Positive selection drives population differentiation in the skeletal genes in modern humans

During the course of evolution, the human skeletal system has evolved rapidly leading to an incredible array of phenotypic diversity, including variations in height and bone mineral density. However, the genetic basis of this phenotypic diversity and the relatively rapid tempo of evolution have remained largely undocumented. Here, we discover that skeletal genes exhibit a significantly greater level of population differentiation among humans compared with other genes in the genome. The pattern is exceptionally evident at amino acid-altering sites within these genes. Divergence is greater between Africans and both Europeans and East Asians. In contrast, relatively weak differentiation is observed between Europeans and East Asians. SNPs with higher levels of differentiation have correspondingly higher derived allele frequencies in Europeans and East Asians. Thus, it appears that positive selection has operated on skeletal genes in the non-African populations and this may have been initiated with the human colonization of Eurasia. In conclusion, we provide genetic evidence supporting the rapid evolution of the human skeletal system and the associated diversity of phenotypes.

Evidence for Positive Selection on the Osteogenin (BMP3) Gene in Human Populations

Background: Human skeletal system has evolved rapidly since the dispersal of modern humans from Africa, potentially driven by selection and adaptation. Osteogenin (BMP3) plays an important role in skeletal development and bone osteogenesis as an antagonist of the osteogenic bone morphogenetic proteins, and negatively regulates bone mineral density. Methodology/Principal Findings: Here, we resequenced the BMP3 gene from individuals in four geographically separated modern human populations. Features supportive of positive selection in the BMP3 gene were found including the presence of an excess of nonsynonymous mutations in modern humans, and a significantly lower genetic diversity that deviates from neutrality. The prevalent haplotypes of the first exon region in Europeans demonstrated features of long-range haplotype homogeneity. In contrast with findings in European, the derived allele SNP Arg192Gln shows higher extended haplotype homozygosity in East Asian. The worldwide allele frequency distribution of SNP shows not only a high-derived allele frequency in Asians, but also in Americans, which is suggestive of functional adaptation. Conclusions/Significance: In conclusion, we provide evidence for recent positive selection operating upon a crucial gene in skeletal development, which may provide new insight into the evolution of the skeletal system and bone development.

Prevalent positive epistasis in Escherichia coli and Saccharomyces cerevisiae metabolic networks

Epistasis refers to the interaction between genes. Although high-throughput epistasis data from model organisms are being generated and used to construct genetic networks(1-3), the extent to which genetic epistasis reflects biologically meaningful interactions remains unclear(4-6). We have addressed this question through in silico mapping of positive and negative epistatic interactions amongst biochemical reactions within the metabolic networks of Escherichia coli and Saccharomyces cerevisiae using flux balance analysis. We found that negative epistasis occurs mainly between nonessential reactions with overlapping functions, whereas positive epistasis usually involves essential reactions, is highly abundant and, unexpectedly, often occurs between reactions without overlapping functions. We offer mechanistic explanations of these findings and experimentally validate them for 61 S. cerevisiae gene pairs.

Transplantable neural progenitor populations derived from Rhesus monkey embryonic stem cells

Cell-based therapies using embryonic stem cells (ESCs) in the treatment of neural disease will require the generation of homogenous donor neural progenitor (NP) populations. Here we describe an efficient culture system containing hepatocyte growth factor (HGF) and G5 supplement for the production of highly enriched (88.3% +/- 8.1%)populations of NPs from rhesus monkey ESCs. Additional purification resulted in NP preparations that were 98% nestin positive. Moreover, NPs, as monolayers or neurospheres, could be maintained for prolonged periods of time in media containing HGF+G5 or G5 alone. In vitro differentiation and in vivo transplantation assays showed that NPs could differentiate into neurons, astrocytes, and oligodendrocytes. The kinds and quantities of differentiated cells derived from NPs were closely correlated with their niches in vivo. Glial differentiation was predominant in periventricular areas, whereas cells migrating into the cortex were mostly neurons. Cell counts showed that 2 months after transplantation, approximately 25% of transplanted NPs survived and 65% - 80% of the surviving transplanted cells migrated along the ventricular wall or in a radial fashion. Subcloning demonstrated that several clonal lines derived from NPs expressed nestin and differentiated into three neural lineages in vitro and in rat brains in vivo. In contrast, some subcloned lines showed restricted differentiation both in vitro and in vivo in rat brains. These observations set the stage for obtaining highly enriched NPs and evaluating the efficacy of NP-based transplantation therapy in the nonhuman primate and will provide a platform for probing the molecular mechanisms that control neural induction.

A microelectrode drive for long term recording of neurons in freely moving and chaired monkeys

An electrode drive is described for recordings of neurons in freely moving and chaired monkeys during the performance of behavioural tasks. The electrode drives are implanted for periods of up to 6 months, and can advance up to 42 electrodes using 14 independent drive mechanisms. The drive samples 288 points within a 12 mm x 12 min region, with 15 min of electrode travel. Major advantages are that recordings are made in freely moving monkeys, and these recordings can be compared with those in chaired experiments; waveforms of single neurons are stable, enabling prolonged recordings of the same neurons across periods of days; recordings can be made throughout the brain, including the dorsolateral prefrontal cortex and hippocampus; the drive accommodates both sharp microelectrodes and fine wire assemblies such as tetrodes. (C) 2003 Elsevier Science B.V; All rights reserved.

Reinforcement-related neurons in the primate basal forebrain respond to the learned significance of task events rather than to the hedonic attributes of reward

The objective of this study was to determine if the responses of basal forebrain neurons are related to the cognitive processes necessary for the performance of behavioural tasks, or to the hedonic attributes of the reinforcers delivered to the monkey as

Mechanisms of H+ modulation of glycinergic response in rat sacral dorsal commissural neurons

Many ionotropic receptors are modulated by extracellular H+. So far, few studies have directly addressed the role of such modulation at synapses. In the present study, we investigated the effects of changes in extracellular pH on glycinergic miniature inhibitory postsynaptic currents (mIPSCs) as well as glycine-evoked currents (I-Gly) in mechanically dissociated spinal neurons with native synaptic boutons preserved. H+ modulated both the mIPSCs and I-Gly, biphasically, although it activated an amiloride-sensitive inward current by itself. Decreasing extracellular pH reversibly inhibited the amplitude of the mIPSCs and I-Gly, while increasing external pH reversibly potentiated these parameters. Blockade of acid-sensing ion channels (ASICs) with amiloride, the selective antagonist of ASICs, or decreasing intracellular pH did not alter the modulatory effect of H+ on either mIPSCs or I-Gly, H+ shifted the EC50 of the glycine concentration-response curve from 49.3 +/- 5.7 muM at external pH 7.4 to 131.5 +/- 8.1 muM at pH 5.5, without altering the Cl- selectivity of the glycine receptor (GlyR), the Hill coefficient and the maximal I-Gly, suggesting a competitive inhibition of I-Gly by H+. Both Zn2+ and H+ inhibited I-Gly. However, H+ induced no further inhibition of I-Gly in the presence of a saturating concentration of Zn2+. In addition, H+ significantly affected the kinetics of glycinergic mIPSCs and I-Gly. It is proposed that H+ and/or Zn2+ compete with glycine binding and inhibit the amplitude of glycinergic mIPSCs and I-Gly. Moreover, binding of H+ induces a global conformational change in GlyR, which closes the GlyR Cl- channel and results in the acceleration of the seeming desensitization of IGly as well as speeding up the decay time constant of glycinergic mIPSCs. However, the deprotonation rate is faster than the unbinding rate of glycine from the GlyR, leading to reactivation of the undesensitized GlyR after washout of agonist and the appearance of a rebound I-Gly. H+ also modulated the glycine cotransmitter, GABA-activated current (I-GABA). Taken together, the results support a 'conformational coupling' model for H+ modulation of the GlyR and suggest that W may act as a novel modulator for inhibitory neurotransmission in the mammalian spinal cord.

Characterization of acid-sensing ion channels in dorsal horn neurons of rat spinal cord

Acid-sensing ion channels (ASICs) are ligand-gated cation channels activated by extracellular protons. In periphery, they contribute to sensory transmission, including that of nociception and pain. Here we characterized ASIC-like currents in dorsal horn neurons of the rat spinal cord and their functional modulation in pathological conditions. Reverse transcriptase-nested PCR and Western blotting showed that three ASIC isoforms, ASIC1a, ASIC2a, and ASIC2b, are expressed at a high level in dorsal horn neurons. Electrophysiological and pharmacological properties of the proton-gated currents suggest that homomeric ASIC1a and/or heteromeric ASIC1a + 2b channels are responsible for the proton-induced currents in the majority of dorsal horn neurons. Acidification-induced action potentials in these neurons were compatible in a pH-dependent manner with the pH dependence of ASIC-like current. Furthermore, peripheral complete Freund's adjuvant-induced inflammation resulted in increased expression of both ASIC1a and ASIC2a in dorsal horn. These results support the idea that the ASICs of dorsal horn neurons participate in central sensory transmission/modulation under physiological conditions and may play important roles in inflammation-related persistent pain.

Properties of the proton-evoked currents and their modulation by Ca2+ and Zn2+ in the acutely dissociated hippocampus CA1 neurons

The characterization of acid-sensing ion channel (ASIC)-like currents has been reported in hippocampal neurons in primary culture. However, it is suggested that the profile of expression of ASICs changes in culture. In this study, we investigated the properties of proton-activated current and its modulation by extracellular Ca2+ and Zn2+ in neurons acutely dissociated from the rat hippocampal CA1 using conventional whole-cell patch-clamp recording. A rapidly decaying inward current and membrane depolarization was induced by exogenous application of acidic solution. The current was sensitive to the extracellular proton with a response threshold of pH 7.0-6.8 and the pH(50) Of 6.1, the reversal potential close to the Na+ equilibrium potential. It had a characteristic of acid-sensing ion channels (ASICs) as demonstrated by its sensitivity to amiloride (IC50 = 19.6 +/- 2.1 muM). Either low [Ca2+](0) or high [Zn2+](0) increased the amplitude of the current. All these characteristics are consistent with a current mediated through a mixture of homomeric ASIC1a and heteromeric ASIC1a + 2a channels and closely replicate many of the characteristics that have been previously reported for hippocampal neurons cultured for a week or more, indicating that culture artifacts do not necessarily flaw the properties of ASICs. Interestingly, we found that high [Zn2+] (>10(-4) M) slowed the decay time constant of the ASIC-like current significantly in both acutely dissociated and cultured hippocampal neurons. In addition, the facilitating effects of low [Ca2+](0) and high [Zn2+](0) on the ASIC-like current were not additive. Since tissue acidosis, extracellular Zn elevation and/or Ca2+ reduction occur concurrently under some physiological and/or pathological conditions, the present observations suggest that hippocampal ASICs may offer a novel pharmacological target for therapeutic invention. (C) 2004 Elsevier B.V. All rights reserved.

Upregulation of acid-sensing ion channel ASIC1a in spinal dorsal horn neurons contributes to inflammatory pain hypersensitivity

Development of chronic pain involves alterations in peripheral nociceptors as well as elevated neuronal activity in multiple regions of the CNS. Previous pharmacological and behavioral studies suggest that peripheral acid-sensing ion channels (ASICs) cont

Statistical analysis of sequence features of introns with positive transcriptional regulation in yeast genes

A great deal of experimental studies have shown that many introns of eukaryotic genes function as regulators of transcription. However, comprehensive studies of this problem have not yet been conducted. After checking the transcription frequencies of some Saccharomyces cerevisiae (yeast), genes and their introns, a remarkable phenomenon was discovered that generally the introns of the genes with higher transcription frequencies are longer, and the introns of the genes with lower transcription frequencies are shorter. This suggests that the longer introns of genes with higher transcription frequencies may contain some characteristic sequence structures, which could enhance the transcription of genes. Therefore, two sets of introns of yeast genes were chosen for further study. The transcription frequencies of the first set of genes are higher (>30), and those of the second set of genes are lower (less than or equal to10). Some oligonucleotides are detected by statistically comparative analyses of the occurrence frequencies of oligonucleotides (mainly tetranucleotides and pentanucleotides), whose occurrence frequencies in the first set of introns; are significantly higher than those in the second set of introns, and are also significantly higher than those in the exons flanking the introns of the first set. Some of these extracted oligonucleotides are the same as the regulatory elements of transcription revealed by experimental analyses. Besides, the distributions of these extracted oligonucleotides in the two sets of introns and the exons show that the sequence structures of the first set of introns are favorable for transcription of genes.

Detection of potential positive regulatory motifs of transcription in yeast introns by comparative analysis of oligonucleotide frequencies

We conducted a comparative statistical analysis of tetra- through hexanucleotide frequencies in two sets of introns of yeast genes. The first set consisted of introns of genes that have transcription rates higher than 30 mRNAs/h while the second set contained introns of genes whose transcription rates were lower than or equal to 10 mRNAs/h. Some oligonucleotides whose occurrence frequencies in the first set of introns are significantly higher than those in the second set of introns were detected. The frequencies of occurrence of most of these detected oligonucleotides are also significantly higher than those in the exons flanking the introns of the first set. Interestingly some of these detected oligonucleotides are the same as well known "signature" sequences of transcriptional regulatory elements. This could imply the existence of potential positive regulatory motifs of transcription in yeast introns. (C) 2003 Elsevier Ltd. All rights reserved.

Developmental expression of CagMdkb during gibel carp embryogenesis

Midkine (Mdk) genes have been revealed to have different expression patterns in vertebrates and therefore, additional studies on Mdk expression patterns are required in more species. In this study, CagMdkb has been cloned and characterized from a SMART cDNA library of 10-somite stage embryos of Carassius auratus gibelio. Its full length cDNA is 1091 bp and encodes a sequence of 147 amino acids, which shows 97.3% identity to zebrafish Mdkb on the amino acid level. RT-PCR analysis reveals that CagMdkb is first transcribed in gastrula embryos and maintains a relatively stable expression level during subsequent embryogenesis. Western blot analysis reveals a 19 kDa maternal CagMdkb protein band and the zygotic CagMdkb protein is expressed from gastrula stage. At around 10 somite stage, the 19 kDa CagMdkb is processed to another protein band of about 17 kDa, which might be the secreted form with the 21-residue signal peptide removed. With immunofluorescence analysis, maternal CagMdkb protein was found to be localized in each blastamere cell of early embryos. The zygotic CagMdkb positive fluorescence signal was detected from a pair of large neurons at 18-somite stage. At the later stages, CagMdkb protein was also extended to numerous small neurons in the forebrain, midbrain and hindbrain, as well as to nerve fibers in the spinal cord. Co-localization with 3A10 antibody revealed CagMdkb immunoreactivity on developing Mauthner neurons, a member of reticulospinal neurons. In addition, ectopic expression of CagMdkb in early embryos of gibel carp and zebrafish suppressed head formation and CagMdkb function was found to depend on secretory activity. All these findings indicate that CagMdkb plays an important role in neural development during gibel carp embryogenesis and there is functional conservation of Mdkb in fish head formation.

Positive effects of continuous low nitrate levels on growth and photosynthesis of Alexandrium tamarense (Gonyaulacales, Dinophyceae)

The growth and photosynthesis of Alexandrium tamarense (Lebour) Balech in different nutrient conditions were investigated. Low nitrate level (0.0882 mmol/L) resulted in the highest average growth rate from day 0 to day 10 (4.58 x 10(2) cells mL(-1) d(-1)), but the lowest cell yield (5420 cells mL(-1)) in three nitrate level cultures. High nitrate-grown cells showed lower levels of chlorophyll a-specific and cell-specific light-saturated photosynthetic rate (P-m(chl a) and P-m(cell)), dark respiration rate (R-d(chl a) and R-d(cell)) and chlorophyll a-specific apparent photosynthetic efficiency (alpha(chl a)) than was seen for low nitrate-grown cells; whereas the cells became light saturated at higher irradiance at low nitrate condition. When cultures at low nitrate were supplemented with nitrate at 0.7938 mmol/L in late exponential growth phase, or with nitrate at 0.7938 mmol/L and phosphate at 0.072 mmol/L in stationary growth phase, the cell yield was drastically enhanced, a 7-9 times increase compared with non-supplemented control culture, achieving 43 540 cells mL(-1) and 52 300 cells mL(-1), respectively; however, supplementation with nitrate in the stationary growth phase or with nitrate and phosphate in the late exponential growth phase increased the cell yield by no more than 2 times. The results suggested that continuous low level of nitrate with sufficient supply of phosphate may facilitate the growth of A. tamarense.

Positive selection on multiple antique allelic lineages of transferrin in the polyploid Carassius auratus

Transferrin polymorphism has been studied in the polyploid Carassius auratus by cloning and sequence analysis of cDNAs from its three subspecies C. auratus gibelio, C. auratus auratus, and C. auratus cuvieri. DNA polymorphism of extremely high extent was shown for the transferrin gene by the 248 segregation sites among coding region sequences of its alleles. The deduced amino acid sequences of the transferrin alleles showed variable theoretical physicochemical parameters, which might constitute molecular basis for their electrophoretic heterogeneity. Positive selection was inferred by the replacement/synonymous ratios larger than 1 in partial allelic lineages which was subsequently confirmed by likelihood simulation under neutral or selection models. Furthermore, the correspondent sites to these selected codons were collectively located at two planes in the crystallographic structure of rabbit transferrin, which suggested that the rapid evolution of C. auratus transferrin might correlate to its adaptation to variable environmental elements such as oxygen pressure. The minimal 26 recombination events were detected among coding sequences of C. auratus transferrin, with partial mosaic sequences and breakpoints identified by identity scanning and information site analyses. Phylogenetic analyses revealed multiple antique allelic lineages of transferrin, which was estimated to diverge fifteen to twenty MYA. All these features strongly suggested the role of balancing selection in long persistence of high transferrin polymorphism in C. auratus. Furthermore, owing to its particular evolutionary backgrounds, the silver crucian carp might possess a distinctive balancing selection mechanism.