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An analysis of tissue-specific alternative splicing at the protein level.

ABSTRACT: The role of alternative splicing is one of the great unanswered questions in cellular biology. There is strong evidence for alternative splicing at the transcript level, and transcriptomics experiments show that many splice events are tissue specific. It has been suggested that alternative splicing evolved in order to remodel tissue-specific protein-protein networks. Here we investigated the evidence for tissue-specific splicing among splice isoforms detected in a large-scale proteomics analysis. Although the data supporting alternative splicing is limited at the protein level, clear patterns emerged among the small numbers of alternative splice events that we could detect in the proteomics data. More than a third of these splice events were tissue-specific and most were ancient: over 95% of splice events that were tissue-specific in both proteomics and RNAseq analyses evolved prior to the ancestors of lobe-finned fish, at least 400 million years ago. By way of contrast, three in four alternative exons in the human gene set arose in the primate lineage, so our results cannot be extrapolated to the whole genome. Tissue-specific alternative protein forms in the proteomics analysis were particularly abundant in nervous and muscle tissues and their genes had roles related to the cytoskeleton and either the structure of muscle fibres or cell-cell connections. Our results suggest that this conserved tissue-specific alternative splicing may have played a role in the development of the vertebrate brain and heart.

SUBMITTER: Rodriguez JM

PROVIDER: S-EPMC7561204 | biostudies-literature | 2020 Oct

REPOSITORIES: biostudies-literature

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An analysis of tissue-specific alternative splicing at the protein level.

Rodriguez Jose Manuel JM Pozo Fernando F di Domenico Tomas T Vazquez Jesus J Tress Michael L ML

PLoS computational biology 20201005 10

The role of alternative splicing is one of the great unanswered questions in cellular biology. There is strong evidence for alternative splicing at the transcript level, and transcriptomics experiments show that many splice events are tissue specific. It has been suggested that alternative splicing evolved in order to remodel tissue-specific protein-protein networks. Here we investigated the evidence for tissue-specific splicing among splice isoforms detected in a large-scale proteomics analysis ...[more]

PMID: 33017396

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Project description:BackgroundAlternative splicing contributes significantly to the complexity of the human transcriptome and proteome. Computational prediction of alternative splice isoforms are usually based on EST sequences that also allow to approximate the expression pattern of the related transcripts. However, the limited number of tissues represented in the EST data as well as the different cDNA construction protocols may influence the predictive capacity of ESTs to unravel tissue-specifically expressed transcripts.MethodsWe predict tissue and tumor specific splice isoforms based on the genomic mapping (SpliceNest) of the EST consensus sequences and library annotation provided in the GeneNest database. We further ascertain the potentially rare tissue specific transcripts as the ones represented only by ESTs derived from normalized libraries. A subset of the predicted tissue and tumor specific isoforms are then validated via RT-PCR experiments over a spectrum of 40 tissue types.ResultsOur strategy revealed 427 genes with at least one tissue specific transcript as well as 1120 genes showing tumor specific isoforms. While our experimental evaluation of computationally predicted tissue-specific isoforms revealed a high success rate in confirming the expression of these isoforms in the respective tissue, the strategy frequently failed to detect the expected restricted expression pattern. The analysis of putative lowly expressed transcripts using normalized cDNA libraries suggests that our ability to detect tissue-specific isoforms strongly depends on the expression level of the respective transcript as well as on the sensitivity of the experimental methods. Especially splice isoforms predicted to be disease-specific tend to represent transcripts that are expressed in a set of healthy tissues rather than novel isoforms.ConclusionsWe propose to combine the computational prediction of alternative splice isoforms with experimental validation for efficient delineation of an accurate set of tissue-specific transcripts.

Project description:Cyclic nucleotide phosphodiesterases (PDEs) hydrolyze the intracellular second messengers: cyclic adenosine monophosphate (cAMP) and cyclic guanine monophosphate (cGMP). The cAMP-specific PDE family 4 (PDE4) is widely expressed in vertebrates. Each of the four PDE4 gene isoforms (PDE4 A-D) undergo extensive alternative splicing via alternative transcription initiation sites, producing unique amino termini and yielding multiple splice variant forms from each gene isoform termed long, short, super-short and truncated super-short. Many species across the vertebrate lineage contain multiple splice variants of each gene type, which are characterized by length and amino termini.A phylogenetic approach was used to visualize splice variant form genesis and identify conserved splice variants (genome conservation with EST support) across the vertebrate taxa. Bayesian and maximum likelihood phylogenetic inference indicated PDE4 gene duplication occurred at the base of the vertebrate lineage and reveals additional gene duplications specific to the teleost lineage. Phylogenetic inference and PDE4 splice variant presence, or absence as determined by EST screens, were further supported by the genomic analysis of select vertebrate taxa. Two conserved PDE4 long form splice variants were found in each of the PDE4A, PDE4B, and PDE4C genes, and eight conserved long forms from the PDE4 D gene. Conserved short and super-short splice variants were found from each of the PDE4A, PDE4B, and PDE4 D genes, while truncated super-short variants were found from the PDE4C and PDE4 D genes. PDE4 long form splice variants were found in all taxa sampled (invertebrate through mammals); short, super-short, and truncated super-short are detected primarily in tetrapods and mammals, indicating an increasing complexity in both alternative splicing and cAMP metabolism through vertebrate evolution.There was a progressive independent incorporation of multiple PDE4 splice variant forms and amino termini, increasing PDE4 proteome complexity from primitive vertebrates to humans. While PDE4 gene isoform duplicates with limited alternative splicing were found in teleosts, an expansion of both PDE4 splice variant forms, and alternatively spliced amino termini predominantly occurs in mammals. Since amino termini have been linked to intracellular targeting of the PDE4 enzymes, the conservation of amino termini in PDE4 splice variants in evolution highlights the importance of compartmentalization of PDE4-mediated cAMP hydrolysis.

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An analysis of tissue-specific alternative splicing at the protein level.

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An analysis of tissue-specific alternative splicing at the protein level.

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