Great ape DNA sequences reveal a reduced diversity and an expansion in humans

The extent of DNA sequence variation of chimpanzees is several-fold greater than that of humans. It is unclear, however, if humans or chimpanzees are exceptional among primates in having low and high amounts of DNA sequence diversity, respectively. To address this, we have determined approximately 10,000 bp of noncoding DNA sequences at Xq13.3 (which has been extensively studied in both humans and chimpanzees) from 10 western lowland gorillas (Gorilla gorilla gorilla) and 1 mountain gorilla (Gorilla gorilla beringei; that is, from 2 of the 3 currently recognized gorilla subspecies), as well as 8 Bornean (Pongo pygmaeus pygmaeus) and 6 Sumatran (Pongo pygmaeus abelii) orang-utans, representing both currently recognized orang-utan subspecies. We show that humans differ from the great apes in having a low level of genetic variation and a signal of population expansion.     

Evidence of widespread selection on standing variation in Europe at height-associated SNPs

Strong signatures of positive selection at newly arising genetic variants are well documented in humans, but this form of selection may not be widespread in recent human evolution. Because many human traits are highly polygenic and partly determined by common, ancient genetic variation, an alternative model for rapid genetic adaptation has been proposed: weak selection acting on many pre-existing (standing) genetic variants, or polygenic adaptation. By studying height, a classic polygenic trait, we demonstrate the first human signature of widespread selection on standing variation. We show that frequencies of alleles associated with increased height, both at known loci and genome wide, are systematically elevated in Northern Europeans compared with Southern Europeans (P < 4.3 × 10(-4)). This pattern mirrors intra-European height differences and is not confounded by ancestry or other ascertainment biases. The systematic frequency differences are consistent with the presence of widespread weak selection (selection coefficients ～10(-3)-10(-5) per allele) rather than genetic drift alone (P < 10(-15)).     

Detection of regulatory variation in mouse genes

Functional polymorphism in genes can be classified as coding variation, altering the amino-acid sequence of the encoded protein, or regulatory variation, affecting the level or pattern of expression of the gene. Coding variation can be recognized directly from DNA sequence, and consequently its frequency and characteristics have been extensively described. By contrast, virtually nothing is known about the extent to which gene regulation varies in populations. Yet it is likely that regulatory variants are important in modulating gene function: alterations in gene regulation have been proposed to influence disease susceptibility and to have been the primary substrate for the evolution of species. Here, we report a systematic study to assess the extent of cis-acting regulatory variation in 69 genes across four inbred mouse strains. We find that at least four of these genes show allelic differences in expression level of 1.5-fold or greater, and that some of these differences are tissue specific. The results show that the impact of regulatory variants can be detected at a significant frequency in a genomic survey and suggest that such variation may have important consequences for organismal phenotype and evolution. The results indicate that larger-scale surveys in both mouse and human could identify a substantial number of genes with common regulatory variation.     

Common genetic variants account for differences in gene expression among ethnic groups

Variation in DNA sequence contributes to individual differences in quantitative traits, but in humans the specific sequence variants are known for very few traits. We characterized variation in gene expression in cells from individuals belonging to three major population groups. This quantitative phenotype differs significantly between European-derived and Asian-derived populations for 1,097 of 4,197 genes tested. For the phenotypes with the strongest evidence of cis determinants, most of the variation is due to allele frequency differences at cis-linked regulators. The results show that specific genetic variation among populations contributes appreciably to differences in gene expression phenotypes. Populations differ in prevalence of many complex genetic diseases, such as diabetes and cardiovascular disease. As some of these are probably influenced by the level of gene expression, our results suggest that allele frequency differences at regulatory polymorphisms also account for some population differences in prevalence of complex diseases.     

Comparison of genome degradation in Paratyphi A and Typhi, human-restricted serovars of Salmonella enterica that cause typhoid

Salmonella enterica serovars often have a broad host range, and some cause both gastrointestinal and systemic disease. But the serovars Paratyphi A and Typhi are restricted to humans and cause only systemic disease. It has been estimated that Typhi arose in the last few thousand years. The sequence and microarray analysis of the Paratyphi A genome indicates that it is similar to the Typhi genome but suggests that it has a more recent evolutionary origin. Both genomes have independently accumulated many pseudogenes among their approximately 4,400 protein coding sequences: 173 in Paratyphi A and approximately 210 in Typhi. The recent convergence of these two similar genomes on a similar phenotype is subtly reflected in their genotypes: only 30 genes are degraded in both serovars. Nevertheless, these 30 genes include three known to be important in gastroenteritis, which does not occur in these serovars, and four for Salmonella-translocated effectors, which are normally secreted into host cells to subvert host functions. Loss of function also occurs by mutation in different genes in the same pathway (e.g., in chemotaxis and in the production of fimbriae).     

High-throughput sequencing provides insights into genome variation and evolution in Salmonella Typhi

Isolates of Salmonella enterica serovar Typhi (Typhi), a human-restricted bacterial pathogen that causes typhoid, show limited genetic variation. We generated whole-genome sequences for 19 Typhi isolates using 454 (Roche) and Solexa (Illumina) technologies. Isolates, including the previously sequenced CT18 and Ty2 isolates, were selected to represent major nodes in the phylogenetic tree. Comparative analysis showed little evidence of purifying selection, antigenic variation or recombination between isolates. Rather, evolution in the Typhi population seems to be characterized by ongoing loss of gene function, consistent with a small effective population size. The lack of evidence for antigenic variation driven by immune selection is in contrast to strong adaptive selection for mutations conferring antibiotic resistance in Typhi. The observed patterns of genetic isolation and drift are consistent with the proposed key role of asymptomatic carriers of Typhi as the main reservoir of this pathogen, highlighting the need for identification and treatment of carriers.     

Systematic screen for human disease genes in yeast

High similarity between yeast and human mitochondria allows functional genomic study of Saccharomyces cerevisiae to be used to identify human genes involved in disease. So far, 102 heritable disorders have been attributed to defects in a quarter of the known nuclear-encoded mitochondrial proteins in humans. Many mitochondrial diseases remain unexplained, however, in part because only 40-60% of the presumed 700-1,000 proteins involved in mitochondrial function and biogenesis have been identified. Here we apply a systematic functional screen using the pre-existing whole-genome pool of yeast deletion mutants to identify mitochondrial proteins. Three million measurements of strain fitness identified 466 genes whose deletions impaired mitochondrial respiration, of which 265 were new. Our approach gave higher selection than other systematic approaches, including fivefold greater selection than gene expression analysis. To apply these advantages to human disorders involving mitochondria, human orthologs were identified and linked to heritable diseases using genomic map positions.     

Analyses of X-linked and autosomal genetic variation in population-scale whole genome sequencing

The ratio of genetic diversity on chromosome X to that on the autosomes is sensitive to both natural selection and demography. On the basis of whole-genome sequences of 69 females, we report that whereas this ratio increases with genetic distance from genes across populations, it is lower in Europeans than in West Africans independent of proximity to genes. This relative reduction is most parsimoniously explained by differences in demographic history without the need to invoke natural selection.     

Comparative genomes of Chlamydia pneumoniae and C. trachomatis

Chlamydia are obligate intracellular eubacteria that are phylogenetically separated from other bacterial divisions. C. trachomatis and C. pneumoniae are both pathogens of humans but differ in their tissue tropism and spectrum of diseases. C. pneumoniae is a newly recognized species of Chlamydia that is a natural pathogen of humans, and causes pneumonia and bronchitis. In the United States, approximately 10% of pneumonia cases and 5% of bronchitis cases are attributed to C. pneumoniae infection. Chronic disease may result following respiratory-acquired infection, such as reactive airway disease, adult-onset asthma and potentially lung cancer. In addition, C. pneumoniae infection has been associated with atherosclerosis. C. trachomatis infection causes trachoma, an ocular infection that leads to blindness, and sexually transmitted diseases such as pelvic inflammatory disease, chronic pelvic pain, ectopic pregnancy and epididymitis. Although relatively little is known about C. trachomatis biology, even less is known concerning C. pneumoniae. Comparison of the C. pneumoniae genome with the C. trachomatis genome will provide an understanding of the common biological processes required for infection and survival in mammalian cells. Genomic differences are implicated in the unique properties that differentiate the two species in disease spectrum. Analysis of the 1,230,230-nt C. pneumoniae genome revealed 214 protein-coding sequences not found in C. trachomatis, most without homologues to other known sequences. Prominent comparative findings include expansion of a novel family of 21 sequence-variant outer-membrane proteins, conservation of a type-III secretion virulence system, three serine/threonine protein kinases and a pair of parologous phospholipase-D-like proteins, additional purine and biotin biosynthetic capability, a homologue for aromatic amino acid (tryptophan) hydroxylase and the loss of tryptophan biosynthesis genes.     

Regulatory changes underlying expression differences within and between Drosophila species

Differences in gene expression are an important source of phenotypic variation, and can be caused by changes in cis and/or trans regulation. cis-regulatory variants alter allele-specific expression, whereas trans-regulatory variants influence expression of both alleles in a diploid cell. Because of this difference, we hypothesize that natural selection may favor one type of change over the other. Here, we investigate contributions of cis- and trans-regulatory changes to variable intra- and interspecific gene expression using four strains of Drosophila melanogaster, three strains of D. simulans and a total of 78 genes. We show that cis-regulatory changes account for a greater proportion of the expression differences observed between rather than within species. These data are inconsistent with a neutral model assuming equal probabilities of fixation for cis- and trans-regulatory polymorphisms, suggesting that natural selection influences the molecular mechanisms underlying divergent gene expression. Specifically, cis-regulatory changes seem to accumulate preferentially over time.     

Adaptive evolution of a duplicated pancreatic ribonuclease gene in a leaf-eating monkey

Although the complete genome sequences of over 50 representative species have revealed the many duplicated genes in all three domains of life, the roles of gene duplication in organismal adaptation and biodiversity are poorly understood. In addition, the evolutionary forces behind the functional divergence of duplicated genes are often unknown, leading to disagreement on the relative importance of positive Darwinian selection versus relaxation of functional constraints in this process. The methodology of earlier studies relied largely on DNA sequence analysis but lacked functional assays of duplicated genes, frequently generating contentious results. Here we use both computational and experimental approaches to address these questions in a study of the pancreatic ribonuclease gene (RNASE1) and its duplicate gene (RNASE1B) in a leaf-eating colobine monkey, douc langur. We show that RNASE1B has evolved rapidly under positive selection for enhanced ribonucleolytic activity in an altered microenvironment, a response to increased demands for the enzyme for digesting bacterial RNA. At the same time, the ability to degrade double-stranded RNA, a non-digestive activity characteristic of primate RNASE1, has been lost in RNASE1B, indicating functional specialization and relaxation of purifying selection. Our findings demonstrate the contribution of gene duplication to organismal adaptation and show the power of combining sequence analysis and functional assays in delineating the molecular basis of adaptive evolution.     

Parallel adaptive origins of digestive RNases in Asian and African leaf monkeys

Similar morphological or physiological changes occurring in multiple evolutionary lineages are not uncommon. Such parallel changes are believed to be adaptive, because a complex character is unlikely to originate more than once by chance. However, the occurrence of adaptive parallel amino acid substitutions is debated. Here I propose four requirements for establishing adaptive parallel evolution at the protein sequence level and use these criteria to demonstrate such a case. I report that the gene encoding pancreatic ribonuclease was duplicated independently in Asian and African leaf-eating monkeys. Statistical analyses of DNA sequences, functional assays of reconstructed ancestral proteins and site-directed mutagenesis show that the new genes acquired enhanced digestive efficiencies through parallel amino acid replacements driven by darwinian selection. They also lost a non-digestive function independently, under a relaxed selective constraint. These results demonstrate that despite the overall stochasticity, even molecular evolution has a certain degree of repeatability and predictability under the pressures of natural selection.     

The genetic basis for differences in leaf form between Arabidopsis thaliana and its wild relative Cardamine hirsuta

A key question in biology is how differences in gene function or regulation produce new morphologies during evolution. Here we investigate the genetic basis for differences in leaf form between two closely related plant species, Arabidopsis thaliana and Cardamine hirsuta. We report that in C. hirsuta, class I KNOTTED1-like homeobox (KNOX) proteins are required in the leaf to delay cellular differentiation and produce a dissected leaf form, in contrast to A. thaliana, in which KNOX exclusion from leaves results in a simple leaf form. These differences in KNOX expression arise through changes in the activity of upstream gene regulatory sequences. The function of ASYMMETRIC LEAVES1/ROUGHSHEATH2/PHANTASTICA (ARP) proteins to repress KNOX expression is conserved between the two species, but in C. hirsuta the ARP-KNOX regulatory module controls new developmental processes in the leaf. Thus, evolutionary tinkering with KNOX regulation, constrained by ARP function, may have produced diverse leaf forms by modulating growth and differentiation patterns in developing leaf primordia.     

Network modeling links breast cancer susceptibility and centrosome dysfunction

Many cancer-associated genes remain to be identified to clarify the underlying molecular mechanisms of cancer susceptibility and progression. Better understanding is also required of how mutations in cancer genes affect their products in the context of complex cellular networks. Here we have used a network modeling strategy to identify genes potentially associated with higher risk of breast cancer. Starting with four known genes encoding tumor suppressors of breast cancer, we combined gene expression profiling with functional genomic and proteomic (or 'omic') data from various species to generate a network containing 118 genes linked by 866 potential functional associations. This network shows higher connectivity than expected by chance, suggesting that its components function in biologically related pathways. One of the components of the network is HMMR, encoding a centrosome subunit, for which we demonstrate previously unknown functional associations with the breast cancer-associated gene BRCA1. Two case-control studies of incident breast cancer indicate that the HMMR locus is associated with higher risk of breast cancer in humans. Our network modeling strategy should be useful for the discovery of additional cancer-associated genes.     

The melanocyte differentiation program predisposes to metastasis after neoplastic transformation

The aggressive clinical behavior of melanoma suggests that the developmental origins of melanocytes in the neural crest might be relevant to their metastatic propensity. Here we show that primary human melanocytes, transformed using a specific set of introduced genes, form melanomas that frequently metastasize to multiple secondary sites, whereas human fibroblasts and epithelial cells transformed using an identical set of genes generate primary tumors that rarely do so. Notably, these melanomas have a metastasis spectrum similar to that observed in humans with melanoma. These observations indicate that part of the metastatic proclivity of melanoma is attributable to lineage-specific factors expressed in melanocytes and not in other cell types analyzed. Analysis of microarray data from human nevi shows that the expression pattern of Slug, a master regulator of neural crest cell specification and migration, correlates with those of other genes that are important for neural crest cell migrations during development. Moreover, Slug is required for the metastasis of the transformed melanoma cells. These findings indicate that melanocyte-specific factors present before neoplastic transformation can have a pivotal role in governing melanoma progression.