Widespread Shortening of 3’ Untranslated Regions and Increased Exon Inclusion Are Evolutionarily Conserved Features of Innate Immune Responses to Infection

Abstract

The contribution of pre-mRNA processing mechanisms to the regulation of immune responses remains poorly studied despite emerging examples of their role as regulators of immune defenses. We sought to investigate the role of mRNA processing in the cellular responses of human macrophages to live bacterial infections. Here, we used mRNA sequencing to quantify gene expression and isoform abundances in primary macrophages from 60 individuals, before and after infection with Listeria monocytogenes and Salmonella typhimurium. In response to both bacteria we identified thousands of genes that significantly change isoform usage in response to infection, characterized by an overall increase in isoform diversity after infection. In response to both bacteria, we found global shifts towards (i) the inclusion of cassette exons and (ii) shorter 3’ UTRs, with near-universal shifts towards usage of more upstream polyadenylation sites. Using complementary data collected in non-human primates, we show that these features are evolutionarily conserved among primates. Following infection, we identify candidate RNA processing factors whose expression is associated with individual-specific variation in isoform abundance. Finally, by profiling microRNA levels, we show that 3’ UTRs with reduced abundance after infection are significantly enriched for target sites for particular miRNAs. These results suggest that the pervasive usage of shorter 3’ UTRs is a mechanism for particular genes to evade repression by immune-activated miRNAs. Collectively, our results suggest that dynamic changes in RNA processing may play key roles in the regulation of innate immune responses.

Environmental influences on RNA processing: Biochemical, molecular and genetic regulators of cellular response

“…Read‐through transcripts may actually function to maintain open chromatin upon stress, suggesting a beneficial role for polyadenylation inhibition (Vilborg et al, ). This regulated defect in 3′ end processing, which is strongly associated with stress response, is similar to observations of 3′ UTR shortening seen in proliferating cells, immune response, and cancer cells (Mayr & Bartel, ; Pai et al, ; Sandberg et al, ).…”

miRNA length variation during macrophage stimulation confounds the interpretation of results: implications for miRNA quantification by RT-qPCR

“…We next searched the literature for other miRNAs reported to be decreased in mouse macrophages upon LPS stimulation, which might have been attributed to an amplification bias of the RT-qPCR assays of a longer minor isoform. Our analyses of miR-27a-3p in the human macrophages and mouse BMDM/BMDC data sets (Dueck et al 2014;Pai et al 2016), demonstrated that the stoichiometry of its isoforms was impacted by bacterial products, but that this did not substantially decrease the overall intracellular concentration of this miRNA family, as the 20 nt Table S4). Black circles highlight miRBase canonical isoforms, while stars refer to abundant isoforms when different from miRBase definition (seen in the right panel).…”

“…Since Seeley et al also reported an induction of miR-222 levels in human monocytes following bacterial infection (Seeley et al 2018), we next revisited an independent small RNA-seq data set of human monocyte-derived macrophages infected by Salmonella enterica serovar Typhimurium for 24 h (which also activates TLR4) (Pai et al 2016). Analyses of these samples at the isoform level revealed a shift of prevalent miR-222 isoforms from 24 to 22 nt, with an overall decrease of miR-222 molecules (Fig.…”

Inflammation-Induced Alternative Pre-mRNA Splicing in Mouse Alveolar Macrophages

“…Much of this occurs in a cell-type-specific and/or signal-induced manner. We and others, have shown that mouse and human macrophages exposed to inflammatory stimuli in vitro undergo substantial alternative pre-mRNA splicing (Beyer et al 2012;Bhatt et al 2012;de Bruin et al 2016;Haque et al 2018;Lin et al 2016;Liu et al 2018;O'Connor et al 2015;Pai et al 2016;Pandya-Jones et al 2013). This can have profound effects on the nature and extent of the inflammatory response (Lynch 2004;Schaub and Glasmacher 2017).…”

“…In a similar fashion, alternative pre-mRNA splicing has been shown to alter cellular metabolism (Clower et al 2010;Yang and Lu 2013;Satoh et al 2015). While inflammation-induced alternative pre-mRNA splicing in macrophages has been investigated on a genome-wide scale in vitro (Beyer et al 2012;Bhatt et al 2012;Lin et al 2016;O'Connor et al 2015;Pai et al 2016;Pandya-Jones et al 2013), to our knowledge it has not been investigated in vivo. Such an approach is necessary to fully appreciate the effect of in vivo physiological context on macrophage pre-mRNA splicing.…”