidepressant that can also enhance plasticity in the adult rodent brain by lowering GABAergic inhibition. For these reasons, we wondered if an adult-onset 4-week oral fluoxetine treatment restores spatial learning and memory impairments in Ts65Dn mice. Fluoxetine did not measurably improve behavioral impairments of Ts65Dn mice. On the contrary, we observed seizures and mortality in fluoxetine-treated Ts65Dn mice, raising the possibility of a drug × genotype interaction with respect to these adverse treatment outcomes. Future studies should re-address this in larger animal cohorts and determine if fluoxetine treatment is associated with adverse treatment effects in individuals with Down syndrome.

Episodic neurological channelopathies.

Inherited episodic neurological disorders are often due to mutations in ion channels or their interacting proteins, termed channelopathies. There are a wide variety of such disorders, from those causing paralysis, to extreme pain, to ataxia. A common theme in these is alteration of action potential properties or synaptic transmission and a resulting increased propensity of the resulting tissue to enter into or stay in an altered excitability state. Manifestations of these disorders are triggered by an array of precipitants, all of which stress the particular affected tissue in some way and aid in propelling its activity into an aberrant state. Study of these disorders has aided in the understanding of disease risk factors and elucidated the cause of clinically related sporadic disorders. The findings from study of these disorders will aid in the diagnosis and efficient targeted treatment of affected patients.

Mutations in potassium channel Kir2.6 cause susceptibility to thyrotoxic hypokalemic periodic paralysis.

Thyrotoxic hypokalemic periodic paralysis (TPP) is characterized by acute attacks of weakness, hypokalemia, and thyrotoxicosis of various etiologies. These transient attacks resemble those of patients with familial hypokalemic periodic paralysis (hypoKPP) and resolve with treatment of the underlying hyperthyroidism. Because of the phenotypic similarity of these conditions, we hypothesized that TPP might also be a channelopathy. While sequencing candidate genes, we identified a previously unreported gene (not present in human sequence databases) that encodes an inwardly rectifying potassium (Kir) channel, Kir2.6. This channel, nearly identical to Kir2.2, is expressed in skeletal muscle and is transcriptionally regulated by thyroid hormone. Expression of Kir2.6 in mammalian cells revealed normal Kir currents in whole-cell and single-channel recordings. Kir2.6 mutations were present in up to 33% of the unrelated TPP patients in our collection. Some of these mutations clearly alter a variety of Kir2.6 properties, all altering muscle membrane excitability leading to paralysis.

Giant excised patch recordings of recombinant ion channel currents expressed in mammalian cells.

The giant excised patch variant of patch clamp recording combines microsecond time resolution of macroscopic currents with rapid exchange of the experimental solutions at the intracellular membrane surface. This technique has been applied to a limited number of cell types, including Xenopus oocytes, muscle cells, and photoreceptors. We have applied this technique to recording recombinant ion channel currents expressed in membrane patches excised from HEK293 cell lines. Giant inside-out patch recordings of Na(+) channels and SK(Ca) type calcium-activated potassium channels show high temporal resolution and excellent signal to noise characteristics. This technique will facilitate the study of recombinant ion channels expressed in mammalian cells.