Pattern and Timing of Gene Duplication in Animal Genomes

Duplication of genes, giving rise to multigene families, has been a characteristic feature of the evolution of eukaryotic genomes. In the case of vertebrates, it has been proposed that an increase in gene number resulted from two rounds of duplication of the entire genome by polyploidization (the 2R hypothesis). In the most extensive test to date of this hypothesis, we compared gene numbers in homologous families and conducted phylogenetic analyses of gene families with two to eight members in the complete genomes of Caenorhabditis elegans and Drosophila melanogaster and the available portion of the human genome. Although the human genome showed a higher proportion of recent gene duplications than the other animal genomes, the proportion of duplications after the deuterostome-protostome split was constant across families, with no peak of such duplications in four-member families, contrary to the expectation of the 2R hypothesis. A substantial majority (70.9%) of human four-member families and four-member clusters in larger families showed topologies inconsistent with two rounds of polyploidization in vertebrates.     

DNA Evidence for Strong Genome-Wide Pleiotropy of Cognitive and Learning Abilities

Very different neurocognitive processes appear to be involved in cognitive abilities such as verbal and non-verbal ability as compared to learning abilities taught in schools such as reading and mathematics. However, twin studies that compare similarity for monozygotic and dizygotic twins suggest that the same genes are largely responsible for genetic influence on these diverse aspects of cognitive function. It is now possible to test this evidence for strong pleiotropy using DNA alone from samples of unrelated individuals. Here we used this new method with 1.7 million DNA markers for a sample of 2,500 unrelated children at age 12 to investigate for the first time the extent of pleiotropy between general cognitive ability (aka intelligence) and learning abilities (reading, mathematics and language skills). We also compared these DNA results to results from twin analyses using the same sample and measures. The DNA-based method revealed strong genome-wide pleiotropy: Genetic correlations were greater than 0.70 between general cognitive ability and language, reading, and mathematics, results that were highly similar to twin study estimates of genetic correlations. These results indicate that genes related to diverse neurocognitive processes have general rather than specific effects.     

Evidence for an Association of the Dopamine D5 Receptor Gene on Age at Onset of Attention Deficit Hyperactivity Disorder

The purpose of this study was to determine whether the single nucleotide polymorphisms (SNPs) within candidate genes for attention deficit hyperactivity disorder (ADHD) are associated with the age at onset for ADHD. One hundred and forty-three SNPs were genotyped across five candidate genes ( DRD5 , SLC6A3 , HTR1B , SNAP25 , DRD4 ) for ADHD in 229 families with at least one affected offspring. SNPs with the highest estimated power to detect an association with age at onset were selected for each candidate gene, using a power-based screening procedure that does not compromise the nominal significance level. A time-to-onset analysis for family-based samples was performed on these SNPs to determine if an association exists with age at onset for ADHD. Seven consecutive SNPs surrounding the D5 dopamine receptor gene ( DRD5 ), were associated with the age at onset for ADHD; FDR adjusted q-values ranged from 0.008 to 0.023. This analysis indicates that individuals with the risk genotype develop ADHD earlier than individuals with any other genotype. A haplotype analysis across the 6 significant SNPs that were in linkage disequilibrium with one another, CTCATA , was also found to be significant (p-value = 0.02). We did not observe significant associations with age at onset for the other candidate loci tested. Although definitive conclusions await independent replication, these results suggest that a variant in DRD5 may affect age at onset for ADHD.     

Repurposing New Use for Old Drug Chloroquine against Metabolic Syndrome: A Review on Animal and Human Evidence

Chloroquine (CQ) and hydroxychloroquine (HCQ) are traditional anti-malarial drugs that have been repurposed for new therapeutic uses in many diseases due to their simple usage and cost-effectiveness. The pleiotropic effects of CQ and HCQ in regulating blood pressure, glucose homeostasis, lipid, and carbohydrate metabolism have been previously described in vivo and in humans, thus suggesting their role in metabolic syndrome (MetS) prevention. The anti-hyperglycaemic, anti-hyperlipidaemic, cardioprotective, anti-hypertensive, and anti-obesity effects of CQ and HCQ might be elicited through reduction of inflammatory response and oxidative stress, improvement of endothelial function, activation of insulin signalling pathway, inhibition of lipogenesis and autophagy, as well as regulation of adipokines and apoptosis. In conclusion, the current state of knowledge supported the repurposing of CQ and HCQ usage in the management of MetS.     

Evidence for Recombination in the Envelope Gene of Maedi-visna Virus

Frequent recombination occurs during replication in all retroviruses examined. This increases the genetic variation in the retroviral population and may be of importance in the evolution of the virus. Maedi-visna virus (MVV), a retrovirus of sheep, has a highly variable envelope gene. In a previous experiment, 20 sheep were infected with an uncloned strain of MVV and virus was isolated at regular intervals for 7 years. We sequenced the envelope genes of a number of these strains and found evidence for recombination that may have contributed to the observed high frequency of antigenic variants.     

Food–Nonfood Discrimination in Ancestral Vertebrates: Gamete Cannibalism and the Origin of the Adaptive Immune System

Adaptive immunity is a complex system that appeared twice in vertebrates (in gnathostomes and in jawless fish) although it is not required for invertebrate defence. The adaptive immune system is tightly associated with self–non‐self discrimination, and it is now clear that this interplay is not limited to the prevention of autoreactivity. Micro‐organisms are usually considered for their pathogenicity or symbiotic ability, but, for most small metazoans, they mainly constitute food. Vertebrates are characterized by feeding by predation on larger preys, when compared to their ancestors who were filter feeders and ate micro‐organisms. Predation gives a strong selective advantage, not only due to the availability of new food resources but also by the ability to eliminate competitors for environmental resources (intraguild predation (IGP)). Unlike size‐structured IGP, intraspecific predation of juveniles, zygotes or gametes can be detrimental for species fitness in some circumstances. The ability of individuals to recognize highly polymorphic molecules on the surface of gametes present in the plankton and so distinguish self versus non‐self gametes might have constituted a strong selective advantage in intraspecific competition. Here, I propose the theory that the capacity to rearrange receptors has been selected in ancestral vertebrates as a consequence of this strong need for discriminating between hetero‐cannibalism versus filial cannibalism. This evolutionary origin sheds light on presently unexplained features of the immune system, including the existence of regulatory T cells and of non‐pathogenic natural autoimmunity.     

Analysis of Evoked Electrical Activity in the Cerebral Cortex Using Wavelet Transformation at Different Stages of a Set to an Emotional Facial Expression

A set to an emotionally negative facial expression was developed in 35 healthy subjects. Wavelet transformation coefficients were used to study the rhythmic components of evoked EEG activity in the alpha and theta ranges at different stages of the set. Individual differences in the perception of facial expressions on activation of the set allowed the subjects to be divided into two groups: those with rapid (plastic sets) and those with prolonged extinction of its effects (rigid sets). At the set formation stage, subjects with rigid sets showed higher maximum coefficient values in the alpha rhythm than subjects with plastic sets. That EEG responses of subjects with plastic sets showed higher mean coefficients in the theta rhythm than subjects with rigid sets. Groups of subjects were characterized by different maximum and mean wavelet transformation coefficients, depending on the stage of the experiment. It is suggested that sets to an emotional facial expression were accompanied by different ratios of cortical-subcortical interactions both in different groups of subjects and at different stages of the experiment.     

Comparison of Genomes of Brucella melitensis M28 and the B. melitensis M5-90 Derivative Vaccine Strain Highlights the Translation Elongation Factor Tu Gene tuf2 as an Attenuation-Related Gene

Brucella melitensis causes brucellosis, a disease affecting sheep, cattle, and sometimes humans. Attenuated B. melitensis strain M5-90, derived from virulent strain M28, is widely used as a live vaccine in ruminants in China. Genetic differences between the strains may cast light on the mechanism of attenuation. We recently reported the complete genomic sequences of M28 and M5-90. Genome organization is highly conserved between these isolates, and also with virulent strains 16 M and ATCC 23457. Analysis revealed 23 open reading frames (ORFs) with consistent differences between M5-90 and the virulent strains. Notably, the tuf2 gene encoding translation elongation factor EF-Tu from M5-90 contained 50 single nucleotide polymorphisms (SNPs) and 9 gaps (indels) compared to tuf2 of M28 or of the other virulent strains. There were no changes in tuf1. To evaluate the potential role of EF-Tu in pathogenesis, tuf1 and tuf2 mutants of M28 and an M5-90 strain harboring wild-type tuf2 were constructed, and their virulence/attenuation was evaluated in vivo. We report that the tuf2 gene plays an important role in the attenuation of M5-90 virulence.     

Duplications and copy number variants of 8p23.1 are cytogenetically indistinguishable but distinct at the molecular level

It has been proposed that duplications of 8p23.1 are either euchromatic variants of the 8p23.1 defensin domain with no phenotypic consequences or true duplications associated with developmental delay and heart defects. Here, we provide evidence for both alternatives in two new families. A duplication of most of band 8p23.1 (circa 5 Mb) was found in a girl of 8 years with pulmonary stenosis and mild language delay. BAC fluorescence in situ hybridisation (FISH) and multiplex amplifiable probe hybridisation (MAPH) showed that the two copies of the duplicated segment were sited, in an alternating fashion, between three copies of a circa 300-450 kb segment from 8p23.1 distal to REPD. Copy number of the variable 8p23.1 defensin domain was consistent with duplication but within the normal range. Duplication of the GATA-binding protein 4 gene (GATA4) in this patient and others with and without heart defects, suggests it is a dosage-sensitive gene with variable penetrance. A cytogenetically similar duplication of 8p23.1 was found at prenatal diagnosis in a fetus, father and grandmother. There was no duplication using BAC FISH but MAPH showed 11 copies of the 360 kb variable defensin domain which is within the expanded range found in previous euchromatic variant carriers. Semiquantitative FISH (SQ-FISH) was consistent with a simultaneous expansion of the adjacent olfactory receptor repeats. These results distinguish duplications of 8p23.1 with clinically significant consequences from benign copy number variants, which have not yet been associated with qualitative or quantitative traits.     

New Evidence for the Prion Nature of Amyotrophic Leukospongiosis

New experimental data from studies of the etiopathogenesis of amyotrophic leukospongiosis (AL) obtained by immunoblotting and electron and atomic force microscopy are presented. This disease is characterized by gradual progression of flaccid paralysis of the limbs and truncal muscles with inevitable lethal outcomes due to spinal respiratory disorders; there are no impairments to cranial nerve function or pyramidal signs and all types of sensation are preserved. The main morphological appearances of AL consist of death of motoneurons throughout the spinal cord, combined with spongiosis of the white matter of the brain (axon degeneration on the background of preserved myelin sheaths). Human brain specimens obtained at autopsy from victims of AL always showed protease-resistant prion protein (PrPAL), aggregating into characteristic polymorphous amyloid-like structures. Previous studies have shown that brain homogenates and their highly purified fractions have in vitro infectivity. The new data provide evidence supporting the view that the disease has a prion nature and is one of the transmissible spongiform encephalopathies.     

Evidence for Non-additive Influence of Single Nucleotide Polymorphisms within the Apolipoprotein E Gene

We analyzed 13 single nucleotide polymorphisms (SNPs) within the apolipoprotein E (APOE) gene, to identify pairs of SNPs that interact in a non‐additive manner to influence genotypic mean levels of the ApoE protein in blood. An overparameterized general linear model of two‐SNP genotype means was applied to data from 456 female and 398 male unrelated European Americans from Rochester, MN, USA. We found statistically significant evidence for non‐additivity between SNPs within the male sample, but not within the female sample. We observed nine pairs of SNPs with evidence of non‐additivity at the α= 0.05 level of statistical significance within the male sample, when approximately three were expected by chance. Five of the nine pairs involved three SNPs (560, 624 and 1163) that did not have a statistically significant influence when considered separately in a single‐site analysis. Three of the nine pairs involving four SNPs (832, 1998, 3937 and 4951) showed significant evidence for non‐additivity in at least one of two other male samples from Jackson, MS, USA and North Karelia, Finland. Although all four of these SNPs had a statistically significant influence in Rochester when considered separately, only SNP 3937 gave a significant result in the other male samples. The four SNPs are located in the promoter, intronic and exonic regions, and 3' to the polyadenylation signal in the APOE gene. Our study suggests that analyses that only consider SNPs located in exons and ignore contexts such as those indexed by gender and population, and disregard non‐additivity of SNP effects, may inappropriately model the contribution of a gene to the genetic architecture of a trait that has a complex multifactorial etiology.     

Competitive ELISA of Chloramphenicol: Influence of Immunoreagent Structure and Application of the Method for the Inspection of Food of Animal Origin

An indirect competitive ELISA for the detection of chloramphenicol (CAP) in food of animal origin (milk, meat, eggs) is described. Influence of immunoreagent structure and composition on the assay sensitivity and specificity was investigated. Two CAP derivatives were used for conjugation with proteins: CAP succinate and a diazo derivative of CAP. Molar incorporation of CAP into the coating conjugates was also varied. To eliminate matrix effect on the assay results, a special casein-containing buffer was used for milk samples, whereas for meat and egg samples a 50-fold dilution of the buffer extracts was needed. The method developed permits CAP concentrations to be determined in the range 0.08-100 μg 1^-1. The detection limit is 0.08 μg kg^-1. Recovery in different food samples averages between 70 and 130%. The method can be applied for inspection of food of animal origin for CAP residues.     

Sequence Analysis of the Gene Encoding VP4 of a Bovine Group C Rotavirus: Molecular Evidence for a New P Genotype

Nucleotide sequence of the bovine group C rotavirus Shintoku strain gene 3 was determined. Segment 3 is 2253 nucleotides (nt) in length and contains a long open reading frame (ORF) beginning at nt 22 and terminating at nt 2223. This ORF encodes a polypeptide of 733 amino acids with a predicted molecular mass of 83 kDa. The deduced gene 3 amino acid sequence shares 79% and 73% identities with VP4 of the porcine Cowden and human Bristol strains, respectively. Lack of high amino acid sequence homology in VP4 of bovine, porcine, and human group C rotaviruses indicates that the Shintoku strain represents a new P genotype.     

Molecular Genetic Evidence Supporting the Clonality and Appendiceal Origin of Pseudomyxoma Peritonei in Women

Pseudomyxoma peritonei (PMP) is a poorly understood condition characterized by mucinous ascites and multifocal peritoneal mucinous tumors. Women with PMP often have mucinous tumors involving both the appendix and the ovaries. Several previous histopathological and immunohistochemical studies of PMP have suggested that most, if not all, cases of PMP in women are derived from mucinous adenomas of the appendix rather than from primary ovarian tumors. A few studies of the molecular genetics of PMP have been recently reported. However, these studies analyzed only a small number of cases and some included a heterogeneous group of mucinous tumors, including both benign and malignant appendiceal and ovarian tumors. We analyzed K-ras mutations and allelic losses of chromosomes 18q, 17p, 5q, and 6q in a substantial number of morphologically uniform cases of PMP with synchronous ovarian and appendiceal tumors as well as in appendiceal mucinous adenomas (MAs) and ovarian mucinous tumors of low malignant potential (MLMPs) unassociated with PMP. Each of the 16 PMP cases (100%) analyzed demonstrated identical K-ras mutations in the appendiceal adenoma and corresponding synchronous ovarian tumor. K-ras mutations were identified in 11 of 16 (69%) appendiceal MAs unassociated with PMP and in 12 of 16 (75%) ovarian MLMPs unassociated with PMP. Two PMP cases showed identical allelic losses in the matched ovarian and appendiceal tumors. A discordant pattern of allelic loss between the ovarian and appendiceal tumors at one or two of the loci tested was observed in six PMP cases. In all but one instance, LOH was observed in the ovarian tumor, whereas both alleles were retained in the matched appendiceal lesion, suggesting tumor progression in a secondary (metastatic) site. Our findings strongly support the conclusion that mucinous tumors involving the appendix and ovaries in women with PMP are clonal and derived from a single site, most likely the appendix.     

Evidence for an eye-centered spherical representation of the visuomotor map

During visually guided movement, visual coordinates of target location must be transformed into coordinates appropriate for movement. To investigate the representation of this visuomotor coordinate transformation, we examined changes in pointing behavior induced by a local visuomotor remapping. The visual feedback of finger position was limited to one location within the workspace, at which a discrepancy was introduced between the actual and visually perceived finger position. This remapping induced a change in pointing that extended over the entire workspace and was best captured by a spherical coordinate system centered near the eyes.     

Sex and Ontogenetic Variation in the Crest of Numida meleagris: Implications for Crested Vertebrates

Crested vertebrates are known from a wide variety of modern and fossil taxa, however, the actual formation and function of the crest is still debatable. Among modern birds, the globally distributed guinea fowl (Numida meleagris) is characterized by having a cranial bony crest (overlain by keratin), but surprisingly little is known about its development. Here, we studied the crest of 202 wild guinea fowl from the same population, using anatomical measurements as well as 2D-morphometry. Our results show that juveniles have smaller skulls than adults and have smaller, simpler crests that are visible even in very young individuals. Among adults, female skulls are smaller than males, and they have smaller, simpler shaped crests, which permit a discrimination between the sexes of 93% when the keratin is preserved with the bony crest, and of 89% when only the bony crest is available. By extrapolation, these results confirm that the crest can be used as an ontogenetic character, as well as for sex discrimination in the fossil record. Our results also show that the overlying keratin does not always mimic the underlying bony crest, which should be considered when reconstructing extinct crested vertebrates. Anat Rec, 303:1018–1034, 2020. © 2019 American Association for Anatomy     

基于靶基因文库的高通量测序平台建立肥厚型心肌病患者致病基因突变检测方法

目的 基于靶基因文库,应用下一代半导体高通量测序平台,建立快速、 准确的肥厚型心肌病(hypertrophic cardiomyopathy,HCM)常见致病基因突变检测方法 ,有利于HCM患者的早期预防及临床分子诊断.方法 选择国内外公认的与HCM致病相关的常见基因(MYH7、MYBPC3、TNNT2、TNNI3、A...     

A Rapid Method for the Separation of Functional Lymphoid Cell Populations of Human and Animal Origin on PVP-Silica (Percoll) Density Gradients

Subclasses of lymphocytes can be separated on gradients of non-toxic polyvinylpyrrolidone-coated colloidal silica (Percoll) by virtue of differential densities. Such gradients can yield functionally active lymphocyte populations after brief centrifugation. Gradients can be generated in a discontinuous step fashion and centrifuged in standard table-top laboratory centrifuges or as self-generating gradients during ultracentrifugation. The density medium has low viscosity and can be made isotonic for virtually any use. Gradients have proved useful in both human and experimental animal studies, and high percentage yields allow for separations from small cell numbers. Methods are described for separation of whole blood and lymphoid subpopuctions. The cytoxic capability of various density fractions was evaluated for mixed lymphocyte culture-induced allogeneic killing and spontaneous, so-called "natural" killer cell activity. The lower density associated with blast transformation allows for significant enrichments of stimulated cells from in vitro cultures. Higher thymidine incorporation, restimualtion in mixed lymphocyte reactions, and greater cytotoxic capacity are associated with these "blast" fractions.