AbstractFunctioning as an essential step of pre‐mRNA processing, polyadenylation has been realized in recent years to play an important regulatory role during eukaryotic gene expression. Such regulation occurs mostly through the use of alternative polyadenylation (APA) sites and generates different transcripts with altered coding capacity for proteins and/or RNA. However, the molecular mechanisms that underlie APAs are poorly understood. Besides APA cases demonstrated in animal embryo development, cancers, and other diseases, there are a number of APA examples reported in plants. The best‐known ones are related to flowering time control pathways and stress responses. Genome‐wide studies have revealed that plants use APA extensively to generate diversity in their transcriptomes. Although each transcript produced by RNA polymerase II has a poly(A) tail, over 50% of plant genes studied possess multiple APA sites in their transcripts. The signals defining poly(A) sites in plants were mostly studied through classical genetic means. Our understanding of these poly(A) signals is enhanced by the tallies of whole plant transcriptomes. The profiles of these signals have been used to build computer models that can predict poly(A) sites in newly sequenced genomes, potential APA sites in genes of interest, and/or to identify, and then mutate, unwanted poly(A) sites in target transgenes to facilitate crop improvements. In this review, we provide readers an update on recent research advances that shed light on the understanding of polyadenylation, APA, and its role in gene expression regulation in plants. WIREs RNA 2011 2 445–458 DOI: 10.1002/wrna.59This article is categorized under: RNA Processing > Splicing Regulation/Alternative Splicing RNA Processing > 3' End Processing