REVIEWS Post-transcriptional regulation of gene expression in innate immunity Susan Carpenter1*, Emiliano P. Ricci2*, Blandine C. Mercier2*, Melissa J. Moore2 and Katherine A. Fitzgerald1,3 Abstract | Innate immune responses combat infectious microorganisms by inducing inflammatory responses, antimicrobial pathways and adaptive immunity. Multiple genes within each of these functional categories are coordinately and temporally regulated in response to distinct external stimuli. The substantial potential of these responses to drive pathological inflammation and tissue damage highlights the need for rigorous control of these responses. Although transcriptional control of inflammatory gene expression has been studied extensively, the importance of post-transcriptional regulation of these processes is less well defined. In this Review, we discuss the regulatory mechanisms that occur at the level of mRNA splicing, mRNA polyadenylation, mRNA stability and protein translation, and that have instrumental roles in controlling both the magnitude and duration of the inflammatory response. A dynamic and coordinately regulated gene expression specific signal that is encountered. Although transcrip- programme lies at the heart of the inflammatory process. tion is an essential first step, and certainly the most well- This response endows the host with a first line of defence scrutinized area in studies of innate immunity5,6, proper against infection and the capacity to repair and restore regulation of immune genes also involves a plethora of damaged tissues. However, unchecked, prolonged or additional post-transcriptional checkpoints. These occur inappropriately scaled inflammation can be detrimental at the level of mRNA splicing, mRNA polyadenylation, to the host and lead to diseases such as atherosclerosis, mRNA stability and protein translation. Many of these arthritis and cancer1,2. mechanisms are particularly important for modulating 1Program in Innate Immunity, Division of Infectious The acute inflammatory programme is initiated the strength and duration of the response and for turn- Diseases and Immunology, when germline-encoded pattern recognition receptors ing the system off in a timely and efficient manner. In Department of Medicine, (PRRs) that are present in distinct cellular compart- this Review, we cover exciting recent developments in this University of Massachusetts ments respond to signs of microbial infection3,4. Once under­explored area. We also highlight the emerging role Medical School, Worcester, Massachusetts 01605, USA. activated, these receptors trigger signalling cascades of long non-coding RNAs (lncRNAs) in controlling the 2Howard Hughes Medical that converge on well-defined transcription factors. inflammatory response. A better understanding of these Institute, University of Mobilization of these factors leads to rapid, dynamic processes could facilitate the development of selective Massachusetts Medical and temporally regulated changes in the expression of therapeutics to prevent damaging inflammation. School, Worcester, hundreds of genes that are involved in antimicrobial Massachusetts 01605, USA. 3Centre of Molecular defence, phagocytosis, cell migration, tissue repair and Alternative splicing in innate immunity Inflammation Research, the regulation of adaptive immunity. Although transcriptional regulation has been at the Department of Cancer Multiple genes within distinct functional categories are forefront of studies of innate immunity, the role of post- Research and Molecular coordinately and temporally regulated by transcriptional transcriptional regulation in controlling gene expres- Medicine, Norwegian University of Science and ‘on’ and ‘off’ switches that account for the specificity of sion in macrophages and other innate immune cells is Technology, 7491 Trondheim, gene expression in response to external stimuli. Multiple equally important. Almost one-fifth of the genes that Norway. layers of regulation — including chromatin state, histone are expressed in human dendritic cells (DCs) undergo *These authors contributed or DNA modifications, and the recruitment of transcrip- alternative splicing upon bacterial challenge. Most of equally to this work. tion factors and of the basal transcription machinery — these genes are involved in general cellular functions Correspondence to K.A.F. 7 e‑mail: kate.fitzgerald@ collaborate to control these pathogen-induced or danger but some participate directly in antimicrobial defence . 5,6 umassmed.edu signal-induced gene expression programmes , which Furthermore, stimulation of human monocytes with the doi:10.1038/nri3682 vary depending on the cell lineage involved and the Toll-like receptor 4 (TLR4) ligand lipopolysaccharide NATURE REVIEWS | IMMUNOLOGY VOLUME 14 | JUNE 2014 | 361 © 2014 Macmillan Publishers Limited. All rights reserved REVIEWS Spliced intron a Mechanism of transcript intron splicing U1 NTC U2 U2 U4 U1 U6 U5 U2 U6 U4 U5 NTC Exon junction 5′ Exon U1 U2 Exon 3′ U6 U5 complex Intron Transcript Splice site definition Exon Exon Spliceosome Splicing assembly b Mechanism of transcript cleavage and polyadenylation CSTF CPSF CSTF Downstream 5′ 3′ sequence elements Transcript Cleavage CPSF PAP site 5′ Poly(A) Downstream AAAAAAAA signal sequence elements Transcript Cleavage 3′ Poly(A) site signal Figure 1 | Pre-mRNA processing into mature mRNAs: intron splicing and polyadenylation. a | Following transcription, pre-mRNA intronic sequences are removed by splicing. The 5ʹ and 3ʹ splice sitesNature of introns Reviews are recognized | Immunology by the small nuclear ribonucleoproteins (snRNPs) U1 and U2, respectively, then the spliceosome assembles and catalyses the excision of the introns and ligation of the flanking exons. A multi-protein complex, the exon junction complex, is deposited on exon–exon junctions. b | A poly(A) tail is also added to the 3ʹ end of transcripts. The poly(A) signal and nearby U-rich or GU‑rich downstream sequence elements are recognized by two multi-protein complexes — namely, cleavage and polyadenylation specificity factor (CPSF) and cleavage stimulating factor (CSTF), respectively — that promote endonucleolytic cleavage of the transcript. Poly(A) polymerase (PAP) catalyses the subsequent addition of a stretch of adenosines from the cleavage site. Pattern recognition (LPS) and with interferon‑γ (IFNγ) causes the poly­ of 5ʹ or 3ʹ splice sites at intron ends. Alternative poly­ receptors adenylation machinery to favour proximal poly(A) site adenylation within an intron can also generate an mRNA (PRRs). Host receptors (such as use in terminal exons that contain two or more poly(A) that encodes a truncated protein product. However, Toll-like receptors (TLRs) or sites8. This type of alternative polyadenylation leads to a alternative processing is by no means limited to internal NOD-like receptors (NLRs)) that can sense pathogen- global shortening of 3ʹ untranslated regions (UTRs) and a sites. Alternative promoter use results in alternative first associated molecular patterns loss of key regulatory elements such as microRNA (miRNA) exons, which changes the length and sequence of the and initiate signalling cascades target sites and AU‑rich elements (AREs). 5ʹ UTR. Similarly, alternative polyadenylation within that lead to an innate immune the last exon can shorten or extend the 3ʹ UTR11 (FIG. 2a). response. These can be Alternative pre-mRNA processing. Following tran- Modifications to UTRs have important consequences membrane-bound (for example, TLRs) or soluble scription, pre-mRNA intronic sequences are removed because they can affect sequences that regulate sub- cytoplasmic receptors (for by splicing. The 5ʹ and 3ʹ splice sites of introns are cellular mRNA localization, translation efficiency and example, retinoic acid- recognized by the small nuclear ribonucleic particles mRNA stability12. inducible protein I (RIG‑I), (snRNPs) U1 and U2, respectively, before the spliceo- melanoma differentiation- associated protein 5 (MDA5) some assembles and catalyses excision of the introns Regulation of TLR signalling by alternative splicing and 9 and NLRs). and the ligation of flanking exons (FIG. 1a). In addi- alternative polyadenylation. The TLR signalling pathway tion, a poly(A) tail is added to the 3ʹ end of transcripts. is subject to extensive post-transcriptional regulation, in microRNA A poly(A) signal and nearby U-rich or GU‑rich down- which more than 256 alternatively processed transcripts (miRNA). Non-coding RNA (21 stream sequence elements (DSEs) are recognized by encode variants of receptors, adaptors and signalling nucleotides in length) that is 13 encoded in the genomes of two multi-protein complexes — namely, cleavage and molecules . Every TLR gene has numerous alternatively 13–18 animals and plants. miRNAs polyadenylation specificity factor (CPSF) and cleavage spliced variants , and TLR1 to TLR7 all have between regulate gene expression by stimulation factor (CSTF), respectively — that promote two and four predicted alternative polyadenylation sites16. binding to the 3ʹ untranslated endonucleolytic cleavage of the pre-mRNAs. Poly(A) These variant transcripts have myriad effects on signal region of target mRNAs. polymerase (PAP; also known as PAPα and PAPOLA) transduction. For example, an alternatively spliced form AU‑rich elements subsequently catalyses the addition of a stretch of of mouse Tlr4 mRNA includes an exon that is not pre- (AREs). Regulatory elements adenosines from the cleavage site10 (FIG. 1b). sent in the canonical mRNA15.