Genome-wide nucleosome map and cytosine methylation levels of an ancient human genome

Epigenetic information is available from contemporary organisms, but is difficult to track back in evolutionary time. Here, we show that genome-wide epigenetic information can be gathered directly from next-generation sequence reads of DNA isolated from ancient remains. Using the genome sequence data generated from hair shafts of a 4000-yr-old Paleo-Eskimo belonging to the Saqqaq culture, we generate the first ancient nucleosome map coupled with a genome-wide survey of cytosine methylation levels. The validity of both nucleosome map and methylation levels were confirmed by the recovery of the expected signals at promoter regions, exon/intron boundaries, and CTCF sites. The top-scoring nucleosome calls revealed distinct DNA positioning biases, attesting to nucleotide-level accuracy. The ancient methylation levels exhibited high conservation over time, clustering closely with modern hair tissues. Using ancient methylation information, we estimated the age at death of the Saqqaq individual and illustrate how epigenetic information can be used to infer ancient gene expression. Similar epigenetic signatures were found in other fossil material, such as 110,000- to 130,000-yr-old bones, supporting the contention that ancient epigenomic information can be reconstructed from a deep past. Our findings lay the foundation for extracting epigenomic information from ancient samples, allowing shifts in epialleles to be tracked through evolutionary time, as well as providing an original window into modern epigenomics.Ancient DNA research started in the mid-1980s with the successful cloning and sequencing of a short mitochondrial DNA fragment from the quagga zebra, a species that became extinct in the early 20th century (Higuchi et al. 1984). Soon after, the invention of PCR unlocked access to this fragmented and degraded DNA material (Pääbo 1989), making it possible to amplify short gene markers of interest and compare their sequence to that from extant organisms. This illuminated a range of topics ranging from the reconstruction of the evolutionary origins of several now-extinct iconic mammals (Orlando et al. 2003; Krause et al. 2006), the evaluation of the possible role played by major past climatic changes in driving mega-fauna extinctions (Shapiro et al. 2004; Campos et al. 2010; Lorenzen et al. 2011), to the identification of the pathogens responsible for massive historical outbreaks (Taubenberger et al. 1997).However, before the advent of next-generation sequencing (NGS) platforms, the amount of ancient sequence information one had access to was limited to several tens of thousands of nucleotides at best (Noonan et al. 2005, 2006), and until very recently, sequencing whole ancient mitochondrial genomes was considered a major achievement (Cooper et al. 2001; Krause et al. 2006). Parallel sequencing of millions to billions of short DNA fragments has revolutionized ancient DNA research, and today a series of ancient genomes has been reconstructed from humans (Rasmussen et al. 2010, 2011; Keller et al. 2012; Raghavan et al. 2013), archaic hominins (Green et al. 2010; Reich et al. 2010; Meyer et al. 2012), the woolly mammoth (Miller et al. 2008), and several microbial pathogens (Bos et al. 2011; Martin et al. 2013; Schuenemann et al. 2013; Yoshida et al. 2013). Those mainly date back to recent historical periods or the Late Pleistocene, but most recently, the characterization of a 560,000- to 780,000-yr-old horse draft genome revealed that genomic information could be retrieved over much longer evolutionary time scales, probably up until the last million years (Orlando et al. 2013).Ancient genomes have provided important new insights into human evolution and dispersals (Rasmussen et al. 2010, 2011; Keller et al. 2012; Raghavan et al. 2013), revealing an admixture between contemporary human ancestors and archaic hominins (Green et al. 2010; Reich et al. 2010; Meyer et al. 2012) and multiple early human expansions into both Asia and North America (Rasmussen et al. 2010, 2011). The information gained from these samples has largely been limited to nucleotide polymorphisms. Unlike mutations, epigenetic modifications do not alter the underlying DNA sequence, but can be inherited across cell divisions and from parents to offspring and can control gene expression by reshaping cytosine methylation landscapes, nucleosome organization, and histone modification patterns. The range of biological processes that depend on some level of epigenetic regulation is diverse and includes imprinting (Bird 2002), transposition (Hollister and Gaut 2009), cell differentiation (Meissner et al. 2008), and cancer (Teschendorff et al. 2011). In this study, we use the Saqqaq genome that was retrieved from an ∼4000-yr-old tuft of hair of a Paleo-Eskimo from Greenland and sequenced to an average depth of 20× (Rasmussen et al. 2010). We demonstrate that NGS data can be used in the absence of bisulfite or further experimental treatment to directly infer genome-wide nucleosome organization and regional methylation levels, thereby providing the first survey of an ancient epigenome.     

In-season strength maintenance training increases well-trained cyclists’ performance

We investigated the effects of strength maintenance training on thigh muscle cross-sectional area (CSA), leg strength, determinants of cycling performance, and cycling performance. Well-trained cyclists completed either (1) usual endurance training supplemented with heavy strength training twice a week during a 12-week preparatory period followed by strength maintenance training once a week during the first 13 weeks of a competition period (E + S; n = 6 [♂ = 6]), or (2) usual endurance training during the whole intervention period (E; n = 6 [♂ = 5, ♀ = 1]). Following the preparatory period, E + S increased thigh muscle CSA and 1RM (p < 0.05), while no changes were observed in E. Both groups increased maximal oxygen consumption and mean power output in the 40-min all-out trial (p < 0.05). At 13 weeks into the competition period, E + S had preserved the increase in CSA and strength from the preparatory period. From the beginning of the preparatory period to 13 weeks into the competition period, E + S increased peak power output in the Wingate test, power output at 2 mmol l⁻¹ [la⁻], maximal aerobic power output (W _max), and mean power output in the 40-min all-out trial (p < 0.05). The relative improvements in the last two measurements were larger than in E (p < 0.05). For E, W _max and power output at 2 mmol l⁻¹ [la⁻] remained unchanged. In conclusion, in well-trained cyclists, strength maintenance training in a competition period preserved increases in thigh muscle CSA and leg strength attained in a preceding preparatory period and further improved cycling performance determinants and performance.     

Effect of heavy strength training on thigh muscle cross-sectional area, performance determinants, and performance in well-trained cyclists

The purpose of this study was to investigate the effect of heavy strength training on thigh muscle cross-sectional area (CSA), determinants of cycling performance, and cycling performance in well-trained cyclists. Twenty well-trained cyclists were assigned to either usual endurance training combined with heavy strength training [E + S; n = 11 (♂ = 11)] or to usual endurance training only [E; n = 9 (♂ = 7, ♀ = 2)]. The strength training performed by E + S consisted of four lower body exercises [3 × 4–10 repetition maximum (RM)], which were performed twice a week for 12 weeks. Thigh muscle CSA, maximal force in isometric half squat, power output in 30 s Wingate test, maximal oxygen consumption (VO_2max), power output at 2 mmol l⁻¹ blood lactate concentration ([la⁻]), and performance, as mean power production, in a 40-min all-out trial were measured before and after the intervention. E + S increased thigh muscle CSA, maximal isometric force, and peak power in the Wingate test more than E. Power output at 2 mmol l⁻¹ [la⁻] and mean power output in the 40-min all-out trial were improved in E + S (P < 0.05). For E, only performance in the 40-min all-out trial tended to improve (P = 0.057). The two groups showed similar increases in VO_2max (P < 0.05). In conclusion, adding strength training to usual endurance training improved determinants of cycling performance as well as performance in well-trained cyclists. Of particular note is that the added strength training increased thigh muscle CSA without causing an increase in body mass.     

Nucleocytoplasmic transport signals affect the age at onset of abnormalities in knock-in mice expressing polyglutamine within an ectopic protein context

In order to better understand the role of nuclear localization of polyglutamine in the human CAG repeat disorders, gene targeting was used to add either nuclear localization (NLS) or nuclear export (NES) signals to versions of the mouse Hprt protein containing expanded polyglutamine (HprtQ150). The NLS increased levels of nuclear HprtQ150 protein in the mouse brain and hastened both the presentation of neuronal intranuclear inclusions (NIIs) and the onset of behavioral abnormalities. The NES reduced levels of nuclear HprtQ150 protein in mouse brain and delayed both the presentation of NIIs and the onset of behavioral abnormalities. Together these results indicate the nucleus is the primary site of toxicity in HprtQ150 mice. Furthermore, the signals did not alter the relative regional distribution of NIIs, suggesting that factors other than nuclear access dictate the regional specificity of NII formation in this mouse model.     

In vitro interactions between aged garlic extract and drugs used for the treatment of cardiovascular and diabetic patients

Background  

Disease preventing effects gained by garlic consumption have been recognized since early period of history, making commercially available garlic supplements attractive to the general public. Possible pharmacokinetic interactions which could occur between applied drugs and aged garlic extract (AGE) are unknown.     

Regionally selective atrophy of subcortical structures in prodromal HD as revealed by statistical shape analysis

Huntington disease (HD) is a neurodegenerative disorder that involves preferential atrophy in the striatal complex and related subcortical nuclei. In this article, which is based on a dataset extracted from the PREDICT‐HD study, we use statistical shape analysis with deformation markers obtained through “Large Deformation Diffeomorphic Metric Mapping” of cortical surfaces to highlight specific atrophy patterns in the caudate, putamen, and globus pallidus, at different prodromal stages of the disease. On the basis of the relation to cortico‐basal ganglia circuitry, we propose that statistical shape analysis, along with other structural and functional imaging studies, may help expand our understanding of the brain circuitry affected and other aspects of the neurobiology of HD, and also guide the most effective strategies for intervention. Hum Brain Mapp 35:792–809, 2014. © 2012 Wiley Periodicals, Inc.     

Mechanism of interaction between cisplatin and human recombinant interferon gamma in human ovarian-cancer cell lines

Human ovarian carcinoma cells (2008 and its cisplatin-resistant sub-line 2008/C13*) were sensitized to cisplatin by treatment with human recombinant gamma interferon (IFNγ). IFNγ produced no significant change in the uptake of CDDP. Exposure of 2008 and 2008/C13* cells to IFNγ resulted in a time-dependent decrease of cellular glutathione and total glutathione-S-transferase activity, principally the π isoform. By contrast, the treatment of 2008 and 2008/C13* cell lines with IFNγ induced rather than suppressed metallothionein II_A mRNA levels. IFNγ changed neither the formation of total platinum-DNA adducts, nor DNA repair. A significant decrease in c-erbB-2 expression was observed both in sensitive and in resistant cell lines after treatment with IFNγ, and this decrease was dose-dependent. Our results indicate that the mechanism of IFNγ-induced sensitization in human ovarian-cancer cell lines is multifactorial. © 1995 Wiley-Liss, Inc.     

Histiocytosis-X in gynecology

(1) Histiocytosis-X can manifest itself in virtually every organ, but in gynecology it is an absolute curiosity. (2) Differential diagnosis must exclude specific and nonspecific ulcerations and granulations such as syphilis, tuberculosis, Boeck's disease, and also neoplastic processes like lymphomas, sarcomas, carcinomas, and malignant diseases of the hemopoietic system. (3) The diagnosis by light microscopy alone, as in our case, may be insufficient; therefore, electron microscopy should be used. As soon as the diagnosis is confirmed histologically, an extensive examination of all organs is necessary in order to establish an exact prognosis and an optimal plan of therapy. (4) Because of the unknown etiology of histiocytosis-X, a causal treatment is not yet possible. In spite of this, with a symptomatic, individualized therapy by means of excision, low-dose irradiation and cytotoxic agents a 5-year survival of 90% was obtained for the patients. (5) Because of its rarity and multidisciplinary character, histiocytosis-X is a challenge to interdisciplinary and interregional cooperation. Though not being a malignoma in the strict sense, diagnosis, therapy, and in part prognosis are not essentially different from a malignant disease.     

Treatment effects on nigrostriatal projection integrity in partial 6-OHDA lesions: comparison of L-DOPA and pramipexole

There is controversy over potential effects of dopaminergic replacement therapies on the partially lesioned nigrostriatal dopaminergic projection. We evaluated indirect (levodopa, L-DOPA) versus direct (pramipexole, PRA) dopaminergic treatment effects on nigrostriatal lesion severity as measured with vesicular monoamine transporter type-2 (VMAT2) binding. Prior studies have shown that striatal VMAT2 density provides an objective estimate of dopaminergic neuronal integrity, without confounding effects of compensatory regulation. Partial unilateral median forebrain bundle lesions were made by injection of 6-hydroxydopamine in adult male Sprague-Dawley rats. Lesion severity was estimated using rotational behavior after injections of apomorphine and amphetamine. Rats were ranked and matched in pairs by rotation and assigned to receive either PRA (1 mg/kg/day) or L-DOPA/benserazide (100/25 mg/kg/day) ip via osmotic pump. After 4 weeks of drug treatment, in vitro autoradiography was performed with [(3)H]methoxytetrabenazine to measure striatal VMAT2 binding density. Lesion-to-intact VMAT2 density correlated with rotation in both treatment groups. There was no treatment effect on VMAT2 expression in the partially lesioned striatum and thus no differential effect of indirect versus direct dopamimetic treatment on nigrostriatal integrity.