Evolution Of Alternative Splicing Research Articles

Catsnap: a user-friendly algorithm for determining the conservation of protein variants reveals extensive parallelisms in the evolution of alternative splicing.

Understanding the evolutionary conservation of complex eukaryotic transcriptomes significantlyilluminates the physiological relevance of alternative splicing (AS). Examining the evolutionary depth of a given AS event with ordinary homology searches is generally challenging and time-consuming. Here, we present Catsnap, an algorithmic pipeline for assessing the conservation of putative protein isoforms generated by AS. It employs a machine learning approach following a database search with the provided pair of protein sequences. We used the Catsnap algorithm for analyzing the conservation of emerging experimentally characterized alternative proteins from plants and animals. Indeed, most of them are conserved among other species. Catsnap can detect the conserved functional protein isoforms regardless of the AS type by which they are generated. Notably,we found that while the primary amino acid sequence is maintained, the type of AS determining the inclusion or exclusion of protein regions varies throughout plant phylogenetic lineages in these proteins. We also document that this phenomenon is less seen among animals. In sum, our algorithm highlights the presence of unexpectedly frequent hotspots where protein isoforms recurrently arise to carry physiologically relevant functions. The user web interface is available at https://catsnap.cesnet.cz/.

The evolution of alternative splicing in glioblastoma under therapy

BackgroundAlternative splicing is a rich source of tumor-specific neoantigen targets for immunotherapy. This holds promise for glioblastomas (GBMs), the most common primary tumors of the adult brain, which are resistant to standard-of-care therapy. Although most clinical trials enroll patients at recurrence, most preclinical studies have been done with specimens from primary disease. There are limited expression data from GBMs at recurrence and surprisingly little is known about the evolution of splicing patterns under therapy.ResultWe profile 37 primary-recurrent paired human GBM specimens via RNA sequencing. We describe the landscape of alternative splicing in GBM at recurrence and contrast that to primary and non-malignant brain-tissue specimens. By screening single-cell atlases, we identify cell-type-specific splicing patterns and novel splicing events in cell-surface proteins that are suitable targets for engineered T cell therapies. We identify recurrent-specific isoforms of mitogen-activated kinase pathway genes that enhance invasiveness and are preferentially expressed by stem-like cells.ConclusionThese studies shed light on gene expression in recurrent GBM and identify novel targets for therapeutic development.

Evolution of Alternative Splicing in Eudicots.

Alternative pre-mRNA splicing (AS) is prevalent in plants and is involved in many interactions between plants and environmental stresses. However, the patterns and underlying mechanisms of AS evolution in plants remain unclear. By analyzing the transcriptomes of four eudicot species, we revealed that the divergence of AS is largely due to the gains and losses of AS events among orthologous genes. Furthermore, based on a subset of AS, in which AS can be directly associated with specific transcripts, we found that AS that generates transcripts containing premature termination codons (PTC), are likely more conserved than those that generate non-PTC containing transcripts. This suggests that AS coupled with nonsense-mediated decay (NMD) might play an important role in affecting mRNA levels post-transcriptionally. To understand the mechanisms underlying the divergence of AS, we analyzed the key determinants of AS using a machine learning approach. We found that the presence/absence of alternative splice site (SS) within the junction, the distance between the authentic SS and the nearest alternative SS, the size of exon–exon junctions were the major determinants for both alternative 5′ donor site and 3′ acceptor site among the studied species, suggesting a relatively conserved AS mechanism. The comparative analysis further demonstrated that variations of the identified AS determinants significantly contributed to the AS divergence among closely related species in both Solanaceae and Brassicaceae taxa. Together, these results provide detailed insights into the evolution of AS in plants.

Role and convergent evolution of competing RNA secondary structures in mutually exclusive splicing

ABSTRACTExon or cassette duplication is an important means of expanding protein and functional diversity through mutually exclusive splicing. However, the mechanistic basis of this process in non-arthropod species remains poorly understood. Here, we demonstrate that MRP1 genes underwent tandem exon duplication in Nematoda, Platyhelminthes, Annelida, Mollusca, Arthropoda, Echinodermata, and early-diverging Chordata but not in late-diverging vertebrates. Interestingly, these events were of independent origin in different phyla, suggesting convergent evolution of alternative splicing. Furthermore, we showed that multiple sets of clade-conserved RNA pairings evolved to guide species-specific mutually exclusive splicing in Arthropoda. Importantly, we also identified a similar structural code in MRP exon clusters of the annelid, Capitella teleta, and chordate, Branchiostoma belcheri, suggesting an evolutionarily conserved competing pairing-guided mechanism in bilaterians. Taken together, these data reveal the molecular determinants and RNA pairing-guided evolution of species-specific mutually exclusive splicing spanning more than 600 million years of bilaterian evolution. These findings have a significant impact on our understanding of the evolution of and mechanism underpinning isoform diversity and complex gene structure.

The Evolutionary Landscape of Alternative Splicing in Vertebrate Species

How species with similar repertoires of protein-coding genes differ so markedly at the phenotypic level is poorly understood. By comparing organ transcriptomes from vertebrate species spanning ~350 million years of evolution, we observed significant differences in alternative splicing complexity between vertebrate lineages, with the highest complexity in primates. Within 6 million years, the splicing profiles of physiologically equivalent organs diverged such that they are more strongly related to the identity of a species than they are to organ type. Most vertebrate species-specific splicing patterns are cis-directed. However, a subset of pronounced splicing changes are predicted to remodel protein interactions involving trans-acting regulators. These events likely further contributed to the diversification of splicing and other transcriptomic changes that underlie phenotypic differences among vertebrate species.

Evolution of alternative splicing in newly evolved genes of Drosophila

New gene origination is a fundamental process underlying evolution of biological diversity. Although new genes usually evolve rapidly in sequences, structure and expression, the evolutionary pattern of alternative splicing (AS) in new genes and the molecular mechanisms involved in this alternation remain to be explored. Here, we used the new genes identified in the Drosophila melanogaster lineage to study alternation of AS and the possible functional consequences of these genes. We found that new genes tended to exhibit low degree of AS, though a few new genes were alternatively spliced. Interestingly loss of introns in retroposed new genes can only account for one third of the low-level AS in new genes, while partial gene duplication without AS exons and mutations in the duplicated AS exons/introns together have resulted in two-third AS losses in new genes, indicating that reducing the degree of AS is a general trend in all categories of new genes. Further investigations on tissue expression patterns of these new genes showed that those with AS alternation had a relatively lower expression level, were expressed in fewer tissues and tended to be more likely expressed in testis than their parental genes. All these observations imply that these new genes may have gained diverged structures and expression patterns from their parental genes after AS alternation.

Evolution of alternative splicing in primate brain transcriptomes

Alternative splicing is a predominant form of gene regulation in higher eukaryotes. The evolution of alternative splicing provides an important mechanism for the acquisition of novel gene functions. In this work, we carried out a genome-wide phylogenetic survey of lineage-specific splicing patterns in the primate brain, via high-density exon junction array profiling of brain transcriptomes of humans, chimpanzees and rhesus macaques. We identified 509 genes showing splicing differences among these species. RT-PCR analysis of 40 exons confirmed the predicted splicing evolution of 33 exons. Of these 33 exons, outgroup analysis using rhesus macaques confirmed 13 exons with human-specific increase or decrease in transcript inclusion levels after humans diverged from chimpanzees. Some of the human-specific brain splicing patterns disrupt domains critical for protein-protein interactions, and some modulate translational efficiency of their host genes. Strikingly, for exons showing splicing differences across species, we observed a significant increase in the rate of silent substitutions within exons, coupled with accelerated sequence divergence in flanking introns. This indicates that evolution of cis-regulatory signals is a major contributor to the emergence of human-specific splicing patterns. In one gene (MAGOH), using minigene reporter assays, we demonstrated that the combination of two human-specific cis-sequence changes created its human-specific splicing pattern. Together, our data reveal widespread human-specific changes of alternative splicing in the brain and suggest an important role of splicing in the evolution of neuronal gene regulation and functions.

Alternative Splicing in the Postgenomic Era, edited by Benjamin J. Blencowe and Brenton R. Graveley. 2007, XXIV, Springer, New York. ISBN: 978-0-387-77373-5.

<i>Alternative Splicing in the Postgenomic Era</i>, edited by Benjamin J. Blencowe and Brenton R. Graveley. 2007, XXIV, Springer, New York. ISBN: 978-0-387-77373-5.

Comparative Genomics and Evolution of Alternative Splicing: The Pessimists′ Science

AbstractChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 200 leading journals. To access a ChemInform Abstract, please click on HTML or PDF.

Comparative Genomics and Evolution of Alternative Splicing: The Pessimists' Science

Comparative Genomics and Evolution of Alternative Splicing: The Pessimists' Science

Functional and evolutionary analysis of alternatively spliced genes is consistent with an early eukaryotic origin of alternative splicing

BackgroundAlternative splicing has been reported in various eukaryotic groups including plants, apicomplexans, diatoms, amoebae, animals and fungi. However, whether widespread alternative splicing has evolved independently in the different eukaryotic groups or was inherited from their last common ancestor, and may therefore predate multicellularity, is still unknown. To better understand the origin and evolution of alternative splicing and its usage in diverse organisms, we studied alternative splicing in 12 eukaryotic species, comparing rates of alternative splicing across genes of different functional classes, cellular locations, intron/exon structures and evolutionary origins.ResultsFor each species, we find that genes from most functional categories are alternatively spliced. Ancient genes (shared between animals, fungi and plants) show high levels of alternative splicing. Genes with products expressed in the nucleus or plasma membrane are generally more alternatively spliced while those expressed in extracellular location show less alternative splicing. We find a clear correspondence between incidence of alternative splicing and intron number per gene both within and between genomes. In general, we find several similarities in patterns of alternative splicing across these diverse eukaryotes.ConclusionAlong with previous studies indicating intron-rich genes with weak intron boundary consensus and complex spliceosomes in ancestral organisms, our results suggest that at least a simple form of alternative splicing may already have been present in the unicellular ancestor of plants, fungi and animals. A role for alternative splicing in the evolution of multicellularity then would largely have arisen by co-opting the preexisting process.

Increase of functional diversity by alternative splicing

A large-scale analysis of protein isoforms arising from alternative splicing shows that alternative splicing tends to insert or delete complete protein domains more frequently than expected by chance, whereas disruption of domains and other structural modules is less frequent. If domain regions are disrupted, the functional effect, as predicted from 3D structure, is frequently equivalent to removal of the entire domain. Also, short alternative splicing events within domains, which might preserve folded structure, target functional residues more frequently than expected. Thus, it seems that positive selection has had a major role in the evolution of alternative splicing.

Evolution of alternative splicing: deletions, insertions and origin of functional parts of proteins from intron sequences

Alternative splicing is thought to be a major source of functional diversity in animal proteins. We analyzed the evolutionary conservation of proteins encoded by alternatively spliced genes and predicted the ancestral state for 73 cases of alternative splicing (25 insertions and 48 deletions). The amino acid sequences of most of the inserts in proteins produced by alternative splicing are as conserved as the surrounding sequences. Thus, alternative splicing often creates novel isoforms by the insertion of new, functional protein sequences that probably originated from noncoding sequences of introns.

Intron self-complementarity enforces exon inclusion in a yeast pre-mRNA.

Skipping of internal exons during removal of introns from pre-mRNA must be avoided for proper expression of most eukaryotic genes. Despite significant understanding of the mechanics of intron removal, mechanisms that ensure inclusion of internal exons in multi-intron pre-mRNAs remain mysterious. Using a natural two-intron yeast gene, we have identified distinct RNA-RNA complementarities within each intron that prevent exon skipping and ensure inclusion of internal exons. We show that these complementarities are positioned to act as intron identity elements, bringing together only the appropriate 5' splice sites and branchpoints. Destroying either intron self-complementarity allows exon skipping to occur, and restoring the complementarity using compensatory mutations rescues exon inclusion, indicating that the elements act through formation of RNA secondary structure. Introducing new pairing potential between regions near the 5' splice site of intron 1 and the branchpoint of intron 2 dramatically enhances exon skipping. Similar elements identified in single intron yeast genes contribute to splicing efficiency. Our results illustrate how intron secondary structure serves to coordinate splice site pairing and enforce exon inclusion. We suggest that similar elements in vertebrate genes could assist in the splicing of very large introns and in the evolution of alternative splicing.

Proto-splice site model of intron origin

It is proposed that nuclear pre-mRNA introns (classical introns) were first generated as by-products during the evolution of alternative splicing. They were formed whenever two splice sites within the coding sequence of ancestral genes were used at a frequency that removed the coding constraint from the intervening sequence. Once introns had evolved, it is suggested that they were spread by the splicing machinery which inserted them into proto or cryptic-splice sites of other genes by reverse splicing, so giving rise to genes that have introns yet are not alternatively spliced. It is argued that 5' and 3' splice sites evolved from common ancestral splice sites, referred to as proto-splice sites, that were bidirectional and had a core consensus sequence of C or A, A, G, R, which remains today as the immediate flanking sequence of most introns. The ancestral splicing machinery, although inefficient, would have been capable of generating vast mRNA diversity by splicing between proto-splice sites. Natural selection would be expected to have preserved mutations that increased the amounts of advantageously spliced mRNA. It is argued that this process drove the evolution of present 5' and 3' splice sites from a subset of proto-splice sites and also drove the evolution of a more efficient splicing machinery. The positions of most introns that evolved directly from the coding sequence would be expected to correlate with protein structure.