Human ribosomal protein S3a: cloning of the cDNA and primary structure of the protein.

The amino acid (aa) sequence of human ribosomal protein S3a (hRPS3a) was deduced partially from the nucleotide sequence of the corresponding cDNA and confirmed by direct aa sequencing from the N terminus of the purified hRPS3a protein. The cDNA clone was isolated from a cDNA expression library in the pEX vector using antibodies. The hRPS3a protein has 263 aa and its calculated M(r) is 29 813.     

The properties of the complex between ribosomal protein L2 and tRNA

Escherichia coli ribosomal protein L2 interacts with fMet-tRNA^Met and NacPhe-tRNA^Phe in solution, protecting their 3'-ends from enzymatic degradation. At the same time L2 enhances the rate of spontaneous hydrolysis of the ester bonds between terminal riboses and amino acyl moieties of these two peptidyl-tRNA analogues. L2 has, however, only a slight effect on the rate of spontaneous deacylation of aminoacyltRNAs. We suggest that the role of L2 is in the fixation of the aminoacyl stem of tRNA to the ribosome at its P-site, and speculate that this protein is directly involved in the peptidyl transferase (PT) reaction. Peptidyl transferase Protein L2 tRNA-protein complex     

The Light Skin Allele of SLC24A5 in South Asians and Europeans Shares Identity by Descent

Skin pigmentation is one of the most variable phenotypic traits in humans. A non-synonymous substitution (rs1426654) in the third exon of SLC24A5 accounts for lighter skin in Europeans but not in East Asians. A previous genome-wide association study carried out in a heterogeneous sample of UK immigrants of South Asian descent suggested that this gene also contributes significantly to skin pigmentation variation among South Asians. In the present study, we have quantitatively assessed skin pigmentation for a largely homogeneous cohort of 1228 individuals from the Southern region of the Indian subcontinent. Our data confirm significant association of rs1426654 SNP with skin pigmentation, explaining about 27% of total phenotypic variation in the cohort studied. Our extensive survey of the polymorphism in 1573 individuals from 54 ethnic populations across the Indian subcontinent reveals wide presence of the derived-A allele, although the frequencies vary substantially among populations. We also show that the geospatial pattern of this allele is complex, but most importantly, reflects strong influence of language, geography and demographic history of the populations. Sequencing 11.74 kb of SLC24A5 in 95 individuals worldwide reveals that the rs1426654-A alleles in South Asian and West Eurasian populations are monophyletic and occur on the background of a common haplotype that is characterized by low genetic diversity. We date the coalescence of the light skin associated allele at 22–28 KYA. Both our sequence and genome-wide genotype data confirm that this gene has been a target for positive selection among Europeans. However, the latter also shows additional evidence of selection in populations of the Middle East, Central Asia, Pakistan and North India but not in South India.     

Uniparental Genetic Heritage of Belarusians: Encounter of Rare Middle Eastern Matrilineages with a Central European Mitochondrial DNA Pool

Ethnic Belarusians make up more than 80% of the nine and half million people inhabiting the Republic of Belarus. Belarusians together with Ukrainians and Russians represent the East Slavic linguistic group, largest both in numbers and territory, inhabiting East Europe alongside Baltic-, Finno-Permic- and Turkic-speaking people. Till date, only a limited number of low resolution genetic studies have been performed on this population. Therefore, with the phylogeographic analysis of 565 Y-chromosomes and 267 mitochondrial DNAs from six well covered geographic sub-regions of Belarus we strove to complement the existing genetic profile of eastern Europeans. Our results reveal that around 80% of the paternal Belarusian gene pool is composed of R1a, I2a and N1c Y-chromosome haplogroups – a profile which is very similar to the two other eastern European populations – Ukrainians and Russians. The maternal Belarusian gene pool encompasses a full range of West Eurasian haplogroups and agrees well with the genetic structure of central-east European populations. Our data attest that latitudinal gradients characterize the variation of the uniparentally transmitted gene pools of modern Belarusians. In particular, the Y-chromosome reflects movements of people in central-east Europe, starting probably as early as the beginning of the Holocene. Furthermore, the matrilineal legacy of Belarusians retains two rare mitochondrial DNA haplogroups, N1a3 and N3, whose phylogeographies were explored in detail after de novo sequencing of 20 and 13 complete mitogenomes, respectively, from all over Eurasia. Our phylogeographic analyses reveal that two mitochondrial DNA lineages, N3 and N1a3, both of Middle Eastern origin, might mark distinct events of matrilineal gene flow to Europe: during the mid-Holocene period and around the Pleistocene-Holocene transition, respectively.     

In-solution hybrid capture of bisulfite-converted DNA for targeted bisulfite sequencing of 174 ADME genes

DNA methylation is one of the most important epigenetic alterations involved in the control of gene expression. Bisulfite sequencing of genomic DNA is currently the only method to study DNA methylation patterns at single-nucleotide resolution. Hence, next-generation sequencing of bisulfite-converted DNA is the method of choice to investigate DNA methylation profiles at the genome-wide scale. Nevertheless, whole genome sequencing for analysis of human methylomes is expensive, and a method for targeted gene analysis would provide a good alternative in many cases where the primary interest is restricted to a set of genes.Here, we report the successful use of a custom Agilent SureSelect Target Enrichment system for the hybrid capture of bisulfite-converted DNA. We prepared bisulfite-converted next-generation sequencing libraries, which are enriched for the coding and regulatory regions of 174 ADME genes (i.e. genes involved in the metabolism and distribution of drugs). Sequencing of these libraries on Illumina’s HiSeq2000 revealed that the method allows a reliable quantification of methylation levels of CpG sites in the selected genes, and validation of the method using pyrosequencing and the Illumina 450K methylation BeadChips revealed good concordance.     

Biomarker Profiling by Nuclear Magnetic Resonance Spectroscopy for the Prediction of All-Cause Mortality: An Observational Study of 17,345 Persons

Background

Early identification of ambulatory persons at high short-term risk of death could benefit targeted prevention. To identify biomarkers for all-cause mortality and enhance risk prediction, we conducted high-throughput profiling of blood specimens in two large population-based cohorts.

Methods and Findings

106 candidate biomarkers were quantified by nuclear magnetic resonance spectroscopy of non-fasting plasma samples from a random subset of the Estonian Biobank (n = 9,842; age range 18–103 y; 508 deaths during a median of 5.4 y of follow-up). Biomarkers for all-cause mortality were examined using stepwise proportional hazards models. Significant biomarkers were validated and incremental predictive utility assessed in a population-based cohort from Finland (n = 7,503; 176 deaths during 5 y of follow-up). Four circulating biomarkers predicted the risk of all-cause mortality among participants from the Estonian Biobank after adjusting for conventional risk factors: alpha-1-acid glycoprotein (hazard ratio [HR] 1.67 per 1–standard deviation increment, 95% CI 1.53–1.82, p = 5×10⁻³¹), albumin (HR 0.70, 95% CI 0.65–0.76, p = 2×10⁻¹⁸), very-low-density lipoprotein particle size (HR 0.69, 95% CI 0.62–0.77, p = 3×10⁻¹²), and citrate (HR 1.33, 95% CI 1.21–1.45, p = 5×10⁻¹⁰). All four biomarkers were predictive of cardiovascular mortality, as well as death from cancer and other nonvascular diseases. One in five participants in the Estonian Biobank cohort with a biomarker summary score within the highest percentile died during the first year of follow-up, indicating prominent systemic reflections of frailty. The biomarker associations all replicated in the Finnish validation cohort. Including the four biomarkers in a risk prediction score improved risk assessment for 5-y mortality (increase in C-statistics 0.031, p = 0.01; continuous reclassification improvement 26.3%, p = 0.001).

Conclusions

Biomarker associations with cardiovascular, nonvascular, and cancer mortality suggest novel systemic connectivities across seemingly disparate morbidities. The biomarker profiling improved prediction of the short-term risk of death from all causes above established risk factors. Further investigations are needed to clarify the biological mechanisms and the utility of these biomarkers for guiding screening and prevention. Please see later in the article for the Editors'' Summary     

Genome-Wide Analysis of Cold Adaptation in Indigenous Siberian Populations

Following the dispersal out of Africa, where hominins evolved in warm environments for millions of years, our species has colonised different climate zones of the world, including high latitudes and cold environments. The extent to which human habitation in (sub-)Arctic regions has been enabled by cultural buffering, short-term acclimatization and genetic adaptations is not clearly understood. Present day indigenous populations of Siberia show a number of phenotypic features, such as increased basal metabolic rate, low serum lipid levels and increased blood pressure that have been attributed to adaptation to the extreme cold climate. In this study we introduce a dataset of 200 individuals from ten indigenous Siberian populations that were genotyped for 730,525 SNPs across the genome to identify genes and non-coding regions that have undergone unusually rapid allele frequency and long-range haplotype homozygosity change in the recent past. At least three distinct population clusters could be identified among the Siberians, each of which showed a number of unique signals of selection. A region on chromosome 11 (chr11:66–69 Mb) contained the largest amount of clustering of significant signals and also the strongest signals in all the different selection tests performed. We present a list of candidate cold adaption genes that showed significant signals of positive selection with our strongest signals associated with genes involved in energy regulation and metabolism (CPT1A, LRP5, THADA) and vascular smooth muscle contraction (PRKG1). By employing a new method that paints phased chromosome chunks by their ancestry we distinguish local Siberian-specific long-range haplotype signals from those introduced by admixture.     

Neuroprotective and anticonvulsant effects of organic and conventional purple grape juices on seizures in Wistar rats induced by pentylenetetrazole

Epilepsy is the most common neurological disorder worldwide. Studies have shown that recurrent seizures may increase the concentration of reactive oxygen species, which can lead to oxidative stress and neuronal damage. These seizures result in substantial deleterious effects on an individual's health. Organic and conventional grape juices are rich in polyphenols, compounds with important antioxidant activity. However, these juices could have differences in their polyphenol content. The aim of this study was to investigate the neuroprotective and anticonvulsant effects of organic and conventional grape juice treatments in Wistar rats against pentylenetetrazole (a convulsant drug)-induced damage. In addition, we evaluated potential behavioral changes in rats treated with the juices and the polyphenolic profile of those samples. Animals (n=16 in each group) received treatment with saline, organic or conventional grape juice for 17 days. On the eighteenth day, behavioral changes were evaluated by an open field test. Afterwards, half of the rats from each group received pentylenetetrazole and were observed for 30 min to evaluate possible seizure characteristics. The animals were subsequently killed by decapitation and their hippocampus, cerebellum and cerebral cortex tissues were isolated. The results of this study showed that neither organic nor conventional grape juice altered the behavior parameters, and no statistical differences were observed in the seizure characteristics of the groups. Nevertheless, both juice types were able to protect from lipid and protein oxidative damage, decrease nitric oxide content and increase enzymatic (superoxide dismutase and catalase) and non-enzymatic (sulfhydryl protein) antioxidant defenses in brain tissues following pentylenetetrazole-induced seizures. In general, organic juice showed superior results in each test, probably due to its higher polyphenol content relative to conventional juice. These results indicate that grape juices can provide further insight into natural neuroprotective compounds and may lead to the development of new therapeutic strategies for epileptic patients.     

Acute Kidney Injury in Severe Sepsis and Septic Shock in Patients with and without Diabetes Mellitus: A Multicenter Study

IntroductionWhether diabetes mellitus increases the risk of acute kidney injury (AKI) during sepsis is controversial.ResultsFirst, we compared 451 patients with severe sepsis or septic shock and diabetes to 3,277 controls with severe sepsis or septic shock and without diabetes. Then, we compared 318 cases (with diabetes) to 746 matched controls (without diabetes). Diabetic patients did not have a higher frequency of AKI (hazard ratio [HR], 1.18; P = 0.05]) or RRT (HR, 1.09; P = 0.6). However, at discharge, diabetic patients with severe sepsis or septic shock who experienced acute kidney injury during the ICU stay and were discharged alive more often required RRT (9.5% vs. 4.8%; P = 0.02), had higher serum creatinine values (134 vs. 103 µmoL/L; P<0.001) and had less often recovered a creatinine level less than 1.25 fold the basal creatinine (41.1% vs. 60.5%; P<0.001).ConclusionsIn patients with severe sepsis or septic shock, diabetes is not associated with occurrence of AKI or need for RRT but is an independent risk factor for persistent renal dysfunction in patients who experience AKI during their ICU stay.     

Sequence variation in nuclear ribosomal small subunit,internal transcribed spacer and large subunit regions of Rhizophagus irregularis and Gigaspora margarita is high and isolate‐dependent

Arbuscular mycorrhizal (AM) fungi are known to exhibit high intra‐organism genetic variation. However, information about intra‐ vs. interspecific variation among the genes commonly used in diversity surveys is limited. Here, the nuclear small subunit (SSU) rRNA gene, internal transcribed spacer (ITS) region and large subunit (LSU) rRNA gene portions were sequenced from 3 to 5 individual spores from each of two isolates of Rhizophagus irregularis and Gigaspora margarita. A total of 1482 Sanger sequences (0.5 Mb) from 239 clones were obtained, spanning ~4370 bp of the ribosomal operon when concatenated. Intrasporal and intra‐isolate sequence variation was high for all three regions even though variant numbers were not exhausted by sequencing 12–40 clones per isolate. Intra‐isolate nucleotide variation levels followed the expected order of ITS > LSU > SSU, but the values were strongly dependent on isolate identity. Single nucleotide polymorphism (SNP) densities over 4 SNP/kb in the ribosomal operon were detected in all four isolates. Automated operational taxonomic unit picking within the sequence set of known identity overestimated species richness with almost all cut‐off levels, markers and isolates. Average intraspecific sequence similarity values were 99%, 96% and 94% for amplicons in SSU, LSU and ITS, respectively. The suitability of the central part of the SSU as a marker for AM fungal community surveys was further supported by its level of nucleotide variation, which is similar to that of the ITS region; its alignability across the entire phylum; its appropriate length for next‐generation sequencing; and its ease of amplification in single‐step PCR.