Genome-wide identification of alternative splicing events that regulate protein transport across the secretory pathway

Abstract

Alternative splicing (AS) strongly increases proteome diversity and functionality in eukaryotic cells. Protein secretion is a tightly controlled process, especially when it occurs in a tissue-specific and differentiation-dependent manner. While previous work has focussed on transcriptional and post-translational regulatory mechanisms, the impact of AS on the secretory pathway remains largely unexplored. Here, we integrate results from a published screen for modulators of protein transport and RNA-Seq analyses to identify over 200 AS events as secretion regulators. We confirm that splicing events along all stages of the secretory pathway regulate the efficiency of membrane trafficking using morpholino and CRISPR/Cas9 experiments. We furthermore show that these events are highly tissue-specific and mediate an adaptation of the secretory pathway during T-cell activation and adipocyte differentiation. Our data substantially advance the understanding of AS functionality, add a new regulatory layer to a fundamental cell biological process and provide a resource of alternative isoforms that control the secretory pathway.

In silico analysis of alternative splicing events implicated in intracellular trafficking during B-lymphocyte differentiation

“…Interestingly, the expression of COPII components during T cell activation stays rather constant, whereas PCs show a substantial upregulation of almost all COPII components when compared to non-antibody-secreting prePB. These data suggest that B cells use changes in gene expression to adjust their secretory machinery during activation and differentiation, whereas T cells rely mostly on alternative splicing of key regulators ( 34 , 35 ). This may also indicate that B cells require stronger adjustments to their secretory system than T cells, which would make B cell differentiation a very interesting model system to study the adaptation of the secretory pathway in the future.…”

“…These analyses together indicate that alternative splicing controls diverse trafficking pathways during B cell activation and differentiation. Furthermore, the adaptation of intracellular trafficking in B cell subsets is different from the situation observed during T cell activation, which largely relies on alternative splicing to control their secretory capacity ( 34 , 35 ). In addition, AS events that change in COPII genes during T-Lymphocyte activation and B-Lymphocyte differentiation are mostly different, pointing to largely different strategies to adapt the secretory machinery to changing conditions.…”

“…Genes down-regulated in plasma cells are shown in blue (up-regulated in memory). (G) Comparison of gene expression levels of COPII-components in PC compared to prePB and T-cell activation [T-cell data was taken from( 34 )].…”

“…To better understand the mechanism that allows PCs to regulate their secretory machinery, we compared the expression level changes of COPII components in PCs and during the activation of CD4+ T cells ( Figure 2G ) ( 34 ). Interestingly, the expression of COPII components during T cell activation stays rather constant, whereas PCs show a substantial upregulation of almost all COPII components when compared to non-antibody-secreting prePB.…”

A Computational Analysis of Alternative Splicing across Mammalian Tissues Reveals Circadian and Ultradian Rhythms in Splicing Events

“…In addition, Golgi vesicle transport was found to be enriched for both the set of candidate genes with 24-h and with 12-h rhythmic phase-shifted isoforms, suggesting a complex temporal regulation of the secretory pathway that may possibly be regulated via circadian and ultradian splice isoform production. AS events have recently been shown to affect the secretory pathway [53] and a circadian control of procollagen transport from ER-to-Golgi and Golgi-to-plasma membrane has been reported in mice [54]. Among the genes with 24-h rhythmic phase-shifted isoform pairs in multiple baboon tissues, we identified several oncogenes and cancer-related genes, including PCBP2 , NELFE , CIRBP , and RACK1 .…”