Date Published: November 19, 2009
Publisher: Public Library of Science
Author(s): Yevgeniy A. Grigoryev, Sunil M. Kurian, Aleksey A. Nakorchevskiy, John P. Burke, Daniel Campbell, Steve R. Head, Jun Deng, Aaron B. Kantor, John R. Yates, Daniel R. Salomon, Juan Valcarcel. http://doi.org/10.1371/journal.pone.0007906
Abstract: Alternative splicing of pre-mRNA is a mechanism that increases the protein diversity of a single gene by differential exon inclusion/exclusion during post-transcriptional processing. While alternative splicing is established to occur during lymphocyte activation, little is known about the role it plays during the immune response. Our study is among the first reports of a systematic genome-wide analysis of activated human T and B lymphocytes using whole exon DNA microarrays integrating alternative splicing and differential gene expression. Purified human CD2+ T or CD19+ B cells were activated using protocols to model the early events in post-transplant allograft immunity and sampled as a function of time during the process of immune activation. Here we show that 3 distinct classes of alternatively spliced and/or differentially expressed genes change in an ordered manner as a function of immune activation. We mapped our results to function-based canonical pathways and demonstrated that some are populated by only one class of genes, like integrin signaling, while other pathways, such as purine metabolism and T cell receptor signaling, are populated by all three classes of genes. Our studies augment the current view of T and B cell activation in immunity that has been based exclusively upon differential gene expression by providing evidence for a large number of molecular networks populated as a function of time and activation by alternatively spliced genes, many of which are constitutively expressed.
Partial Text: The technology for investigating gene expression in cells and tissues has developed significantly over the last decade, making global gene expression profiling using microarrays relatively straightforward. However, until recently, the field has concentrated largely on studies of differential gene expression and discovering signatures that correlate with various biological challenges or disease states. Unfortunately, the premise of analyzing differential gene expression is limited by the view that molecular mechanisms or biomarkers are represented by classes of genes either up- or down-regulated in a given situation. Alternative splicing (AS) is a process by which a single pre-mRNA transcript can give rise to multiple protein isoforms through the mechanism of coordinated intron removal and differential exon joining. AS is a major source of diversity in the human proteome; as many as 75% ,  of all human genes are alternatively spliced and the most recent study using next-generation sequencing technology indicates that 92–94% of human genes undergo alternative splicing , . Splicing can modulate protein function by changing functional domains, affinities for assembly of heteromeric complexes, or altering mRNA stability. The advent of high-throughput genomics has dramatically changed the view of alternative splicing from a single gene perspective to the level of genome-wide discovery and quantification.
Despite many recent reports on alternative splicing, studies of genome-wide alternative splicing in the immune system are still limited and, while alternative splicing (AS) is established to occur during lymphocyte activation, little is known about the role AS plays or its impact on immunity. Alternative splicing of pre-mRNA is a mechanism that increases the protein repertoire of a single gene sequence by including or excluding exons during post-transcriptional processing. There is evidence in different model systems for alternative splicing of as many as 75% to 94% of all human genes , , . Splicing can modulate protein function by changing functional domains, affinities for assembly of heteromeric complexes, or altering mRNA stability.