Alternative polyadenylation of mRNA precursors

Abstract

Alternative polyadenylation (APA) is an RNA-processing mechanism that generates distinct 3' termini on mRNAs and other RNA polymerase II transcripts. It is widespread across all eukaryotic species and is recognized as a major mechanism of gene regulation. APA exhibits tissue specificity and is important for cell proliferation and differentiation. In this Review, we discuss the roles of APA in diverse cellular processes, including mRNA metabolism, protein diversification and protein localization, and more generally in gene regulation. We also discuss the molecular mechanisms underlying APA, such as variation in the concentration of core processing factors and RNA-binding proteins, as well as transcription-based regulation.

Figure 1. 3′ UTR-APA

a | Alternative polyadenylation (APA) leading to the production of two mRNA isoforms with different 3′ untranslated regions (3′ UTRs) — termed 3′ UTR-APA here — is shown. The 3′ UTR region upstream of the proximal polyadenylation site (PAS) is found in both short (top) and long (bottom) isoforms and is denoted the constitutive UTR (cUTR), whereas the downstream region is present in the long isoform only and is termed the alternative UTR (aUTR). Interactions between the aUTR and RNA-binding proteins (RBPs), microRNAs (miRNAs) and long non-coding RNAs (lncRNAs) can have various functional consequences. The poly(A) tail is represented as AAA. b | In the case of the gene encoding human IFN-regulatory factor 5 (IRF5), APA of the transcript produces a long 3′ UTR isoform that is more rapidly degraded owing to the presence of a destabilizing AU-rich element (ARE) in the aUTR. The ARE and cytoplasmic exosome mediate mRNA decay. In patients with systemic lupus erythematosus (SLE), a single nucleotide polymorphism reducing the use of the proximal PAS leads to the production of long isoforms at the expense of short isoforms, which results in reduced IRF5 levels. c | Differential mRNA localization of brain-derived neurotrophic factor (BDNF) 3′ UTR-APA isoforms in neurons. The long isoform localizes to dendrites more than the short isoform, and this supports dendrite-localized protein synthesis. d | Differential localization of the transmembrane CD47 proteins encoded by long or short APA isoforms. Both isoforms are translated on the ER membrane. The aUTR of the long isoform is bound by the RBP Hu antigen R (HUR), which leads to the localization of CD47 protein to the cell membrane through a cascade of interactions (dashed arrow) involving the phosphatase 2A inhibitor SET and RAC1. The protein generated from the short isoform remains in the ER. CDS, coding sequence.

Figure 2. UR-APA

a | Alternative polyadenylation (APA) in upstream regions (URs) of mRNAs — termed UR-APA here — can lead to the production of isoforms with different 3′-terminal exons and, hence, different coding sequences and 3′ untranslated regions (3′ UTRs). Three isoforms are shown, with their respective terminal exon types indicated. Splicing is indicated by a dashed line. The ‘canonical’ isoform (top) is formed by the use of the polyadenylation site (PAS) in the 3′-most exon. The use of a PAS in an alternative exon that is excluded from the canonical isoform generates a transcript containing a skipped terminal exon (middle). Inhibition of splicing at the indicated 5′ splicing site (5′SS) results in the inclusion of part of the downstream intron and use of a PAS within that intron; such a transcript is described as containing a composite terminal exon (bottom). Regions not present in the canonical isoform are shown in red. The functional consequences of UR-APA are indicated. b | UR-APA of the transcript encoding polyadenylation factor retinoblastoma-binding protein 6 (RBBP6) produces an isoform encoding a dominant negative protein, Iso3. c | UR-APA of the mRNA encoding polyadenylation factor cleavage stimulation factor 77 kDa subunit (CSTF77) produces a short isoform that encodes a truncated protein with no apparent functions (not shown). The full-length protein activates the usage of the upstream PAS, thereby increasing the levels of the short mRNA and forming a negative feedback loop. CDS, coding sequence.

Figure 3. Regulation of APA

a | The choice of polyadenylation site (PAS) during alternative polyadenylation (APA) can be influenced by various factors, including the gene promoter at the transcription start site (TSS); recruitment of polyadenylation factors directly or of proteins that influence PAS choice; nucleosome density in the region around the PAS; RNA polymerase II (Pol II)-mediated transcription elongation by the Pol II-associated factor (PAF) complex; the function of various RNA-binding proteins (RBPs) associated with the nascent transcript; the presence of N⁶-methyladenosine (m⁶A); and inhibition of polyadenylation by the splicing factor U1 small nuclear ribonucleoprotein (U1 snRNP). See the main text for more details. b | A proposed model for the regulation of APA by the cleavage factor I (CFI) complex. Two UGUA elements upstream and downstream of a proximal PAS are recognized by the heterodimeric CFI complex, which consists of CFI 68 kDa subunit (CFI68) and CFI25, leading to skipping of the PAS. c | Regulation of neuronal APA in Drosophila melanogaster by the RBP Embryonic-lethal abnormal visual (Elav). Elav is recruited to Pol II at promoter regions that contain a GAGA sequence, which can cause Pol II pausing. Elav inhibits proximal PAS usage, leading to the expression of long APA isoforms during neurogenesis. PA complex, polyadenylation complex.