The influence of copy number polymorphism on the human phenotype

The variability of human populations in a large part is determined by two complementary factors: environment and genetic information. Genetic variation is caused by different genetic variants (polymorphisms and mutations) present in the human genome. Until recently it was thought that most of these variants are small changes of one or several nucleotides (SNPs) which in their millions are present in the human genome. However, it was recently shown that there are also polymorphisms that extend over hundreds of thousands of DNA base pairs in the human genome. Such alternations called copy number variation (CNV) often include genes and other functional genetic elements. In this article we present the general characteristics of copy number polymorphism and we discuss some examples of CNVs that influence human phenotypes.     

Genetic architecture and balancing selection: the life and death of differentiated variants

Balancing selection describes any form of natural selection, which results in the persistence of multiple variants of a trait at intermediate frequencies within populations. By offering up a snapshot of multiple co‐occurring functional variants and their interactions, systems under balancing selection can reveal the evolutionary mechanisms favouring the emergence and persistence of adaptive variation in natural populations. We here focus on the mechanisms by which several functional variants for a given trait can arise, a process typically requiring multiple epistatic mutations. We highlight how balancing selection can favour specific features in the genetic architecture and review the evolutionary and molecular mechanisms shaping this architecture. First, balancing selection affects the number of loci underlying differentiated traits and their respective effects. Control by one or few loci favours the persistence of differentiated functional variants by limiting intergenic recombination, or its impact, and may sometimes lead to the evolution of supergenes. Chromosomal rearrangements, particularly inversions, preventing adaptive combinations from being dissociated are increasingly being noted as features of such systems. Similarly, due to the frequency of heterozygotes maintained by balancing selection, dominance may be a key property of adaptive variants. High heterozygosity and limited recombination also influence associated genetic load, as linked recessive deleterious mutations may be sheltered. The capture of deleterious elements in a locus under balancing selection may reinforce polymorphism by further promoting heterozygotes. Finally, according to recent genomewide scans, balanced polymorphism might be more pervasive than generally thought. We stress the need for both functional and ecological studies to characterize the evolutionary mechanisms operating in these systems.     

Colour polymorphism is likely to be disadvantageous to some populations and species due to genetic architecture and morph interactions

Polymorphism describes two or more distinct, genetically determined, phenotypes that co‐occur in the same population, where the rarest morph is maintained at a frequency above the mutation rate (Ford 1945; Huxley 1955). In a recent opinion piece, we explored a new idea regarding the role of genetic architectures and morph interactions in colour polymorphisms and how this can negatively affect population performance (Bolton et al. 2015). In this issue of Molecular Ecology, Forsman (2016) thoroughly discusses the current evidence for polymorphisms enhancing population performance and critiques the validity of the definitions of polymorphism we use in our original paper. We respond by clarifying that the negative consequences of polymorphisms that we discussed are likely to be most pertinent in species that have a particular set of characteristics, such as strong sexual or social interactions between morphs and discrete genetic architectures. Although it was not our intention to redefine polymorphism, we do believe that there should be further discussion about refining or characterizing balanced polymorphisms with respect to the degree of morph sympatry, discreteness of traits and their underlying genetic architecture, and the types of selection that drive and maintain the variation. The latter describes whether polymorphism is primarily maintained by external factors such as predation pressure or internal factors such as interactions with members of the same species. The contribution of Forsman (2016) is useful to this discussion, and we hope that our exchange of opinions will inspire new empirical and theoretical ideas on the origin and maintenance of colour polymorphisms.     

No mate preference associated with the supergene controlling social organization in Alpine silver ants

Disassortative mating is a powerful mechanism stabilizing polymorphisms at sex chromosomes and other supergenes. The Alpine silver ant, Formica selysi, has two forms of social organization—single‐queen and multiple‐queen colonies—determined by alternate haplotypes at a large supergene. Here, we explore whether mate preference contributes to the maintenance of the genetic polymorphism at the social supergene. With mate choice experiments, we found that females and males mated randomly with respect to social form. Moreover, queens were able to produce offspring irrespective of whether they had mated with a male from the same or the alternative social form. Yet, females originating from single‐queen colonies were more fertile, suggesting that they may be more successful at independent colony founding. We conclude that the pattern of asymmetric assortative mating documented from mature F. selysi colonies in the field is not caused by mate preferences or major genetic incompatibilities between social forms. More generally, we found no evidence that disassortative mate preference contributes to the maintenance of polymorphism at this supergene controlling ant social organization.     

Sexual selection and genetic colour polymorphisms in animals

Genetic colour polymorphisms are widespread across animals and often subjected to complex selection regimes. Traditionally, colour morphs were used as simple visual markers to measure allele frequency changes in nature, selection, population divergence and speciation. With advances in sequencing technology and analysis methods, several model systems are emerging where the molecular targets of selection are being described. Here, we discuss recent studies on the genetics of sexually selected colour polymorphisms, aiming at (i) reviewing the evidence of sexual selection on colour polymorphisms, (ii) highlighting the genetic architecture, molecular and developmental basis underlying phenotypic colour diversification and (iii) discuss how the maintenance of such polymorphisms might be facilitated or constrained by these. Studies of the genetic architecture of colour polymorphism point towards the importance of tight clustering of colour loci with other trait loci, such as in the case of inversions and supergene structures. Other interesting findings include linkage between colour loci and mate preferences or sex determination, and the role of introgression and regulatory variation in fuelling polymorphisms. We highlight that more studies are needed that explicitly integrate fitness consequences of sexual selection on colour with the underlying molecular targets of colour to gain insights into the evolutionary consequences of sexual selection on polymorphism maintenance.     

Evolution of balanced genetic polymorphism

Extreme genetic polymorphism maintained by balancing selection (so called because many alleles are maintained in a balance by a mechanism of rare allele advantage) is intimately associated with the important task of self/non-self-discrimination. Widely disparate self-recognition systems of plants, animals and fungi share several general features, including the maintenance of large numbers of alleles at relatively even frequency, and persistence of this variation over very long time periods. Because the evolutionary dynamics of balanced polymorphism are very different from those of neutral genetic variation, data on balanced polymorphism have been used as a novel source for inference of the history of populations. This review highlights the unique evolutionary properties of balanced genetic polymorphism, and the use of theoretical understanding in analysis and application of empirical data for inference of population history. However, a second goal of this review is to point out where current theory is incomplete. Recent observations suggest that entirely novel selective forces may act in concert with balancing selection, and these novel forces may be extremely potent in shaping genetic variation at self-recognition loci.     

Association of Hck genetic polymorphisms with gene expression and COPD

Polymorphonuclear leukocytes (PMNs) are major effector cells in the chronic airway inflammation in chronic obstructive pulmonary disease (COPD). PMN degranulation is associated with degradation of extracellular matrix and tissue damage. Hck is an essential molecule in the signaling pathway regulating PMN degranulation. We hypothesized that polymorphisms affect the expression level of Hck, which, in turn, modulates PMN mediator release and tissue damage and influences the development of COPD. Here we systematically investigated genetic tag polymorphisms of the Hck gene, Hck mRNA and protein expression pattern in PMNs, and PMN mediator release (myeloperoxidase) in 60 healthy white subjects, and assessed their association with the use of several genetic models. The association of genetic polymorphisms with COPD-related phenotypes was determined in the lung healthy study cohort (LHS). We identified a novel 15 bp insertion/deletion polymorphism (8,656 L/S) in intron 1 of the Hck gene, which was associated with differential expression of Hck protein and PMN myeloperoxidase release. In the LHS cohort, there was significant interaction between the 8,656 L/S polymorphism and smoking on baseline lung function and 8,656 L/S was associated with bronchodilator response. These data suggest that the insertion/deletion polymorphism could be a functional polymorphism of the Hck gene, may contribute to COPD pathogenesis and modify COPD-related phenotypes.     

The diversity of microbes: resurgence of the phenotype

The introduction of molecular genetic methods has caused confusion about the nature of microbial species. Environmental DNA extraction has indicated the existence of a vast diversity of genotypes, but how this relates to functional and phenotypic diversity has not been sufficiently explored. It has been implied that genetic distance per se correlates with phenotypic differentiation and thus reflects subtle (but undiscovered) adaptive fine-tuning to the environment, and that microbes may show biogeographic patterns at the genetic level. Here, we argue that no theoretically based species concept exists; species represent only the basic unit in the taxonomic hierarchy. The significance of naming species is that it organizes biological information. The reason why microbial species collectively represent large genetic differences is owing to huge absolute population sizes, absence of allopatric speciation and low extinction rates. Microbial phenotypes are, therefore, ancient in terms of the geological time-scale and have been maintained through stabilizing selection. These problems are discussed with special reference to eukaryotic micro-organisms.     

Molecular polymorphism distribution in phenotypically distinct populations of wine yeast strains.

Electrophoretic karyotyping and mitochondrial DNA restriction analysis were used to analyze natural yeast populations from fermenting musts in El Penedès, Spain. Both analyses revealed a considerable degree of polymorphism, indicating heterogeneous natural populations. By specifically designed genetic selection protocols, strains showing potentially interesting phenotypes, such as high tolerance to ethanol and temperature or the ability to grow and to ferment in wine-water-sugar mixtures, were isolated from these natural populations. Genetic analysis showed a strong correlation between the selected phenotypes and mitochondrial DNA polymorphisms. Karyotype analysis revealed several genetically similar yeast lineages in the natural yeast microflora, which we interpret as genetically isolated subpopulations of yeast strains with distinct genetic traits, which may correspond to specific microenvironments. Thus, molecular polymorphism analysis may be useful not only to study the geographical distribution of natural yeast strains but also to identify strains with specific phenotypic properties.     

Polymorphism of melanic ladybirds maintained by frequency-dependent sexual selection

Sexual selection, whether by female preference or male competition, is almost inevitably frequency-dependent. Female preference gives rise to a 'rare male effect', by which the rarer male phenotypes gain a relatively greater selective advantage. In addition to this effect, the proportion of females expressing a preference may also be frequency-dependent.
Frequency-dependent expression of mating preference can arise in at least two ways: (1) when females encounter a succession of courting males while searching for a male they prefer; (2) when females chose a male from within a lek. Models of mating behaviour reveal a clear distinction between the frequency dependence in the expression of female preference and the frequency dependence in the consequent selection of the males. When expression of preference is highly dependent on frequency, the selection of males is constant or only slightly frequency-dependent: constant expression of preference produces high frequency dependence of selection. Analysis of general models shows that genetic polymorphisms can be maintained under a wide range of conditions.
The ladybird, Adalia bipunctata , is polymorphic for several melanic and non-melanic phenotypes. Females have a genetically determined preference for melanic males. Non-melanic phenotypes mate assortatively. By estimating the parameters of a detailed model of natural selection, sexual selection and assortative mating, it has been shown that the Adalia bipunctata polymorphism will be maintained at frequencies observed in the wild.     

Tracking human migrations by the analysis of the distribution of HLA alleles, lineages and haplotypes in closed and open populations

The human leucocyte antigen (HLA) system shows extensive variation in the number and function of loci and the number of alleles present at any one locus. Allele distribution has been analysed in many populations through the course of several decades, and the implementation of molecular typing has significantly increased the level of diversity revealing that many serotypes have multiple functional variants. While the degree of diversity in many populations is equivalent and may result from functional polymorphism(s) in peptide presentation, homogeneous and heterogeneous populations present contrasting numbers of alleles and lineages at the loci with high-density expression products. In spite of these differences, the homozygosity levels are comparable in almost all of them. The balanced distribution of HLA alleles is consistent with overdominant selection. The genetic distances between outbred populations correlate with their geographical locations; the formal genetic distance measurements are larger than expected between inbred populations in the same region. The latter present many unique alleles grouped in a few lineages consistent with limited founder polymorphism in which any novel allele may have been positively selected to enlarge the communal peptide-binding repertoire of a given population. On the other hand, it has been observed that some alleles are found in multiple populations with distinctive haplotypic associations suggesting that convergent evolution events may have taken place as well. It appears that the HLA system has been under strong selection, probably owing to its fundamental role in varying immune responses. Therefore, allelic diversity in HLA should be analysed in conjunction with other genetic markers to accurately track the migrations of modern humans.     

Functional and geographical differentiation of candidate balanced polymorphisms in Arabidopsis thaliana

Molecular population genetic analysis of three chromosomal regions in Arabidopsis thaliana suggested that balancing selection might operate to maintain variation at three novel candidate adaptive trait genes, including SOLUBLE STARCH SYNTHASE I (SSI) , PLASTID TRANSCRIPTIONALLY ACTIVE 7(PTAC7) , and BELL-LIKE HOMEODOMAIN 10 (BLH10). If balanced polymorphisms are indeed maintained at these loci, then we would expect to observe functional variation underlying the previously detected signatures of selection. We observe multiple replacement polymorphisms within and in the 32 amino acids just upstream of the protein–protein interacting BELL domain at the BLH10 locus. While no clear protein sequence differences are found between allele types in SSI and PTAC7, these two genes show evidence for allele-specific variation in expression levels. Geographical patterns of allelic differentiation seem consistent with population stratification in this species and a significant longitudinal cline was observed at all three candidate loci. These data support a hypothesis of balancing selection at all three candidate loci and provide a basis for more detailed functional work by identifying possible functional differences that might be selectively maintained.     

Tissue-specific apoptotic effects of the p53 codon 72 polymorphism in a mouse model

Currently there are several dozen human polymorphisms that have been loosely associated with cancer risk. Correlating such variants with cancer risk has been challenging, primarily due to factors such as genetic heterogeneity, contributions of diet and environmental factors, and the difficulty in obtaining large sample sizes for analysis. Such difficulties can be circumvented with the establishment of mouse models for human variants. Recently, several groups have modeled human cancer susceptibility polymorphisms in the mouse. Remarkably, in each case these mouse models have accurately reflected human phenotypes, and clarified the contribution of these variants to cancer risk. We recently reported on a mouse model for the codon 72 polymorphism in p53, and found that this polymorphism regulates the ability to cooperate with NF-kB and induce apoptosis. Here-in we present evidence that this polymorphism impacts the apoptotic function of p53 in a tissue-specific manner; such tissue-specific effects of polymorphic variants represent an added challenge to human cancer risk association studies. The data presented here support the premise that modeling human polymorphisms in the mouse represents a powerful tool to assess the impact of these variants on cancer risk, progression and therapy.     

Genomic Evidence of Rapid and Stable Adaptive Oscillations over Seasonal Time Scales in Drosophila

In many species, genomic data have revealed pervasive adaptive evolution indicated by the fixation of beneficial alleles. However, when selection pressures are highly variable along a species'' range or through time adaptive alleles may persist at intermediate frequencies for long periods. So called “balanced polymorphisms” have long been understood to be an important component of standing genetic variation, yet direct evidence of the strength of balancing selection and the stability and prevalence of balanced polymorphisms has remained elusive. We hypothesized that environmental fluctuations among seasons in a North American orchard would impose temporally variable selection on Drosophila melanogaster that would drive repeatable adaptive oscillations at balanced polymorphisms. We identified hundreds of polymorphisms whose frequency oscillates among seasons and argue that these loci are subject to strong, temporally variable selection. We show that these polymorphisms respond to acute and persistent changes in climate and are associated in predictable ways with seasonally variable phenotypes. In addition, our results suggest that adaptively oscillating polymorphisms are likely millions of years old, with some possibly predating the divergence between D. melanogaster and D. simulans. Taken together, our results are consistent with a model of balancing selection wherein rapid temporal fluctuations in climate over generational time promotes adaptive genetic diversity at loci underlying polygenic variation in fitness related phenotypes.     

Effective utilization of genetic information for athletes and coaches: focus on ACTN3 R577X polymorphism

Training variants (type, intensity, and duration of exercise) can be selected according to individual aims and fitness assessment. Recently, various methods of resistance and endurance training have been used for muscle hypertrophy and VO₂max improvement. Although several genetic variants are associated with elite athletic performance and muscle phenotypes, genetic background has not been used as variant for physical training. ACTN3 R577X is a well-studied genetic polymorphism. It is the only genotype associated with elite athletic performance in multiple cohorts. This association is strongly supported by mechanistic data from an Actn3-knockout mouse model. In this review, possible guidelines are discussed for effective utilization of ACTN3 R577X polymorphism for physical training.     

Tissue-specific apoptotic effects of the p53 codon 72 polymorphism in a mouse model

Currently there are several dozen human polymorphisms that have been loosely associated with cancer risk. Correlating such variants with cancer risk has been challenging, primarily due to factors such as genetic heterogeneity, contributions of diet and environmental factors, and the difficulty in obtaining large sample sizes for analysis. Such difficulties can be circumvented with the establishment of mouse models for human variants. Recently, several groups have modeled human cancer susceptibility polymorphisms in the mouse. Remarkably, in each case these mouse models have accurately reflected human phenotypes, and clarified the contribution of these variants to cancer risk. We recently reported on a mouse model for the codon 72 polymorphism in p53, and found that this polymorphism regulates the ability to cooperate with NFκB and induce apoptosis. Here-in we present evidence that this polymorphism impacts the apoptotic function of p53 in a tissue-specific manner; such tissue-specific effects of polymorphic variants represent an added challenge to human cancer risk association studies. The data presented here support the premise that modeling human polymorphisms in the mouse represents a powerful tool to assess the impact of these variants on cancer risk, progression and therapy.Key words: p53, polymorphism, apoptosis, codon 72, NFκB     

DNA polymorphisms as modulators of genotoxicity and cancer

Cancer arises as a result of several factors, including multiple genes and environmental exposures. It is generally accepted that genetic polymorphisms are associated with most common disorders like cancer. The majority of polymorphisms are single nucleotide polymorphisms (SNPs) which occur with a frequency of 10(-6). Susceptibility-conferring alleles are not sufficient to cause disease, but modulate the risk in combination with other alleles and environmental exposures, except in the extreme case of Mendelian cancer syndromes (e.g. FAP, HNPCC, Rb). The Environmental Genome Project identifies, among others, two lines of research along which we have been working and are the topic of the present paper, namely (i) allele-disease associations and (ii) functional studies of allelic variants. Case-control association studies conducted by us and others showed that polymorphism at a single site could increase risk-predictability by a factor < 2. It is known, however, that the individual risk predictability increases by associating multiple genetic polymorphisms as was demonstrated for breast, renal and thyroid cancer. Functional genomics of the putative susceptibility-alleles involved in cancers can improve substantially the strength of association studies. This calls for cell-systems capable of tracking different gene activities, which may clarify the possible role of allelic variants in certain cancers. This endeavour is likely to be met by the bacterial tester strain, MTC, described here.     

Simple inheritance,complex regulation: Supergene‐mediated fire ant queen polymorphism

The fire ant Solenopsis invicta exists in two alternate social forms: monogyne nests contain a single reproductive queen and polygyne nests contain multiple reproductive queens. This colony‐level social polymorphism corresponds with individual differences in queen physiology, queen dispersal patterns and worker discrimination behaviours, all evidently regulated by an inversion‐based supergene that spans more than 13 Mb of a “social chromosome,” contains over 400 protein‐coding genes and rarely undergoes recombination. The specific mechanisms by which this supergene influences expression of the many distinctive features that characterize the alternate forms remain almost wholly unknown. To advance our understanding of these mechanisms, we explore the effects of social chromosome genotype and natal colony social form on gene expression in queens sampled as they embarked on nuptial flights, using RNA‐sequencing of brains and ovaries. We observe a large effect of natal social form, that is, of the social/developmental environment, on gene expression profiles, with similarly substantial effects of genotype, including: (a) supergene‐associated gene upregulation, (b) allele‐specific expression and (c) pronounced extra‐supergene trans‐regulatory effects. These findings, along with observed spatial variation in differential and allele‐specific expression within the supergene region, highlight the complex gene regulatory landscape that emerged following divergence of the inversion‐mediated Sb haplotype from its homologue, which presumably largely retained the ancestral gene order. The distinctive supergene‐associated gene expression trajectories we document at the onset of a queen’s reproductive life expand the known record of relevant molecular correlates of a complex social polymorphism and point to putative genetic factors underpinning the alternate social syndromes.