Selective maintenance of Drosophila tandemly arranged duplicated genes during evolution

Background

The physical organization and chromosomal localization of genes within genomes is known to play an important role in their function. Most genes arise by duplication and move along the genome by random shuffling of DNA segments. Higher order structuring of the genome occurs in eukaryotes, where groups of physically linked genes are co-expressed. However, the contribution of gene duplication to gene order has not been analyzed in detail, as it is believed that co-expression due to recent duplicates would obscure other domains of co-expression.

Results

We have catalogued ordered duplicated genes in Drosophila melanogaster, and found that one in five of all genes is organized as tandem arrays. Furthermore, among arrays that have been spatially conserved over longer periods than would be expected on the basis of random shuffling, a disproportionate number contain genes encoding developmental regulators. Using in situ gene expression data for more than half of the Drosophila genome, we find that genes in these conserved clusters are co-expressed to a much higher extent than other duplicated genes.

Conclusions

These results reveal the existence of functional constraints in insects that retain copies of genes encoding developmental and regulatory proteins as neighbors, allowing their co-expression. This co-expression may be the result of shared cis-regulatory elements or a shared need for a specific chromatin structure. Our results highlight the association between genome architecture and the gene regulatory networks involved in the construction of the body plan.     

Singular value decomposition-based regression identifies activation of endogenous signaling pathways in vivo

Background

The physical organization and chromosomal localization of genes within genomes is known to play an important role in their function. Most genes arise by duplication and move along the genome by random shuffling of DNA segments. Higher order structuring of the genome occurs in eukaryotes, where groups of physically linked genes are co-expressed. However, the contribution of gene duplication to gene order has not been analyzed in detail, as it is believed that co-expression due to recent duplicates would obscure other domains of co-expression.

Results

We have catalogued ordered duplicated genes in Drosophila melanogaster, and found that one in five of all genes is organized as tandem arrays. Furthermore, among arrays that have been spatially conserved over longer periods than would be expected on the basis of random shuffling, a disproportionate number contain genes encoding developmental regulators. Using in situ gene expression data for more than half of the Drosophila genome, we find that genes in these conserved clusters are co-expressed to a much higher extent than other duplicated genes.

Conclusions

These results reveal the existence of functional constraints in insects that retain copies of genes encoding developmental and regulatory proteins as neighbors, allowing their co-expression. This co-expression may be the result of shared cis-regulatory elements or a shared need for a specific chromatin structure. Our results highlight the association between genome architecture and the gene regulatory networks involved in the construction of the body plan.     

Selective maintenance of Drosophila tandemly arranged duplicated genes during evolution

Background  

The physical organization and chromosomal localization of genes within genomes is known to play an important role in their function. Most genes arise by duplication and move along the genome by random shuffling of DNA segments. Higher order structuring of the genome occurs in eukaryotes, where groups of physically linked genes are co-expressed. However, the contribution of gene duplication to gene order has not been analyzed in detail, as it is believed that co-expression due to recent duplicates would obscure other domains of co-expression.     

Revealing biases inherent in recombination protocols

Background  

The recombination of homologous genes is an effective protein engineering tool to evolve proteins. DNA shuffling by gene fragmentation and reassembly has dominated the literature since its first publication, but this fragmentation-based method is labor intensive. Recently, a fragmentation-free PCR based protocol has been published, termed recombination-dependent PCR, which is easy to perform. However, a detailed comparison of both methods is still missing.     

Phylum-wide comparative genomics unravel the diversity of secondary metabolism in Cyanobacteria

Background

Cyanobacteria are an ancient lineage of photosynthetic bacteria from which hundreds of natural products have been described, including many notorious toxins but also potent natural products of interest to the pharmaceutical and biotechnological industries. Many of these compounds are the products of non-ribosomal peptide synthetase (NRPS) or polyketide synthase (PKS) pathways. However, current understanding of the diversification of these pathways is largely based on the chemical structure of the bioactive compounds, while the evolutionary forces driving their remarkable chemical diversity are poorly understood.

Results

We carried out a phylum-wide investigation of genetic diversification of the cyanobacterial NRPS and PKS pathways for the production of bioactive compounds. 452 NRPS and PKS gene clusters were identified from 89 cyanobacterial genomes, revealing a clear burst in late-branching lineages. Our genomic analysis further grouped the clusters into 286 highly diversified cluster families (CF) of pathways. Some CFs appeared vertically inherited, while others presented a more complex evolutionary history. Only a few horizontal gene transfers were evidenced amongst strongly conserved CFs in the phylum, while several others have undergone drastic gene shuffling events, which could result in the observed diversification of the pathways.

Conclusions

Therefore, in addition to toxin production, several NRPS and PKS gene clusters are devoted to important cellular processes of these bacteria such as nitrogen fixation and iron uptake. The majority of the biosynthetic clusters identified here have unknown end products, highlighting the power of genome mining for the discovery of new natural products.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-977) contains supplementary material, which is available to authorized users.     

Recombination of chl-fus gene (Plastid Origin) downstream of hop: a locus of chromosomal instability

Background

The co-chaperone Hop [heat shock protein (HSP) organizing protein] has been shown to act as an adaptor for protein folding and maturation, in concert with Hsp70 and Hsp90. The hop gene is of eukaryotic origin. Likewise, the chloroplast elongation factor G (cEF-G) catalyzes the translocation step in chloroplast protein synthesis. The chl-fus gene, which encodes the cEF-G protein, is of plastid origin. Both proteins, Hop and cEF-G, derived from domain duplications. It was demonstrated that the nuclear chl-fus gene locates in opposite orientation to a hop gene in Glycine max. We explored 53 available plant genomes from Chlorophyta to higher plants, to determine whether the chl-fus gene was transferred directly downstream of the primordial hop in the proto-eukaryote host cell. Since both genes came from exon/module duplication events, we wanted to explore the involvement of introns in the early origin and the ensuing evolutionary changes in gene structure.

Results

We reconstructed the evolutionary history of the two convergent plant genes, on the basis of their gene structure, microsynteny and microcolinearity, from 53 plant nuclear genomes. Despite a high degree (72 %) of microcolinearity among vascular plants, our results demonstrate that their adjacency was a product of chromosomal rearrangements. Based on predicted exon − intron structures, we inferred the molecular events giving rise to the current form of genes. Therefore, we propose a simple model of exon/module shuffling by intronic recombinations in which phase-0 introns were essential for domain duplication, and a phase-1 intron for transit peptide recruiting. Finally, we demonstrate a natural susceptibility of the intergenic region to recombine or delete, seriously threatening the integrity of the chl-fus gene for the future.

Conclusions

Our results are consistent with the interpretation that the chl-fus gene was transferred from the chloroplast to a chromosome different from that of hop, in the primitive photosynthetic eukaryote, and much later before the appearance of angiosperms, it was recombined downstream of hop. Exon/module shuffling mediated by symmetric intron phases (i.e., phase-0 introns) was essential for gene evolution. The intergenic region is prone to recombine, risking the integrity of both genes.

Electronic supplementary material

The online version of this article (doi:10.1186/s12864-015-1780-1) contains supplementary material, which is available to authorized users.     

Organization and evolution of transposable elements along the bread wheat chromosome 3B

Background

The 17 Gb bread wheat genome has massively expanded through the proliferation of transposable elements (TEs) and two recent rounds of polyploidization. The assembly of a 774 Mb reference sequence of wheat chromosome 3B provided us with the opportunity to explore the impact of TEs on the complex wheat genome structure and evolution at a resolution and scale not reached so far.

Results

We develop an automated workflow, CLARI-TE, for TE modeling in complex genomes. We delineate precisely 56,488 intact and 196,391 fragmented TEs along the 3B pseudomolecule, accounting for 85% of the sequence, and reconstruct 30,199 nested insertions. TEs have been mostly silent for the last one million years, and the 3B chromosome has been shaped by a succession of bursts that occurred between 1 to 3 million years ago. Accelerated TE elimination in the high-recombination distal regions is a driving force towards chromosome partitioning. CACTAs overrepresented in the high-recombination distal regions are significantly associated with recently duplicated genes. In addition, we identify 140 CACTA-mediated gene capture events with 17 genes potentially created by exon shuffling and show that 19 captured genes are transcribed and under selection pressure, suggesting the important role of CACTAs in the recent wheat adaptation.

Conclusion

Accurate TE modeling uncovers the dynamics of TEs in a highly complex and polyploid genome. It provides novel insights into chromosome partitioning and highlights the role of CACTA transposons in the high level of gene duplication in wheat.

Electronic supplementary material

The online version of this article (doi:10.1186/s13059-014-0546-4) contains supplementary material, which is available to authorized users.     

Sequence permutations in the molecular evolution of DNA methyltransferases

Background  

DNA methyltransferases (MTases), unlike MTases acting on other substrates, exhibit sequence permutation. Based on the sequential order of the cofactor-binding subdomain, the catalytic subdomain, and the target recognition domain (TRD), several classes of permutants have been proposed. The majority of known DNA MTases fall into the α, β, and γ classes. There is only one member of the ζ class known and no members of the δ and ε classes have been identified to date. Two mechanisms of permutation have been proposed: one involving gene duplication and in-frame fusion, and the other involving inter- and intragenic shuffling of gene segments.     

Evolution of GHF5 endoglucanase gene structure in plant-parasitic nematodes: no evidence for an early domain shuffling event

Background  

Endo-1,4-beta-glucanases or cellulases from the glycosyl hydrolase family 5 (GHF5) have been found in numerous bacteria and fungi, and recently also in higher eukaryotes, particularly in plant-parasitic nematodes (PPN). The origin of these genes has been attributed to horizontal gene transfer from bacteria, although there still is a lot of uncertainty about the origin and structure of the ancestral GHF5 PPN endoglucanase. It is not clear whether this ancestral endoglucanase consisted of the whole gene cassette, containing a catalytic domain and a carbohydrate-binding module (CBM, type 2 in PPN and bacteria) or only of the catalytic domain while the CBM2 was retrieved by domain shuffling later in evolution. Previous studies on the evolution of these genes have focused primarily on data of sedentary nematodes, while in this study, extra data from migratory nematodes were included.     

Conserved intron positions in ancient protein modules

Background  

The timing of the origin of introns is of crucial importance for an understanding of early genome architecture. The Exon theory of genes proposed a role for introns in the formation of multi-exon proteins by exon shuffling and predicts the presence of conserved splice sites in ancient genes. In this study, large-scale analysis of potential conserved splice sites was performed using an intron-exon database (ExInt) derived from GenBank.     

uShuffle: A useful tool for shuffling biological sequences while preserving the k-let counts

Background  

Randomly shuffled sequences are routinely used in sequence analysis to evaluate the statistical significance of a biological sequence. In many cases, biologists need sophisticated shuffling tools that preserve not only the counts of distinct letters but also higher-order statistics such as doublet counts, triplet counts, and, in general, k-let counts.     

Construction and Immunogenicity Evaluation of Recombinant Influenza A Viruses Containing Chimeric Hemagglutinin Genes Derived from Genetically Divergent Influenza A H1N1 Subtype Viruses

Background and Objectives

Influenza A viruses cause highly contagious diseases in a variety of hosts, including humans and pigs. To develop a vaccine that can be broadly effective against genetically divergent strains of the virus, in this study we employed molecular breeding (DNA shuffling) technology to create a panel of chimeric HA genes.

Methods and Results

Each chimeric HA gene contained genetic elements from parental swine influenza A viruses that had a history of zoonotic transmission, and also from a 2009 pandemic virus. Each parental virus represents a major phylogenetic clade of influenza A H1N1 viruses. Nine shuffled HA constructs were initially screened for immunogenicity in mice by DNA immunization, and one chimeric HA (HA-129) was expressed on both a A/Puerto Rico/8/34 backbone with mutations associated with a live, attenuated phenotype (PR8_LAIV-129) and a A/swine/Texas/4199-2/98 backbone (TX98-129). When delivered to mice, the PR8_LAIV-129 induced antibodies against all four parental viruses, which was similar to the breadth of immunity observed when HA-129 was delivered as a DNA vaccine. This chimeric HA was then tested as a candidate vaccine in a nursery pig model, using inactivated TX98-129 virus as the backbone. The results demonstrate that pigs immunized with HA-129 developed antibodies against all four parental viruses, as well as additional primary swine H1N1 influenza virus field isolates.

Conclusion

This study established a platform for creating novel genes of influenza viruses using a molecular breeding approach, which will have important applications toward future development of broadly protective influenza virus vaccines.     

Different dynamics of genome content shuffling among host-specificity groups of the symbiotic actinobacterium Frankia

Background

Frankia is a genus of soil actinobacteria forming nitrogen-fixing root-nodule symbiotic relationships with non-leguminous woody plant species, collectively called actinorhizals, from eight dicotyledonous families. Frankia strains are classified into four host-specificity groups (HSGs), each of which exhibits a distinct host range. Genome sizes of representative strains of Alnus, Casuarina, and Elaeagnus HSGs are highly diverged and are positively correlated with the size of their host ranges.

Results

The content and size of 12 Frankia genomes were investigated by in silico comparative genome hybridization and pulsed-field gel electrophoresis, respectively. Data were collected from four query strains of each HSG and compared with those of reference strains possessing completely sequenced genomes. The degree of difference in genome content between query and reference strains varied depending on HSG. Elaeagnus query strains were missing the greatest number (22–32%) of genes compared with the corresponding reference genome; Casuarina query strains lacked the fewest (0–4%), with Alnus query strains intermediate (14–18%). In spite of the remarkable gene loss, genome sizes of Alnus and Elaeagnus query strains were larger than would be expected based on total length of the absent genes. In contrast, Casuarina query strains had smaller genomes than expected.

Conclusions

The positive correlation between genome size and host range held true across all investigated strains, supporting the hypothesis that size and genome content differences are responsible for observed diversity in host plants and host plant biogeography among Frankia strains. In addition, our results suggest that different dynamics of shuffling of genome content have contributed to these symbiotic and biogeographic adaptations. Elaeagnus strains, and to a lesser extent Alnus strains, have gained and lost many genes to adapt to a wide range of environments and host plants. Conversely, rather than acquiring new genes, Casuarina strains have discarded genes to reduce genome size, suggesting an evolutionary orientation towards existence as specialist symbionts.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-609) contains supplementary material, which is available to authorized users.     

Genome shuffling of Lactobacillus plantarum C88 improves adhesion

Genome shuffling is an important method for rapid improvement in microbial strains for desired phenotypes. In this study, ultraviolet irradiation and nitrosoguanidine were used as mutagens to enhance the adhesion of the wild-type Lactobacillus plantarum C88. Four strains with better property were screened after mutagenesis to develop a library of parent strains for three rounds of genome shuffling. Fusants F3-1, F3-2, F3-3, and F3-4 were screened as the improved strains. The in vivo and in vitro tests results indicated that the population after three rounds of genome shuffling exhibited improved adhesive property. Random Amplified Polymorphic DNA results showed significant differences between the parent strain and recombinant strains at DNA level. These results suggest that the adhesive property of L. plantarum C88 can be significantly improved by genome shuffling. Improvement in the adhesive property of bacterial cells by genome shuffling enhances the colonization of probiotic strains which further benefits to exist probiotic function.     

Somatic gene transfer into the lactating ovine mammary gland

Background

Somatic gene therapy requires safe and efficient techniques for the gene transfer procedure. The ovine mammary gland is described as a model system for the evaluation of somatic gene transfer methods.

Methods

Different gene delivery formulations were retrogradely injected into the mammary gland of lactating sheep. The efficiency of the gene transfer was subsequently measured by the detection of the secreted transgene products in the milk. To counteract the milk flow in the lactating gland caused by the permanent milk production, a newly developed pretreatment of the mammary gland with hyperosmotic solutions was applied. In addition, in vivo electroporation of DNA into the mammary gland is described.

Results

Gene transfer using naked DNA or simple complexes of DNA with polycations did not result in traceable amounts of reporter gene products. However, utilizing the complex cationic lipid DOSPER, a peak expression of about 400 ng/ml was observed 6 days after transfection. Maximum expression rates of more than 1 µg/ml were obtained by combining hyperosmotic pretreatment and receptor‐mediated gene transfer. For the in vivo electroporation, the proof of principle for this technique in the mammary gland is reported.

Conclusions

The ovine mammary gland turned out to be a very well suited as a model system for evaluation and optimization of various gene transfer protocols. Copyright © 2002 John Wiley & Sons, Ltd.

     

Unraveling the effect of genomic structural changes in the rhesus macaque - implications for the adaptive role of inversions

Background

By reshuffling genomes, structural genomic reorganizations provide genetic variation on which natural selection can work. Understanding the mechanisms underlying this process has been a long-standing question in evolutionary biology. In this context, our purpose in this study is to characterize the genomic regions involved in structural rearrangements between human and macaque genomes and determine their influence on meiotic recombination as a way to explore the adaptive role of genome shuffling in mammalian evolution.

Results

We first constructed a highly refined map of the structural rearrangements and evolutionary breakpoint regions in the human and rhesus macaque genomes based on orthologous genes and whole-genome sequence alignments. Using two different algorithms, we refined the genomic position of known rearrangements previously reported by cytogenetic approaches and described new putative micro-rearrangements (inversions and indels) in both genomes. A detailed analysis of the rhesus macaque genome showed that evolutionary breakpoints are in gene-rich regions, being enriched in GO terms related to immune system. We also identified defense-response genes within a chromosome inversion fixed in the macaque lineage, underlying the relevance of structural genomic changes in evolutionary and/or adaptation processes. Moreover, by combining in silico and experimental approaches, we studied the recombination pattern of specific chromosomes that have suffered rearrangements between human and macaque lineages.

Conclusions

Our data suggest that adaptive alleles – in this case, genes involved in the immune response – might have been favored by genome rearrangements in the macaque lineage.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-530) contains supplementary material, which is available to authorized users.     

Intraspecific variation of recombination rate in maize

Background

In sexually reproducing organisms, meiotic crossovers ensure the proper segregation of chromosomes and contribute to genetic diversity by shuffling allelic combinations. Such genetic reassortment is exploited in breeding to combine favorable alleles, and in genetic research to identify genetic factors underlying traits of interest via linkage or association-based approaches. Crossover numbers and distributions along chromosomes vary between species, but little is known about their intraspecies variation.

Results

Here, we report on the variation of recombination rates between 22 European maize inbred lines that belong to the Dent and Flint gene pools. We genotype 23 doubled-haploid populations derived from crosses between these lines with a 50 k-SNP array and construct high-density genetic maps, showing good correspondence with the maize B73 genome sequence assembly. By aligning each genetic map to the B73 sequence, we obtain the recombination rates along chromosomes specific to each population. We identify significant differences in recombination rates at the genome-wide, chromosome, and intrachromosomal levels between populations, as well as significant variation for genome-wide recombination rates among maize lines. Crossover interference analysis using a two-pathway modeling framework reveals a negative association between recombination rate and interference strength.

Conclusions

To our knowledge, the present work provides the most comprehensive study on intraspecific variation of recombination rates and crossover interference strength in eukaryotes. Differences found in recombination rates will allow for selection of high or low recombining lines in crossing programs. Our methodology should pave the way for precise identification of genes controlling recombination rates in maize and other organisms.