The Alernative Splicing Evolution Server allows you to rapidly and easily explore how alternative splicing has generated diversity in evolution at the protein level. Ases primary source of information are the gene annotations avalaible in the Ensembl database. It runs ThorAxe( Zea et al. 2021, check out also our preprint and ISMB-ECCB 2019 talk) and PhyloSofS ( Ait-hamlat et al.2020, PDF), a couple of tools we developed to identify transcripts minimal building blocks in evolution and reconstruct plausible scenarios explaining the transcripts we observe today.

For a quick start, you can see our YouTube tutorial:

You want to know more, collaborate, or you encounter a problem with the server? Please feel free to contact us!

Input

Please provide the name of the gene you are interested in (e.g. MAPK8), or preferably, its Ensembl Stable ID (e.g. ENSG00000107643). Note that gene names can be ambiguous, so you can use the Ensembl Stable IDs to avoid that problem. Guidelines for gene naming in Ensembl can be found here. By default, the analysis will be conducted across twelve species, namely human, gorilla, macaque, opossum, rat, mouse, cow, boar, platypus, xenopus, zebrafish and nematode, and human serves as reference.

Optional parameters for ThorAxe

ThorAxe will dynamically retrieve all complete protein coding transcripts annotated in the Ensembl database for the query gene and its one-to-one orthologs in the considered species. If a species does not have a one-to-one ortholog of the input gene, it will be excluded from the analysis, and a warning will be issued on the result page. By default, transcripts with low level of support ( TSL value lower than 3) are filtered out. Then ThorAxe will identify ortology relationships between the exons composing the input transcripts and will build an evolutionary splicing graph summarizing them. The first step of this procedure consists in grouping the exons sharing more than 30% sequence identity, by default.

Optional parameters for PhyloSofS

PhyloSofS will reconstruct plausible evolutionary scenarios explaining the input transcripts using the maximum parsimony principle. Three eveolutionary events are considered, namely transcript appearance, mutation and death. By default, death is not penalized ( death cost is equal to zero) to account for the incompleteness of annotation data. Moreover, the birth to mutation cost ratio is set to 3/2, which implies that only a few mutations will be tolerated along a phylogeny. This ratio is typically well suited for genes that have not diverged much in evolution, and it should be adapted for highly divergent genes.

Advanced parameters

In addition to the optional parameters, more advanced parameters can be tuned. Most of the default values have general validity. In ThorAxe, it may be useful to consider more than one orthologs per species. This can be done by choosing the 1:n or m:n orthology relationships instead of 1:1. You might also want to increase the minimum species number or minimum transcript number to be more stringent on the detection of alternative splicing events. In PhyloSofS, you might want to increase the number of iterations for very complex cases with a very large number of s-exons , transcripts and/or species.

Output

Please check our example result page.

Ases provides a comprehensive visualization of the results with pre-tuned parameters (nodes and edges colors and shapes) and with clear connections between the different representations of the biological objects (graph, forest of trees, multiple sequence alignments, sequences, genes and tables with ids and annotations).

Evolutionary splicing graph

This interactive graph summarizes the transcript variability observed for the query gene across the selected species. Each transcript corresponds to a path in the graph, divided into minimal building blocks called the s-exons. Each s-exon corresponds to a node in the graph. Formally, a s-exon is defined by a multiple sequence alignment where each sequence comes from a species. Upon clicking on a node in the interactive graph, you will be able to visualize the corresponding sequence logo and alignment in the panel below. You can easily move the nodes around to get a clearer picture of the graph topology and of the alternative splicing events. Each event is defined by a pair of canonical and alternative subpaths, and thus appears as a bubble in the graph. The colors of the nodes and the edges indicate the conservation levels of the s-exons and their junctions: number of species for each s-exon, and number of transcripts for each junction. The start and stop nodes are added for convenience, and remain hidden by default.

ThorAxe procedure unfolds in five main steps: (1) cluster genomic exons sharing some similarity, (2) define sub-exons within each species, (3) align all sub-exons belonging to the same cluster, (4) identify s-exons as continuous blocks in the alignment, (5) refine s-exons. The identifiers of the s-exons reflect this procedure. Specifically, the s-exon with id XX_YY corresponds to the YYth block in the alignment built for the XXth cluster. Hence, the s-exons sharing the same prefix come from the same cluster. The s-exons starting with "0" originate from genomic exons that were not included in the clustering (typically because they are very small, see the advanced parameters), and thus they are not associated with an alignment. Nevertheless, you can get access to their sequences in the s-exon table (see below).

Transcript phylogenetic forest

In this interactive forest, each tree represents the phylogeny of a transcript. More precisely, the root of the tree indicates the emergence of a new transcript in evolution, and the changes along the branches represent plausible scenarios explaining how this ancestral transcript has led to a set of observed transcripts in current-day species (the leaves). Hence, the internal nodes and the leaves represent transcripts. Dead ends (shown as triangles) indicate transcript losses. The different trees can be easily distinguished thanks to their different colors. Note that the whole forest is embedded in the gene tree retrieved from Ensembl. The visualisation of the forest is dynamically linked to the visualisation of the evolutionary splicing graph on the left. For instance, upon clicking on a node or a leaf in the forest, the corresponding path in the evolutionary splicing graph will be emphasised by grey boxes. Note that by default, the s-exons appearing only in one transcript from one species have been removed prior to the construction of the forest (see the advanced option no pruning). These s-exons will thus not appear as part of the path in the ESG. Moreover, upon clicking specifically on a leaf, the corresponding transcript sequence will be shown in the Transcript sequence panel at the bottom. You may simply copy-paste it in a domain-annotation or 3D-structure prediction web server. Transcripts can mutate along the branches by gaining or losing s-exons. If you click on a branch, you will get access to the description of the parent and child node/transcript and of the change(s) from one to the other.

Transcript sequence

This panel will show you the sequence and s-exon composition of a given transcript. You can select a transcript from the dropdown menu at the top or by clicking on a leaf of the Transcript phylogenetic forest. When a transcript is selected from the dropdown menu, the s-exon that compound the sequence will be highlighted on the Evolutionary splicing graph. The colour changes on the sequence indicate the s-exon changes. You can see the s-exon identifier for a given residue by keeping the mouse over a residue for some seconds.

Genomic coordinates

This plot shows the gene structure in terms of s-exons (coloured boxes). The s-exons are ordered in increasing (respectively decreasing) genomic coordinates for positive (resp. negative) stranded genes. The arrows indicate the strand direction. You can click on a s-exon box in a gene to see its information, including its genomic coordinates and nucleotide sequence. You can also click on the s-exon in the list on the right to highlight it on the genes. The entire list does not always fit the screen, but you can zoom out and pan the plot to see the s-exons. To unselect an item, click on it.

S-exon and alternative splicing event tables

These two interactive tables give detailed information about the s-exons and the alternative splicing events. You can for example select all transcripts annotated for a given species, get access to the list of building blocks for a given transcript, or select all insertions...etc.

Browser compatibility

The server's JavaScript code uses ECMAScript 2015 (ES6) features; therefore, it needs Chrome 58+, Edge 15+, Firefox 54+, Safari 10+, or Opera 55+. We have tested this server using the following browsers:

If you find any problem, please report it to us.