Concerted Evolution Research Articles

Genetic homogenization of the nuclear ITS loci across two morphologically distinct gentians in their overlapping distributions in the Qinghai-Tibet Plateau.

Interspecific hybridization and introgression can lead to partial genetic homogenization at certain neutral loci between morphologically distinct species and may obscure the species delimitation using nuclear genes. In this study, we investigated this phenomenon through population genetic survey of two alpine plants (Gentiana siphonantha and G. straminea) in the Qinghai-Tibet Plateau, where the distributions of two species are partly overlapped. We identified two clusters of chloroplast DNA haplotypes which correspond to the two species, and three clusters of ITS ribotypes. In addition to clusters specific to each species, the third ITS cluster, which was most likely derived from hybridization between the other two clusters and subsequent recombination and concerted evolution, was widely shared by two species in their adjacent areas. In contrast to the morphological distinctiveness of the two species, interspecific gene flow possibly led to genetic homogenization at their ITS loci. The new ITS lineage recovered for species in adjacent areas is distinctly different from original lineages found in allopatric areas. These findings may have general implications for our understanding of cryptic changes at some genetic loci caused by interspecific gene flow in the history, and they indicate that species delimitation should be based on a combination of both nuclear and chloroplast DNA sequence variations.

Barcoding lichen-forming fungi using 454 pyrosequencing is challenged by artifactual and biological sequence variation.

Although lichens (lichen-forming fungi) play an important role in the ecological integrity of many vulnerable landscapes, only a minority of lichen-forming fungi have been barcoded out of the currently accepted ∼18 000 species. Regular Sanger sequencing can be problematic when analyzing lichens since saprophytic, endophytic, and parasitic fungi live intimately admixed, resulting in low-quality sequencing reads. Here, high-throughput, long-read 454 pyrosequencing in a GS FLX+ System was tested to barcode the fungal partner of 100 epiphytic lichen species from Switzerland using fungal-specific primers when amplifying the full internal transcribed spacer region (ITS). The present study shows the potential of DNA barcoding using pyrosequencing, in that the expected lichen fungus was successfully sequenced for all samples except one. Alignment solutions such as BLAST were found to be largely adequate for the generated long reads. In addition, the NCBI nucleotide database-currently the most complete database for lichen-forming fungi-can be used as a reference database when identifying common species, since the majority of analyzed lichens were identified correctly to the species or at least to the genus level. However, several issues were encountered, including a high sequencing error rate, multiple ITS versions in a genome (incomplete concerted evolution), and in some samples the presence of mixed lichen-forming fungi (possible lichen chimeras).

Novel duplication pattern of the mitochondrial control region in Cantor's Giant softshell turtle Pelochelys cantorii

Cantor's Giant Softshell Turtle, Pelochelys cantorii has become one of the most critically endangered species in the world. When comparative analyses of the P. cantorii complete mitochondrial genome sequences were conducted, we discovered a duplication of a segment of the control region in the mitochondrial genome of P. cantorii. The duplication is characterized by two copies of conserved sequence box 2 (CSB2) and CSB3 in a single control region. In contrast to previous reports of duplications involving the control regions of other animals, this particular pattern of duplications appears to be unique to P. cantorii. Copies of the CSB2 and CSB3 show many of the conserved sequence features typically found in mitochondrial control regions, and rare differences were found between the paralogous copies. Using the primer design principle of simple sequence repeats (SSR) and the reference sequence of the duplicated CSBs, specific primers were designed to amplify the duplicated CSBs. These primers were validated among different individuals and populations of P. cantorii. This unique duplication structure suggests the two copies of the CSB2 and CSB3 may have arisen through occasional tandem duplication and subsequent concerted evolution.

Orthologs, turn-over, and remolding of tRNAs in primates and fruit flies.

BackgroundTransfer RNAs (tRNAs) are ubiquitous in all living organism. They implement the genetic code so that most genomes contain distinct tRNAs for almost all 61 codons. They behave similar to mobile elements and proliferate in genomes spawning both local and non-local copies. Most tRNA families are therefore typically present as multicopy genes. The members of the individual tRNA families evolve under concerted or rapid birth-death evolution, so that paralogous copies maintain almost identical sequences over long evolutionary time-scales. To a good approximation these are functionally equivalent. Individual tRNA copies thus are evolutionary unstable and easily turn into pseudogenes and disappear. This leads to a rapid turnover of tRNAs and often large differences in the tRNA complements of closely related species. Since tRNA paralogs are not distinguished by sequence, common methods cannot not be used to establish orthology between tRNA genes.ResultsIn this contribution we introduce a general framework to distinguish orthologs and paralogs in gene families that are subject to concerted evolution. It is based on the use of uniquely aligned adjacent sequence elements as anchors to establish syntenic conservation of sequence intervals. In practice, anchors and intervals can be extracted from genome-wide multiple sequence alignments. Syntenic clusters of concertedly evolving genes of different families can then be subdivided by list alignments, leading to usually small clusters of candidate co-orthologs. On the basis of recent advances in phylogenetic combinatorics, these candidate clusters can be further processed by cograph editing to recover their duplication histories. We developed a workflow that can be conceptualized as stepwise refinement of a graph of homologous genes. We apply this analysis strategy with different types of synteny anchors to investigate the evolution of tRNAs in primates and fruit flies. We identified a large number of tRNA remolding events concentrated at the tips of the phylogeny. With one notable exception all phylogenetically old tRNA remoldings do not change the isoacceptor class.ConclusionsGene families evolving under concerted evolution are not amenable to classical phylogenetic analyses since paralogs maintain identical, species-specific sequences, precluding the estimation of correct gene trees from sequence differences. This leaves conservation of syntenic arrangements with respect to “anchor elements” that are not subject to concerted evolution as the only viable source of phylogenetic information. We have demonstrated here that a purely synteny-based analysis of tRNA gene histories is indeed feasible. Although the choice of synteny anchors influences the resolution in particular when tight gene clusters are present, and the quality of sequence alignments, genome assemblies, and genome rearrangements limits the scope of the analysis, largely coherent results can be obtained for tRNAs. In particular, we conclude that a large fraction of the tRNAs are recent copies. This proliferation is compensated by rapid pseudogenization as exemplified by many very recent alloacceptor remoldings.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-016-2927-4) contains supplementary material, which is available to authorized users.

Contrasting Patterns of rDNA Homogenization within the Zygosaccharomyces rouxii Species Complex.

Arrays of repetitive ribosomal DNA (rDNA) sequences are generally expected to evolve as a coherent family, where repeats within such a family are more similar to each other than to orthologs in related species. The continuous homogenization of repeats within individual genomes is a recombination process termed concerted evolution. Here, we investigated the extent and the direction of concerted evolution in 43 yeast strains of the Zygosaccharomyces rouxii species complex (Z. rouxii, Z. sapae, Z. mellis), by analyzing two portions of the 35S rDNA cistron, namely the D1/D2 domains at the 5’ end of the 26S rRNA gene and the segment including the internal transcribed spacers (ITS) 1 and 2 (ITS regions). We demonstrate that intra-genomic rDNA sequence variation is unusually frequent in this clade and that rDNA arrays in single genomes consist of an intermixing of Z. rouxii, Z. sapae and Z. mellis-like sequences, putatively evolved by reticulate evolutionary events that involved repeated hybridization between lineages. The levels and distribution of sequence polymorphisms vary across rDNA repeats in different individuals, reflecting four patterns of rDNA evolution: I) rDNA repeats that are homogeneous within a genome but are chimeras derived from two parental lineages via recombination: Z. rouxii in the ITS region and Z. sapae in the D1/D2 region; II) intra-genomic rDNA repeats that retain polymorphisms only in ITS regions; III) rDNA repeats that vary only in their D1/D2 domains; IV) heterogeneous rDNA arrays that have both polymorphic ITS and D1/D2 regions. We argue that an ongoing process of homogenization following allodiplodization or incomplete lineage sorting gave rise to divergent evolutionary trajectories in different strains, depending upon temporal, structural and functional constraints. We discuss the consequences of these findings for Zygosaccharomyces species delineation and, more in general, for yeast barcoding.

Sequence diversity and positive selection at the Duffy-binding protein genes of Plasmodium knowlesi and P. cynomolgi: Analysis of the complete coding sequences of Thai isolates

Plasmodium knowlesi and P. cynomolgi are simian malaria parasites capable of causing symptomatic human infections. The interaction between the Duffy binding protein alpha on P. knowlesi merozoite and the Duffy-antigen receptor for chemokine (DARC) on human and macaque erythrocyte membrane is prerequisite for establishment of blood stage infection whereas DARC is not required for erythrocyte invasion by P. cynomolgi. To gain insights into the evolution of the PkDBP gene family comprising PkDBPα, PkDBPβ and PkDBPγ, and a member of the DBP gene family of P. cynomolgi (PcyDBP1), the complete coding sequences of these genes were analyzed from Thai field isolates and compared with the publicly available DBP sequences of P. vivax (PvDBP). The complete coding sequences of PkDBPα (n=11), PkDBPβ (n=11), PkDBPγ (n=10) and PcyDBP1 (n=11) were obtained from direct sequencing of the PCR products. Nucleotide diversity of DBP is highly variable across malaria species. PcyDBP1 displayed the greatest level of nucleotide diversity while all PkDBP gene members exhibited comparable levels of diversity. Positive selection occurred in domains I, II and IV of PvDBP and in domain V of PcyDBP1. Although deviation from neutrality was not detected in domain II of PkDBPα, a signature of positive selection was identified in the putative DARC binding site in this domain. The DBP gene families seem to have arisen following the model of concerted evolution because paralogs rather than orthologs are clustered in the phylogenetic tree. The presence of identical or closely related repeats exclusive for the PkDBP gene family suggests that duplication of gene members postdated their divergence from the ancestral PcyDBP and PvDBP lineages. Intragenic recombination was detected in all DBP genes of these malaria species. Despite the limited number of isolates, P. knowlesi from Thailand shared phylogenetically related domain II sequences of both PkDBPα and PkDBPγ with those from Peninsular Malaysia, consistent with their geographic proximity.

Polyandry, Predation, and the Evolution of Frog Reproductive Modes.

Frog reproductive modes are complex phenotypes that include egg/clutch characteristics, oviposition site, larval development, and sometimes, parental care. Two evident patterns in the evolution of these traits are the higher diversity of reproductive modes in the tropics and the apparent progression from aquatic to terrestrial reproduction, often attributed to higher fitness resulting from decreased predation on terrestrial eggs and tadpoles. Here, we propose that sexual selection-and not only natural selection due to predation-favors terrestrial breeding by reducing the loss of fitness due to polyandry. To examine this novel selective mechanism, we reconstructed the evolution of reproductive diversity in two frog families (Hylidae and Leptodactylidae) and tested for concerted evolution of egg and tadpole development sites with specific mating behaviors. We found that oviposition and tadpole development sites are evolving independently, do not show the same diversity and/or directionality in terms of terrestriality, and thus may be diversifying due to different selective mechanisms. In both families, terrestrial egg deposition is correlated with amplexus that is hidden from competing males, and in hylids, testes mass was significantly larger and more variable in males with exposed amplexus that are vulnerable to polyandry. Our results indicate that intrasexual selection has been an underappreciated mechanism promoting diversification of frog reproductive modes.

Intragenomic polymorphism and intergenomic recombination in the ribosomal RNA genes of strains belonging to a yeast species Pichia membranifaciens

ABSTRACTA concerted evolution model has been proposed to explain the observed lack of sequence variation among the multiple ribosomal RNA (rRNA) gene copies in many different eukaryotic species. Recent studies on the level of intragenomic variations in the rRNA gene repeats of fungi resulted in controversial conclusions. In this study, we clearly showed that significant polymorphisms of the internal transcribed spacers (ITS1 and ITS2) of ribosomal DNA (rDNA) exist within the genome of a strain of the yeast species Pichia membranifaciens. More interestingly, we showed that the intragenomic ITS sequence polymorphisms were formed by intergenomic rDNA recombination among different P. membranifaciens strains with significantly different ITS sequences. Intergenomic rDNA recombination was also responsible for the diversification of rDNA sequences in different strains of the species. After the events bring together different rDNA types in individual genomes of the P. membranifaciens strains compared, rDNA sequence heterogeneity has remained in the genome of one but eliminated by homogenisation in the genomes of other strains. Our findings show new clue for further investigation on the mechanism of concerted evolution of rRNA genes in eukaryotes.

Extremophile Poeciliidae: multivariate insights into the complexity of speciation along replicated ecological gradients.

BackgroundReplicate population pairs that diverge in response to similar selective regimes allow for an investigation of (a) whether phenotypic traits diverge in a similar and predictable fashion, (b) whether there is gradual variation in phenotypic divergence reflecting variation in the strength of natural selection among populations, (c) whether the extent of this divergence is correlated between multiple character suites (i.e., concerted evolution), and (d) whether gradual variation in phenotypic divergence predicts the degree of reproductive isolation, pointing towards a role for adaptation as a driver of (ecological) speciation. Here, we use poeciliid fishes of the genera Gambusia and Poecilia that have repeatedly evolved extremophile lineages able to tolerate high and sustained levels of toxic hydrogen sulfide (H2S) to answer these questions.ResultsWe investigated evolutionary divergence in response to H2S in Gambusia spp. (and to a lesser extent Poecilia spp.) using a multivariate approach considering the interplay of life history, body shape, and population genetics (nuclear miscrosatellites to infer population genetic differentiation as a proxy for reproductive isolation). We uncovered both shared and unique patterns of evolution: most extremophile Gambusia predictably evolved larger heads and offspring size, matching a priori predictions for adaptation to sulfidic waters, while variation in adult life histories was idiosyncratic. When investigating patterns for both genera (Gambusia and Poecilia), we found that divergence in offspring-related life histories and body shape were positively correlated across populations, but evidence for individual-level associations between the two character suites was limited, suggesting that genetic linkage, developmental interdependencies, or pleiotropic effects do not explain patterns of concerted evolution. We further found that phenotypic divergence was positively correlated with both environmental H2S-concentration and neutral genetic differentiation (a proxy for gene flow).ConclusionsOur results suggest that higher toxicity exerts stronger selection, and that divergent selection appears to constrain gene flow, supporting a scenario of ecological speciation. Nonetheless, progress toward ecological speciation was variable, partially reflecting variation in the strength of divergent selection, highlighting the complexity of selective regimes even in natural systems that are seemingly governed by a single, strong selective agent.Electronic supplementary materialThe online version of this article (doi:10.1186/s12862-016-0705-1) contains supplementary material, which is available to authorized users.

Prevalence and Dynamics of Ribosomal DNA Micro-heterogeneity Are Linked to Population History in Two Contrasting Yeast Species.

Despite the considerable number and taxonomic breadth of past and current genome sequencing projects, many of which necessarily encompass the ribosomal DNA, detailed information on the prevalence and evolutionary significance of sequence variation in this ubiquitous genomic region are severely lacking. Here, we attempt to address this issue in two closely related yet contrasting yeast species, the baker’s yeast Saccharomyces cerevisiae and the wild yeast Saccharomyces paradoxus. By drawing on existing datasets from the Saccharomyces Genome Resequencing Project, we identify a rich seam of ribosomal DNA sequence variation, characterising 1,068 and 970 polymorphisms in 34 S. cerevisiae and 26 S. paradoxus strains respectively. We discover the two species sets exhibit distinct mutational profiles. Furthermore, we show for the first time that unresolved rDNA sequence variation resulting from imperfect concerted evolution of the ribosomal DNA region follows a U-shaped allele frequency distribution in each species, similar to loci that evolve under non-concerted mechanisms but arising through rather different evolutionary processes. Finally, we link differences between the shapes of these allele frequency distributions to the two species’ contrasting population histories.

Concerted evolution in the mitochondrial control region of the Amazon small-bodied frog Pseudopaludicola canga (Anura, Leiuperidae)

This study presents evidence of concerted evolution in the mitochondrial control region of the frog Pseudopaludicola canga. Four repeat units of 88 bp (as well as a fifth, incomplete unit) were observed in the 5′ domain, with the duplicated segments of the same specimen being more related to one another than to the equivalent regions in other specimens, as a result of concerted evolution. We highlight that drawing conclusions from phylogeographical analysis using the control region containing VNTRs must be interpreted with caution, because it violated a basic assumption of phylogeny, since the regions cannot be treated as independent characters.

Zinc Finger Domain of the PRDM9 Gene on Chromosome 1 Exhibits High Diversity in Ruminants but Its Paralog PRDM7 Contains Multiple Disruptive Mutations.

PRDM9 is the sole hybrid sterility gene identified so far in vertebrates. PRDM9 gene encodes a protein with an immensely variable zinc-finger (ZF) domain that determines the site of meiotic recombination hotspots genome-wide. In this study, the terminal ZF domain of PRDM9 on bovine chromosome 1 and its paralog on chromosome 22 were characterized in 225 samples from five ruminant species (cattle, yak, mithun, sheep and goat). We found extraordinary variation in the number of PRDM9 zinc fingers (6 to 12). We sequenced PRDM9 ZF encoding region from 15 individuals (carrying the same ZF number in both copies) and found 43 different ZF domain sequences. Ruminant zinc fingers of PRDM9 were found to be diversifying under positive selection and concerted evolution, specifically at positions involved in defining their DNA-binding specificity, consistent with the reports from other vertebrates such as mice, humans, equids and chimpanzees. ZF-encoding regions of the PRDM7, a paralog of PRDM9 on bovine chromosome 22 and on unknown chromosomes in other studied species were found to contain 84 base repeat units as in PRDM9, but there were multiple disruptive mutations after the first repeat unit. The diversity of the ZFs suggests that PRDM9 may activate recombination hotspots that are largely unique to each ruminant species.

Non-concerted evolution in ribosomal ITS2 sequence in Cynoglossus zanzibarensis (Pleuronectiformes: Cynoglossidae)

The nuclear ribosomal RNA genes (nrDNA) include three rRNA genes (18S, 5.8S, and 28S) and two internal transcribed spacers (ITS1 and ITS2). The lack of variability in nrDNA has been explained as the result of concerted evolution. In fact, many examples of variation have been discovered, suggesting a non-concerted evolution process. There are few studies on ITS2 in teleostean fishes; particularly no reports on flatfishes. In this study, we focused on ITS2 in Zanzibar tonguesole, Cynoglossus zanzibarensis (Pleuronectiformes: Cynoglossidae). Three types of ITS2 sequences (Type A, B, and C) coexisted and mainly differ in length, nucleotide diversity (π), secondary structure, and minimum free energy. Based on these different features, Type B and C were speculated as the putative pseudogenic ITS2 region. Moreover, Type C sequences were detected as recombinant from crossing over between Type A and B sequences. To our knowledge, this is the first report on ITS2 recombination in animal ribosomal RNA genes. The high degree of ITS2 sequence polymorphism at the intraindividual level supported a non-concerted evolution pattern in the C. zanzibarensis genome. These results provide useful data for further studies on ITS2 in flatfishes and contribute to polymorphism analyses of ribosomal RNA genes in teleostean fishes.

A molecular phylogeny of rose chafers (Coleoptera: Scarabaeidae: Cetoniinae) reveals a complex and concerted morphological evolution related to their flight mode

Rose chafers (Cetoniinae) are a large group of flower visitors within the pleurostict Scarabaeidae that are characterized by their distinctive flight mode with nearly closed forewings. Despite their popularity, this is the first study to use molecular data to infer their phylogenetic relationships. We used partial gene sequences for 28S rRNA, cytochrome oxidase I (cox1) and 16S rRNA (rrnL) for 299 species, representing most recognized subfamilies of Scarabaeidae, including 125 species of Cetoniinae. Combined analyses using maximum parsimony, maximum likelihood and Bayesian inferences recovered Cetoniinae as monophyletic in all analyses, with the sister clade composed of Rutelinae and Dynastinae. Rutelinae was always recovered as paraphyletic with respect to Dynastinae. Trichiini sensu lato (s.l.) was recovered as a polyphyletic clade, while Cetoniini s.l. was recovered as paraphyletic. The inferred topologies were also supported by site bootstrapping of the ML trees. With the exception of Cremastochelini, most tribes of Cetoniinae were poly- or paraphyletic, indicating the critical need for a careful revision of rose chafer classification. Analysis of elytral base structure (including 11 scored characters) in the context of phylogeny, revealed a complex, concerted and rapid transformation of the single trait elements linked to a modified flight mode with closed elytra. This appears to be unlinked to the lateral sinuation of the elytra, which originated independently several times at later stages in the evolution of the group.

Sperm Bindin Divergence under Sexual Selection and Concerted Evolution in Sea Stars.

Selection associated with competition among males or sexual conflict between mates can create positive selection for high rates of molecular evolution of gamete recognition genes and lead to reproductive isolation between species. We analyzed coding sequence and repetitive domain variation in the gene encoding the sperm acrosomal protein bindin in 13 diverse sea star species. We found that bindin has a conserved coding sequence domain structure in all 13 species, with several repeated motifs in a large central region that is similar among all sea stars in organization but highly divergent among genera in nucleotide and predicted amino acid sequence. More bindin codons and lineages showed positive selection for high relative rates of amino acid substitution in genera with gonochoric outcrossing adults (and greater expected strength of sexual selection) than in selfing hermaphrodites. That difference is consistent with the expectation that selfing (a highly derived mating system) may moderate the strength of sexual selection and limit the accumulation of bindin amino acid differences. The results implicate both positive selection on single codons and concerted evolution within the repetitive region in bindin divergence, and suggest that both single amino acid differences and repeat differences may affect sperm-egg binding and reproductive compatibility.

627. Concerted Evolution of Pseudogenes Facilitated Contiguous Deletion of the Canine Dystrophin and TMEM47 Genes by Homologous Recombination

Duchenne Muscular Dystrophy (DMD) is an X-linked recessive disorder caused by sporadic mutations in the 2.4 Mb dystrophin gene. The large size of the gene is responsible for one of the highest loss-of-function mutation rates in the entire human genome. The majority of dystrophin mutations that cause DMD are deletions that span multiple exons. As the resulting absence of central immunological tolerance has the documented potential to complicate gene therapies in which recombinant dystrophin is expressed, there is an urgent need for well characterized, deletional null animal models for DMD. Preliminary characterization of a naturally occurring mutation in a German shorthaired Pointer (GSHPMD) suggested that the canine dystrophin gene was disrupted by a single, large deletion. Here, we further characterize the GSHPMD deletion and determine the breakpoints to be within short, highly homologous DNA elements spaced over 5.6 Mb apart, suggesting homologous recombination as the most plausible mechanism. The deletion generously flanks the entire dystrophin gene, and additionally encompasses the TMEM47 gene. This is reminiscent of the famous B.B. deletion in the DMD patient whose DNA was used for subtractive hybridization in the cloning of the DMD and nearby CGD loci. Further investigation revealed that the homologous canine DNA elements are ferritin-like pseudogenes, and surprisingly that there are several additional copies present on the X-chromosome. Phylogenomic comparison of the physical map positions of the orthologous pseudogenes in proximity to the dystrophin gene shows a pattern of positional synteny, consistent with the expansion of this pseudogene family prior to the mammalian radiation. Unexpectedly, only in the canine lineage have the pseudogene sequences remained virtually identical to one another, suggesting the existence of a heretofore uncharacterized process of concerted molecular evolution spanning large physical distances. The fully characterized deletional null animal model represents an invaluable resource for the evaluation of the immunological consequences of recombinant dystrophin expression via gene therapy, as it provides the only mammalian system in which the complete absence of dystrophin prevents the development of central tolerance, as would be expected in most cases of DMD. Thus the GSHPMD model can be uniquely leveraged to address a major safety concern as the field pursues strategies for systemic gene delivery in DMD. Of note, published studies have revealed evidence for deleterious immunological responses following administration of vectors expressing recombinant HUMAN dystrophin in both DMD and the Golden Retriever canine model (NEJM 363: 1429-1437 and Mol. Ther. 18:1501-1508).

Sericin Composition in the Silk of Antheraea yamamai.

The silks produced by caterpillars consist of fibroin proteins that form two core filaments, and sericin proteins that seal filaments into a fiber and conglutinate fibers in the cocoon. Sericin genes are well-known in Bombyx mori (Bombycidae) but have received little attention in other insects. This paper shows that Antheraea yamamai (Saturniidae) contains five sericin genes very different from the three sericin genes of B. mori. In spite of differences, all known sericins are characterized by short exons 1 and 2 (out of 3-12 exons), expression in the middle silk gland section, presence of repeats with high contents of Ser and charged amino acid residues, and secretion as a sticky silk component soluble in hot water. The B. mori sericins represent tentative phylogenetic lineages (I) BmSer1 and orthologs in Saturniidae, (II) BmSer2, and (III) BmSer3 and related sericins of Saturniidae and of the pyralid Galleria mellonella. The lineage (IV) seems to be limited to Saturniidae. Concerted evolution of the sericin genes was apparently associated with gene amplifications as well as gene loses. Differences in the silk fiber morphology indicate that the cocktail of sericins linking the filaments and coating the fiber is modified during spinning. Silks are composite biomaterials of conserved function in spite of great diversity of their composition.

Human Xq28 inversion polymorphism: From sex linkage to Genomics--A genetic mother lode.

An inversion polymorphism of the filamin and emerin genes at the tip of the long arm of the human X-chromosome serves as the basis of an investigative laboratory in which students learn something new about their own genomes. Long, nearly identical inverted repeats flanking the filamin and emerin genes illustrate how repetitive elements can lead to alterations in genome structure (inversions) through nonallelic homologous recombination. The near identity of the inverted repeats is an example of concerted evolution through gene conversion. While the laboratory in its entirety is designed for college level genetics courses, portions of the laboratory are appropriate for courses at other levels. Because the polymorphism is on the X-chromosome, the laboratory can be used in introductory biology courses to enhance understanding of sex-linkage and to test for Hardy-Weinberg equilibrium in females. More advanced topics, such as chromosome interference, the molecular model for recombination, and inversion heterozygosity suppression of recombination can be explored in upper-level genetics and evolution courses. DNA isolation, restriction digests, ligation, long PCR, and iPCR provide experience with techniques in molecular biology. This investigative laboratory weaves together topics stretching from molecular genetics to cytogenetics and sex-linkage, population genetics and evolutionary genetics.

Contrasting evolutionary patterns of 28S and ITS rRNA genes reveal high intragenomic variation in Cephalenchus (Nematoda): Implications for species delimitation

Concerted evolution is often assumed to be the evolutionary force driving multi-family genes, including those from ribosomal DNA (rDNA) repeat, to complete homogenization within a species, although cases of non-concerted evolution have been also documented. In this study, sequence variation of 28S and ITS ribosomal RNA (rRNA) genes in the genus Cephalenchus is assessed at three different levels, intragenomic, intraspecific, and interspecific. The findings suggest that not all Cephalenchus species undergo concerted evolution. High levels of intraspecific polymorphism, mostly due to intragenomic variation, are found in Cephalenchus sp1 (BRA-01). Secondary structure analyses of both rRNA genes and across different species show a similar substitution pattern, including mostly compensatory (CBC) and semi-compensatory (SBC) base changes, thus suggesting the functionality of these rRNA copies despite the variation found in some species. This view is also supported by low sequence variation in the 5.8S gene in relation to the flanking ITS-1 and ITS-2 as well as by the existence of conserved motifs in the former gene. It is suggested that potential cross-fertilization in some Cephalenchus species, based on inspection of female reproductive system, might contribute to both intragenomic and intraspecific polymorphism of their rRNA genes. These results reinforce the potential implications of intragenomic and intraspecific genetic diversity on species delimitation, especially in biodiversity studies based solely on metagenetic approaches. Knowledge of sequence variation will be crucial for accurate species diversity estimation using molecular methods.

Evolutionary history of double-stranded RNA binding proteins in plants: identification of new cofactors involved in easiRNA biogenesis.

In this work, we retrace the evolutionary history of plant double-stranded RNA binding proteins (DRBs), a group of non-catalytic factors containing one or more double-stranded RNA binding motif (dsRBM) that play important roles in small RNA biogenesis and functions. Using a phylogenetic approach, we show that multiple dsRBM DRBs are systematically composed of two different types of dsRBMs evolving under different constraints and likely fulfilling complementary functions. In vascular plants, four distinct clades of multiple dsRBM DRBs are always present with the exception of Brassicaceae species, that do not possess member of the newly identified clade we named DRB6. We also identified a second new and highly conserved DRB family (we named DRB7) whose members possess a single dsRBM that shows concerted evolution with the most C-terminal dsRBM domain of the Dicer-like 4 (DCL4) proteins. Using a BiFC approach, we observed that Arabidopsis thaliana DRB7.2 (AtDRB7.2) can directly interact with AtDRB4 but not with AtDCL4 and we provide evidence that both AtDRB7.2 and AtDRB4 participate in the epigenetically activated siRNAs pathway.