Intron length evolution in Drosophila

I present data on the evolution of intron lengths among 3 closely related Drosophila species, D. melanogaster, Drosophila simulans, and Drosophila yakuba. Using D. yakuba as an outgroup, I mapped insertion and deletion mutations in 148 introns (spanning approximately 30 kb) to the D. melanogaster and D. simulans lineages. Intron length evolution in the 2 sister species has been different: in D. melanogaster, X-linked introns have increased slightly in size, whereas autosomal ones have decreased slightly in size; in D. simulans, both X-linked and autosomal introns have decreased in size. To understand the possible evolutionary causes of these lineage- and chromosome-specific patterns of intron evolution, I studied insertion-deletion (indel) polymorphism and divergence in D. melanogaster. Small insertion mutations segregate at elevated frequencies and enjoy elevated probabilities of fixation, particularly on the X chromosome. In contrast, there is no detectable X chromosome effect on fixations in D. simulans. These findings suggest X chromosome-specific selection or biased gene conversion-gap repair favoring insertions in D. melanogaster but not in D. simulans. These chromosome- and lineage-specific patterns of indel substitution are not easily explained by existing general population genetic models of intron length evolution. Genomic data from D. melanogaster further suggest that the forces described here affect introns and intergenic regions similarly.     

Insertions and deletions trigger adaptive walks in Drosophila proteins

Maps that relate all possible genotypes or phenotypes to fitness--fitness landscapes--are central to the evolution of life, but remain poorly known. An insertion or a deletion (indel) of one or several amino acids constitutes a substantial leap of a protein within the space of amino acid sequences, and it is unlikely that after such a leap the new sequence corresponds precisely to a fitness peak. Thus, one can expect an indel in the protein-coding sequence that gets fixed in a population to be followed by some number of adaptive amino acid substitutions, which move the new sequence towards a nearby fitness peak. Here, we study substitutions that occur after a frame-preserving indel in evolving proteins of Drosophila. An insertion triggers 1.03 ± 0.75 amino acid substitutions within the protein region centred at the site of insertion, and a deletion triggers 4.77 ± 1.03 substitutions within such a region. The difference between these values is probably owing to a higher fraction of effectively neutral insertions. Almost all of the triggered amino acid substitutions can be attributed to positive selection, and most of them occur relatively soon after the triggering indel and take place upstream of its site. A high fraction of substitutions that follow an indel occur at previously conserved sites, suggesting that an indel substantially changes selection that shapes the protein region around it. Thus, an indel is often followed by an adaptive walk of length that is in agreement with the theory of molecular adaptation.     

Is the rate of insertion and deletion mutation male biased?: Molecular evolutionary analysis of avian and primate sex chromosome sequences.

The rate of mutation for nucleotide substitution is generally higher among males than among females, likely owing to the larger number of DNA replications in spermatogenesis than in oogenesis. For insertion and deletion (indel) mutations, data from a few human genetic disease loci indicate that the two sexes may mutate at similar rates, possibly because such mutations arise in connection with meiotic crossing over. To address origin- and sex-specific rates of indel mutation we have conducted the first large-scale molecular evolutionary analysis of indels in noncoding DNA sequences from sex chromosomes. The rates are similar on the X and Y chromosomes of primates but about twice as high on the avian Z chromosome as on the W chromosome. The fact that indels are not uncommon on the nonrecombining Y and W chromosomes excludes meiotic crossing over as the main cause of indel mutation. On the other hand, the similar rates on X and Y indicate that the number of DNA replications (higher for Y than for X) is also not the main factor. Our observations are therefore consistent with a role of both DNA replication and recombination in the generation of short insertion and deletion mutations. A significant excess of deletion compared to insertion events is observed on the avian W chromosome, consistent with gradual DNA loss on a nonrecombining chromosome.     

Positive selection for indel substitutions in the rodent sperm protein catsper1

Catsper1 is a voltage-gated calcium channel located in the plasma membrane of the sperm tail and is necessary for sperm motility and fertility in mice. We here examine the evolutionary pattern of Catsper1 from nine species of the rodent subfamily Murinae of family Muridae. We show that the rate of insertion/deletion (indel) substitutions in exon 1 of the gene is 4-15 times that in introns or neutral genomic regions, suggesting the presence of strong positive selection that promotes fixations of indel mutations in exon 1. The number of indel polymorphisms within species appears higher than expected from interspecific comparisons, although there are too little data to provide a statistically significant conclusion. These results, together with an earlier report in primates, indicate that positive selection promoting length variation in Catsper1 may be widespread in mammals. A structural model of Catsper1 suggested the importance of the exon 1-encoded region in regulating channel inactivation, which may affect sperm mobility and sperm competition. Our findings provide a necessary foundation for future experimental investigations of Catsper1's function in sperm physiology and role in sperm competition using rodent models.     

基因组水平的插入和缺失变异研究进展

插入和缺失(insertion and deletion)是DNA和蛋白质在进化过程中发生的序列长度上的改变,由于缺乏祖先序列的信息,不能肯定其到底是插入事件还是缺失事件,故统称之为增减(indel)。indel是分子水平进化变异的主要来源之一,近年来对这种进化事件的研究已经涵盖了其发生频率、大小、分布模式、序列进化模型及应用等各个方面。该文总结了基因组水平上插入和缺失的研究进展和发生机制;介绍了已经提出的插入和缺失进化模型,包括TKF91、TKF92、"Long Indel"模型和序列环境模型;讨论了插入和缺失作为分子标记在分子进化、基因分型和药物设计等方面的应用。     

Interspecific evolution in plant microsatellite structure

Several intragenically linked microsatellites have been identified in the floral regulatory genes A. sandwicense APETALA1 (ASAP1) and A. sandwicense APETALA3/TM6 (ASAP3/TM6) in 17 species of the Hawaiian and North American Madiinae (Asteraceae). Thirty-nine microsatellite loci were observed in the introns of these two genes, suggesting that they are hotspots for microsatellite formation. The sequences of four of these microsatellites were mapped onto the phylogenies of these floral regulatory genes, and the structural evolution of these repeat loci was traced. Both nucleotide substitutions and insertion/deletion mutations may be responsible for the formation of perfect microsatellites from imperfect repeat regions (and vice versa).     

SHORT INDELS ARE SUBJECT TO INSERTION‐BIASED GENE CONVERSION

Recombination between homologous loci is accompanied by formation of heteroduplexes. Repairing mismatches in heteroduplexes often leads to single nucleotide substitutions in a process known as gene conversion. Gene conversion was shown to be GC‐biased in different organisms; that is, a W(A or T)→S(G or C) substitution is more likely in this process than a S→W substitution. Here, we show that the insertion/deletion ratio for short noncoding indels that reach fixation between species is positively correlated with the recombination rate in Drosophila melanogaster, Homo sapiens, and Saccharomyces cerevisiae. This correlation is both due to an increase of the fixation rate of insertions and decrease of the fixation rate of deletions in regions of high recombination. Whole‐genome data on indel polymorphism and divergence in D. melanogaster rule out mutation biases and selection as the cause of this trend, pointing to insertion‐biased gene conversion as the most likely explanation. The bias toward insertions is the strongest for single‐nucleotide indels, and decreases with indel length. In regions of high recombination rate this bias leads to an up to ～5‐fold excess of fixed short insertions over deletions, and substantially affects the evolution of DNA segments.     

The genomic landscape of short insertion and deletion polymorphisms in the chicken (Gallus gallus) Genome: a high frequency of deletions in tandem duplicates

It is increasingly recognized that insertions and deletions (indels) are an important source of genetic as well as phenotypic divergence and diversity. We analyzed length polymorphisms identified through partial (0.25x) shotgun sequencing of three breeds of domestic chicken made by the International Chicken Polymorphism Map Consortium. A data set of 140,484 short indel polymorphisms in unique DNA was identified after filtering for microsatellite structures. There was a significant excess of tandem duplicates at indel sites, with deletions of a duplicate motif outnumbering the generation of duplicates through insertion. Indel density was lower in microchromosomes than in macrochromosomes, in the Z chromosome than in autosomes, and in 100 bp of upstream sequence, 5'-UTR, and first introns than in intergenic DNA and in other introns. Indel density was highly correlated with single nucleotide polymorphism (SNP) density. The mean density of indels in pairwise sequence comparisons was 1.9 x 10(-4) indel events/bp, approximately 5% the density of SNPs segregating in the chicken genome. The great majority of indels involved a limited number of nucleotides (median 1 bp), with A-rich motifs being overrepresented at indel sites. The overrepresentation of deletions at tandem duplicates indicates that replication slippage in duplicate sequences is a common mechanism behind indel mutation. The correlation between indel and SNP density indicates common effects of mutation and/or selection on the occurrence of indels and point mutations.     

An analysis of substitution, deletion and insertion mutations in cancer genes

Cancer-associated mutations in cancer genes constitute a diverse set of mutations associated with the disease. To gain insight into features of the set, substitution, deletion and insertion mutations were analysed at the nucleotide level, from the COSMIC database. The most frequent substitutions were c → t, g → a, g → t, and the most frequent codon changes were to termination codons. Deletions more than insertions, FS (frameshift) indels more than I-F (in-frame) ones, and single-nucleotide indels, were frequent. FS indels cause loss of significant fractions of proteins. The 5'-cut in FS deletions, and 5'-ligation in FS insertions, often occur between pairs of identical bases. Interestingly, the cut-site and 3'-ligation in insertions, and 3'-cut and join-pair in deletions, were each found to be the same significantly often (p < 0.001). It is suggested that these features aid the incorporation of indel mutations. Tumor suppressors undergo larger numbers of mutations, especially disruptive ones, over the entire protein length, to inactivate two alleles. Proto-oncogenes undergo fewer, less-disruptive mutations, in selected protein regions, to activate a single allele. Finally, catalogues, in ranked order, of genes mutated in each cancer, and cancers in which each gene is mutated, were created. The study highlights the nucleotide level preferences and disruptive nature of cancer mutations.     

Molecular evolution and phylogenetic utility of the petD group II intron: a case study in basal angiosperms

Sequences of spacers and group I introns in plant chloroplast genomes have recently been shown to be very effective in phylogenetic reconstruction at higher taxonomic levels and not only for inferring relationships among species. Group II introns, being more frequent in those genomes than group I introns, may be further promising markers. Because group II introns are structurally constrained, we assumed that sequences of a group II intron should be alignable across seed plants. We designed universal amplification primers for the petD intron and sequenced this intron in a representative selection of 47 angiosperms and three gymnosperms. Our sampling of taxa is the most representative of major seed plant lineages to date for group II introns. Through differential analysis of structural partitions, we studied patterns of molecular evolution and their contribution to phylogenetic signal. Nonpairing stretches (loops, bulges, and interhelical nucleotides) were considerably more variable in both substitutions and indels than in helical elements. Differences among the domains are basically a function of their structural composition. After the exclusion of four mutational hotspots accounting for less than 18% of sequence length, which are located in loops of domains I and IV, all sequences could be aligned unambiguously across seed plants. Microstructural changes predominantly occurred in loop regions and are mostly simple sequence repeats. An indel matrix comprising 241 characters revealed microstructural changes to be of lower homoplasy than are substitutions. In showing Amborella first branching and providing support for a magnoliid clade through a synapomorphic indel, the petD data set proved effective in testing between alternative hypotheses on the basal nodes of the angiosperm tree. Within angiosperms, group II introns offer phylogenetic signal that is intermediate in information content between that of spacers and group I introns on the one hand and coding sequences on the other.     

Comparative analysis of the structural organization of two closely related vitellogenin genes in X. laevis

The structural organization of the two closely related vitellogenin genes A1 and A2 has been determined and compared by electron microscopy. In both genes the mRNA-coding sequence of 6 kb is interrupted 33 times, leading to a total gene length of 21 kb for gene A1 and 16 kb for gene A2. Thus both genes have a mean exon length of 0.175 kb, while the mean intron length is 0.45 kb in gene A1 and 0.31 kb in gene A2. Because the introns interrupt the structural sequence at homologous positions in genes A1 and A2, we suggest that these two genes are the products of a duplication of an ancestral gene which had an intron-exon arrangement similar to that of the extant genes. Since the duplication event, the sequence and length of the analogous introns have changed rapidly, whereas homologous exons have diverged to an extent of only 5% of their sequences. The results suggest different mechanisms of evolution for exons and introns. While the exons evolved primarily by point mutations, such mutations, as well as deletion, insertion and duplication events, were important in the evolution of the introns.     

Insertions, substitutions, and the origin of microsatellites

This paper uses data from the Human Gene Mutation Database to contrast two hypotheses for the origin of short DNA repeats: substitutions and insertions that duplicate adjacent sequences. Because substitutions are much more common than insertions, they are the dominant source of new 2-repeat loci. Insertions are rarer, but over 70% of the 2-4 base insertion mutations are duplications of adjacent sequences, and over half of these generate new repeat regions. Insertions contribute fewer new repeat loci than substitutions, but their relative importance increases rapidly with repeat number so that all new 4-5-repeat mutations come from insertions, as do all 3-repeat mutations of tetranucleotide repeats. This suggests that the process of repeat duplication that dominates microsatellite evolution at high repeat numbers is also important very early in microsatellite evolution. This result sheds light on the puzzle of the origin of short tandem repeats. It also suggests that most short insertion mutations derive from a slippage-like process during replication.     

Evolution of the mitochondrial rps3 intron in perennial and annual angiosperms and homology to nad5 intron 1.

The plant mitochondrial rps3 intron was analyzed for substitution and indel rate variation among 15 monocot and dicot angiosperms from 10 genera, including perennial and annual taxa. Overall, the intron sequence was very conserved among angiosperms. Based on length polymorphism, 10 different alleles were identified among the 10 genera. These allelic differences were mainly attributable to large indels. An insertion of 133 nucleotides, observed in the Alnus intron was partially or completely absent in the other lineages of the family Betulaceae. This insertion was located within domain IV of the secondary-structure model of this group IIA intron. A mobile element of 47 nucleotides that showed homology to sequences located in rice rps3 intron and in intergenic plant mitochondrial genomes was found within this insertion. Both substitution and indel rates were low among the Betulaceae sequences, but substitution rates were increasingly larger than indel rates in comparisons involving more distantly related taxa. From a secondary-structure model, regions involved in helical structures were shown to be well preserved from indels as compared to substitutions, but compensatory changes were not observed among the angiosperm sequences analyzed. Using approximate divergence times based on the fossil record, substitution and indel rate heterogeneity was observed between different pairs of annual and perennial taxa. In particular, the annual petunia and primrose evolved more than 15 and 10 times faster, for substitution and indel rates respectively, than the perennial birch and alder. This is the first demonstration of an evolutionary rate difference between perennial and annual forms in noncoding DNA, lending support to neutral causes such as the generation time, population size, and speciation rate effects to explain such rate heterogeneity. Surprisingly, the sequence from the rps3 intron had a high identity with the sequence of intron 1 from the angiosperm mitochondrial nad5 gene, suggesting a common origin of these two group IIA introns.     

The pattern of insertion/deletion polymorphism in Arabidopsis thaliana

Little is known about variation of nucleotide insertion/deletions (indels) within species. In Arabidopsis thaliana, we investigated indel polymorphism patterns between two genome sequences and among 96 accessions at 1215 loci. Our study identified patterns in the variation of indel density, size, GC content and distribution, and a correlation between indels and substitutions. We found that the GC content in indel sequences was lower than that in non-indel sequences and that indels typically occur in regions with lower GC content. Patterns of indel frequency distribution among populations were more consistent with neutral expectation than substitution patterns. We also found that the local level of substitutions is positively correlated with indel density and negatively correlated with their distance to the closed indel, suggesting that indels play an important role in nucleotide variation.     

Simulating DNA coding sequence evolution with EvolveAGene 3

Phylogenetic reconstruction based upon multiple alignments ofmolecular sequences is important to most branches of modernbiology and is central to molecular evolution. Understandingthe historical relationships among macromolecules depends uponcomputer programs that implement a variety of analytical methods.Because it is impossible to know those historical relationshipswith certainty, assessment of the accuracy of methods and theprograms that implement them requires the use of programs thatrealistically simulate the evolution of DNA sequences. EvolveAGene3 is a realistic coding sequence simulation program that separatesmutation from selection and allows the user to set selectionconditions, including variable regions of selection intensitywithin the sequence and variation in intensity of selectionover branches. Variation includes base substitutions, insertions,and deletions. To the best of my knowledge, it is the only programavailable that simulates the evolution of intact coding sequences.Output includes the true tree and true alignments of the resultingcoding sequence and corresponding protein sequences. A log filereports the frequencies of each kind of base substitution, theratio of transition to transversion substitutions, the ratioof indel to base substitution mutations, and the numbers ofsilent and amino acid replacement mutations. The realism ofthe data sets has been assessed by comparing the d_N/d_S ratio,the ratio of transition to transversion substitutions, and theratio of indel to base substitution mutations of the simulateddata sets with those parameters of real data sets from the "goldstandard" BaliBase collection of structural alignments. Resultsshow that the data sets produced by EvolveAGene 3 are very similarto real data sets, and EvolveAGene 3 is therefore a realisticsimulation program that can be used to evaluate a variety ofprograms and methods in molecular evolution.     

Bovine spongiform encephalopathy associated insertion/deletion polymorphisms of the prion protein gene in the four beef cattle breeds from North China

Two insertion/deletion (indel) polymorphisms of the prion protein gene (PRNP), a 23-bp indel in the putative promoter region and a 12-bp indel within intron I, are associated with the susceptibility to bovine spongiform encephalopathy (BSE) in cattle. In the present study, the polymorphism frequencies of the two indels in four main beef cattle breeds (Hereford, Simmental, Black Angus, and Mongolian) from North China were studied. The results showed that the frequencies of deletion genotypes and alleles of 23- and 12-bp indels were lower, whereas the frequencies of insertion genotypes and alleles of the two indels were higher in Mongolian cattle than in the other three cattle breeds. In Mongolian cattle, the 23-bp insertion / 12-bp insertion was the major haplotype, whereas in Hereford, Simmental, and Black Angus cattle, the 23-bp deletion / 12-bp deletion was the major haplotype. These results demonstrated that Mongolian cattle could be more resistant to BSE, compared with the other three cattle breeds, because of its relatively low frequencies of deletion genotypes and alleles of 23- and 12-bp indel polymorphisms. Thus, this race could be important for selective breeding to improve resistance against BSE in this area.