Changes in exon-intron structure during vertebrate evolution affect the splicing pattern of exons

Exon-intron architecture is one of the major features directing the splicing machinery to the short exons that are located within long flanking introns. However, the evolutionary dynamics of exon-intron architecture and its impact on splicing is largely unknown. Using a comparative genomic approach, we analyzed 17 vertebrate genomes and reconstructed the ancestral motifs of both 3' and 5' splice sites, as also the ancestral length of exons and introns. Our analyses suggest that vertebrate introns increased in length from the shortest ancestral introns to the longest primate introns. An evolutionary analysis of splice sites revealed that weak splice sites act as a restrictive force keeping introns short. In contrast, strong splice sites allow recognition of exons flanked by long introns. Reconstruction of the ancestral state suggests these phenomena were not prevalent in the vertebrate ancestor, but appeared during vertebrate evolution. By calculating evolutionary rate shifts in exons, we identified cis-acting regulatory sequences that became fixed during the transition from early vertebrates to mammals. Experimental validations performed on a selection of these hexamers confirmed their regulatory function. We additionally revealed many features of exons that can discriminate alternative from constitutive exons. These features were integrated into a machine-learning approach to predict whether an exon is alternative. Our algorithm obtains very high predictive power (AUC of 0.91), and using these predictions we have identified and successfully validated novel alternatively spliced exons. Overall, we provide novel insights regarding the evolutionary constraints acting upon exons and their recognition by the splicing machinery.     

Origin and evolution of operons and metabolic pathways

The emergence and evolution of metabolic pathways represented a crucial step in molecular and cellular evolution, allowing primitive organisms to become less dependent on exogenous sources of organic compounds. This work will review the main theories accounting for their assembly and for the origin and evolution of prokaryotic operons.     

Prediction of many new exons and introns in Plasmodium falciparum chromosome 2.

The current prediction of genes in the Plasmodium falciparum genome database relies upon a limited number of specially developed computer algorithms. We have re-annotated the sequence of chromosome 2 of P. falciparum by a computer-assisted manual analysis, which is described here. Of 161 newly predicted introns, we have experimentally confirmed 98. We regard 110 introns from the previously published analyses as probable, we delete 3, change 26 and add 135. We recognise 214 genes in chromosome 2. We have predicted introns in 121 genes. The increased complexity of gene structure on chromosome 2 is likely to be mirrored by the entire genome.     

Origin and evolution of the integumentary skeleton in non-tetrapod vertebrates

Most non-tetrapod vertebrates develop mineralized extra-oral elements within the integument. Known collectively as the integumentary skeleton, these elements represent the structurally diverse skin-bound contribution to the dermal skeleton. In this review we begin by summarizing what is known about the histological diversity of the four main groups of integumentary skeletal tissues: hypermineralized (capping) tissues; dentine; plywood-like tissues; and bone. For most modern taxa, the integumentary skeleton has undergone widespread reduction and modification often rendering the homology and relationships of these elements confused and uncertain. Fundamentally, however, all integumentary skeletal elements are derived (alone or in combination) from only two types of cell condensations: odontogenic and osteogenic condensations. We review the origin and diversification of the integumentary skeleton in aquatic non-tetrapods (including stem gnathostomes), focusing on tissues derived from odontogenic (hypermineralized tissues, dentines and elasmodine) and osteogenic (bone tissues) cell condensations. The novelty of our new scenario of integumentary skeletal evolution resides in the demonstration that elasmodine, the main component of elasmoid scales, is odontogenic in origin. Based on available data we propose that elasmodine is a form of lamellar dentine. Given its widespread distribution in non-tetrapod lineages we further propose that elasmodine is a very ancient tissue in vertebrates and predict that it will be found in ancestral rhombic scales and cosmoid scales.     

Origin and evolution of viroids and viroid-like satellite RNAs

Viroids, the smallest and simplest agents of infectious disease, cause a number of economically important diseases of crop plants. Present evidence indicates that most of these diseases originated recently (in the 20th century) by chance transfer of viroids from endemically infected wild plants or by use of viroid-infected germplasm during plant breeding. Modern agricultural practices, such as widespread monoculture of genetically identical plants, and worldwide distribution of planting material, has made it possible for the pathogens to maintain themselves in the crop plants and to conquer new territories. Phylogenetic analysis of their nucleotide sequences indicates that viroids and satellite RNAs represent a monophyletic group, with all but the two self-cleaving viroids forming one cluster and the satellite RNAs another. The two self-cleaving viroids are phylogenetically distant from either cluster; they may represent ancestral forms. Results from site-directed mutagenesis experiments indicate that, upon exposure to selective pressures, viroids can evolve extremely rapidly, with another, fitter, component of the quasi-species often becoming dominant within days or weeks. This extreme plasticity of their nucleotide sequences establishes viroids as the most rapidly evolving biological system known.     

Origin and evolution of TNF and TNF receptor superfamilies

The tumor necrosis factor superfamily (TNFSF) and the TNF receptor superfamily (TNFRSF) have an ancient evolutionary origin that can be traced back to single copy genes within Arthropods. In humans, 18 TNFSF and 29 TNFRSF genes have been identified. Evolutionary models account for the increase in gene number primarily through multiple whole genome duplication events as well as by lineage and/or species-specific tandem duplication and translocation. The identification and functional analyses of teleost ligands and receptors provide insight into the critical transition between invertebrates and higher vertebrates. Bioinformatic analyses of fish genomes and EST datasets identify 14 distinct ligand groups, some of which are novel to teleosts, while to date, only limited numbers of receptors have been characterized in fish. The most studied ligand is TNF of which teleost species possess between 1 and 3 copies as well as a receptor similar to TNFR1. Functional studies using zebrafish indicate a conserved role of this ligand–receptor system in the regulation of cell survival and resistance to infectious disease. The increasing interest and use of TNFSF and TNFRSF modulators in human and animal medicine underscores the need to understand the evolutionary origins as well as conserved and novel functions of these biologically important molecules.     

Origin and evolution of TNF and TNF receptor superfamilies

The tumor necrosis factor superfamily (TNFSF) and the TNF receptor superfamily (TNFRSF) have an ancient evolutionary origin that can be traced back to single copy genes within Arthropods. In humans, 18 TNFSF and 29 TNFRSF genes have been identified. Evolutionary models account for the increase in gene number primarily through multiple whole genome duplication events as well as by lineage and/or species-specific tandem duplication and translocation. The identification and functional analyses of teleost ligands and receptors provide insight into the critical transition between invertebrates and higher vertebrates. Bioinformatic analyses of fish genomes and EST datasets identify 14 distinct ligand groups, some of which are novel to teleosts, while to date, only limited numbers of receptors have been characterized in fish. The most studied ligand is TNF of which teleost species possess between 1 and 3 copies as well as a receptor similar to TNFR1. Functional studies using zebrafish indicate a conserved role of this ligand-receptor system in the regulation of cell survival and resistance to infectious disease. The increasing interest and use of TNFSF and TNFRSF modulators in human and animal medicine underscores the need to understand the evolutionary origins as well as conserved and novel functions of these biologically important molecules.     

Origin and evolution of the vertebrate immune system.

The immune system is a complex evolutionary unit and it would be simplistic to conclude that the immune systems of all primitive vertebrates are primitive. Because of the large number of elements involved, many evolutionary events must have taken place, some of them neutral, some of them selected, to constitute the systems that we are looking at towards the end of the 20th century. All these systems have perhaps evolved beyond the apparent evolutionary state of the species in which they are found. They have been modulated by factors linked not only to the internal evolution of their elementary genes, but also by coevolution with factors in the internal environment, such as cellular constraints, metabolism, mode of reproduction and progeny size. It seems that good inventions are long lasting, which is the reason why some elements of the invertebrate immune system can be found with similar functions in vertebrates (defensins). It is also the reason why Ig domains have been exploited in so many ways, whether for the immune system or not. Again, they had an evolution of their own. The comparative study of the immune systems carried out on the occasion of this phylogenetic survey shows a world particularly dynamic and diverse. The comparisons between the solutions chosen by the various phyla of the animal kingdom, or closer to us by the various classes of vertebrates, allow us to distinguish the essential features of the immune system. From this viewpoint, this approach is not only of phylogenetic interest, but also has an applied aspect. Increasing our knowledge in this area could help suggest solutions to clinicians when they are faced with deficiencies and abnormalities in the immune system of man.     

Origin and evolution of Georgia 98 (GA98), a new serotype of avian infectious bronchitis virus

We previously identified GA98, a new serotype of infectious bronchitis virus (IBV), which is closely related to the DE072 serotype of IBV genetically, but not antigenically. Herein, we analyzed the 421bp sequence of a hypervariable region (HVR) (position 114–534, counting from the ATG start site) of the S1 subunit of GA98 IBVs to further examine the evolution of these viruses. These viruses were isolated between the years 1997 and 2000. Phylogenetic analysis of the deduced amino acid sequence on that region indicated that GA98 isolates from different regions of Georgia were the result of a single introduction of the S1 gene of the DE072 serotype progenitor. Most of the mutations were nonsynonymous and had become fixed in a progressive manner. The evolutionary and mutation rates in the HVR was calculated as 2.5 and 1.5% per year, respectively. This new serotype of IBV appears to be evolving very fast compared with other serotypes of IBV. We further determined the complete coding sequence of the S1 gene of seven isolates obtained from one selected region in North Georgia. Together with virus neutralization data, it appears that GA98 arose from immune selection caused by DE072 vaccine use. Reasons for this conclusion are discussed.     

Origin and evolution of somatic cell testicular tumours in transgenic mice

Transgenic mice bearing a construct in which the expression of the SV40 oncogene is directed by the AMH promoter (AT mice) develop testicular tumours in adult life. We aimed to study early steps of tumour development and characterize tumours at different ages by histological, morphometric, and immunohistochemical techniques. One‐ to 3‐month‐old AT mice depicted multifocal Leydig cell hyperplasia. The testicular volume occupied by interstitial tissue was significantly higher in 3‐month‐old AT mice in comparison with littermate controls. Between 5 1/2 and 7 months, microscopic interstitial tumours developed that progressively evolved to form large confluent areas of high mitotic index in 7‐ to 14‐month‐old AT mice. Tumour cells had the characteristics and histoarchitecture of Leydig cells, or formed solid cord‐like structures reminiscent of those seen in Sertoli cell tumours. Hyperplastic areas and tumours diffusely expressed 3β‐hydroxysteroid dehydrogenase (3β‐HSD) in Leydig cell areas. AMH expression was negative in Leydig cell conglomerates and tumours and variable in cord‐like tumours. The SV40 T antigen and markers of cell proliferation (PCNA) were intensely positive in hyperplastic cells and tumours. Control mice of similar ages showed neither hyperplasia nor tumours, and SV40 T expression was always negative. In conclusion, transgenic mice develop large testicular tumours that are preceded by interstitial hyperplasia and microtumours. The histological and immunohistochemical phenotype of tumours (Leydig and Sertoli cell differentiation, positive 3β‐HSD, and variable AMH) suggests a mixed differentiation of somatic cells of the specialized gonadal stroma. The finding that an oncogene directed by a promoter specifically active in fetal Sertoli cells has given rise to testicular tumours of mixed differentiation is compatible with a common origin of Leydig and Sertoli cells from the specific stroma of the gonadal ridge, as supported by double labelling experiments in fetal mice showing co‐localization of the transgene with Sertoli and Leydig cell markers. Copyright © 2010 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

Origin and evolution of viruses causing classical swine fever in Cuba

We have analyzed the origin and evolution of viruses from the classical swine fever (CSF) epidemic that affects Cuba since 2001 by nucleotide sequencing of regions within the E2 glycoprotein and the NS5B (polymerase) genes. The sequence of 190 nucleotides from E2 gene was determined for 10 CSF viruses isolated at different locations of the island, and used for phylogenetic analyses, including sequences from viruses of the 1993--1997 epizootic, previously determined, as well as those from representatives of the different CSFV genotypes. The phylogenetic tree obtained indicates that viruses circulating at present belong to the subgroup 1.2 and are closely related to those isolated during the 1993--1997 epizootic, including the strain Margarita used for vaccine potency tests in Cuba. However, the pattern of evolution revealed by these analyses was different than that observed previously, in which western isolates were almost identical to Margarita strain, while eastern isolates showed a higher level of genetic diversification. In this case, all the viruses analyzed grouped in an independent, define cluster that is closely related, albeit distinguishable, from that of Margarita-related viruses that previously circulated in the western part of Cuba. In addition, the 2001--2003 viruses showed a branched pattern with a level of sequence diversification similar to that observed in the eastern 1993--1997 viruses. Interestingly, a significant fraction (about 54%) of the mutations found in the E2 sequence led to amino acid replacements. This high rate of non-synonymous mutations was not found in the previous Cuban epizootic and has not been reported for other CSF outbreaks. In spite of these amino acid replacements, no antigenic changes were observed in the reactivity of different isolates with CSFV-specific MAbs and polyclonal sera. The phylogenetic tree derived from 409 nucleotides of NS5B gene of seven isolates and Margarita strain, was consistent with that obtained from E2 sequences. In this region, encoding a non-structural protein, a low level of fixation of non-synonymous mutations was observed. The results obtained suggests that epidemiological factors affecting CSFV spread during the current epizootic in Cuba can favour the fixation of non-synonymous mutation in the E2 gene, which could be associated with a lower severity in the clinical signs developed by most of the affected animals.     

一种用于基因5'exons预测的新方法

基因预测是生物信息学领域中的一个重要研究方向.本文在研究了基因局部特征的基础上,针对现有遗传算法在预测基因5'exons方面的不足,从生物免疫机制出发,构建了一种用于基因5'exons预测的免疫遗传算法.该算法利用T细胞发育过程中强大的多样性维持机制来设计算法的选择机制,提高算法的求解性能.实验结果表明该算法提高了基因预测的精度,提供了一种可能的研究基因预测方案.     

Origin and structural evolution of the early proliferating oval cells in rat liver

We have analyzed the histological changes in rat liver after 2-acetylaminofluorene (AAF) administration. The data demonstrate that AAF-induced oval cells were preferentially generated by proliferation of the terminal biliary ductules that we suggest constitute the primary hepatic stem cell niche. The oval cells formed ductular structures, representing an extension of the canals of Hering. This histological organization provides continuous bile drainage of the hepatocytes and uninterrupted blood flow in the sinusoids. The oval cell ductules are surrounded by a continuous basement membrane that is intermittently disrupted by processes of stellate cells that form direct cell-cell contact with the oval cells. Although both AAF treatment and bile duct ligation results in proliferation of biliary epithelial cells, the mechanism(s) responsible for the proliferation of the biliary epithelium seems to differ in the two models. In contrast to the biliary proliferation stimulated by bile ligation, AAF-induced oval cell proliferation as well as the capacity of these cells to differentiate into hepatocytes, bile epithelial cells and possibly other cell lineages can be blocked by administration of dexamethasone.     

B*27 in molecular diagnostics: impact of new alleles and polymorphism outside exons 2 and 3

HLA-B*27 is known to be associated with ankylosing spondylitis and several methods have been applied to determine its presence or absence. In this report two molecular methods were used for detection of B*27. The polymerase chain reaction sequence-specific primer (PCR-SSP) method was performed to detect the presence or absence of B*27, whereas the sequence-based typing method (SBT) was used to identify the B*27 subtype. The PCR-SSP method used to detect B*27 was updated to enable the detection of all B*27 alleles. The typing results obtained by this method were compared with the serological typings of 262 individuals. Fifty of them were found to be B*27 positive by PCR-SSP and 46 also showed positive serological reactions with B27-specific sera. The four discrepancies were the result of the presence of B*2712 in three individuals and B*2715 in one individual; both alleles showed no serological reactions with B27-specific antisera. With SBT the sequences of exons 1 through 4 were determined to unequivocally assign the B*27 alleles. Eleven different subtypes were detected in 78 individuals, including three new B*27 alleles: B*27054, B*2715 and B*2717. The allele B*27054 showed an allelic drop out when exon 3 was amplified. Three differences with B*27052 were demonstrated; one in exon 1, one in intron 1 and one in intron 2, the latter being responsible for the allelic drop out. The B*2715 allele was serologically not detectable with several B27-specific sera, but showed Bw4-positive reactions. The sequence of B*2715 showed two mismatches with B*2704. The sequence of B*2717 showed one mismatch with B*27052 at position 248 (A-->T), which was considered to be a conserved position in all B alleles.     

Adaptive evolution of the MHC class III‐encoded receptor RAGE in primates and murine rodents

The receptor for advanced glycation end products is associated with a series of physiological and pathological processes. Here, we studied the evolution of this multiligand receptor in primates and murine rodents. The evolutionary analyses reveal that adaptive selection had contributed to the variation at a number of amino acid sites in both taxa. Further, the major adaptively selected sites of both taxa are located on the extracellular ligand‐ and intracellular adaptor‐binding regions and receptor oligomerization‐related surfaces. The co‐occurrence of adaptive evolution on the homologue domains suggests that they could play similar roles in these taxa. In terms of advantage fitness, the adaptive changes at these sites could contribute to host defence against the potential challenges towards these interactions and relevant signalling pathways, or the specificity of these essential points.