Alternative polyadenylation (APA) is a key RNA-processing mechanism that generates distinct 3' termini on mRNAs and other RNA polymerase II transcripts. It is widespread in eukaryotic species and plays a major role in gene regulation. APA is tissue-specific and important for cell proliferation and differentiation. This review discusses the roles of APA in various cellular processes, including mRNA metabolism, protein diversification, and protein localization. It also covers the molecular mechanisms underlying APA, such as variations in core processing factors and RNA-binding proteins, as well as transcription-based regulation. APA occurs most frequently in the 3' untranslated region (3' UTR) of mRNAs and is used in nearly all eukaryotic species. For example, at least 70% of mammalian mRNA-encoding genes express APA isoforms. APA can significantly affect post-transcriptional gene regulation through modulation of mRNA stability, translation, nuclear export, and protein localization. One remarkable feature of 3' UTR-APA is its global regulation, involving numerous transcripts in a cell. APA can influence microRNA (miRNA) functions by affecting the stability and/or translation of target mRNAs. miRNA target sites are often located in 3' UTRs. Differential targeting of 3' UTR-APA isoforms was first demonstrated in activated T cells and cancer cells. APA can also affect mRNA stability through destabilizing elements, such as AU-rich elements (AREs), GU-rich elements (GREs), and PUF protein-binding elements. Long 3' UTRs are generally less stable than short ones, but this view has been challenged by genome-wide studies. APA can also affect mRNA translation, with long isoforms associated with more ribosomes than short ones. However, another study using human cells reported that 3' UTR length can suppress translation. APA can influence mRNA nuclear export and localization, with long 3' UTRs more abundant in the nucleus than in the cytoplasm. APA can also affect protein localization, with sequences in 3' UTRs regulating protein localization independently of mRNA localization. APA upstream of the last exon (UR-APA) can lead to the expression of alternative terminal exons and can result in changes to both the coding sequence and 3' UTR of an mRNA. UR-APA is generally upregulated in proliferating cells and suppressed during cell differentiation, mirroring the use of proximal PASs in 3' UTRs. UR-APA can affect gene expression in various ways, including the regulation of protein isoform switching. APA can lead to the expression of truncated proteins with dominant negative functions. For example, retinoblastoma-binding protein 6 (RBBP6) produces several isoforms through differential RNA processing. One of these isoforms, Iso3, is severely truncated and can compete with full-length RBBP6 for association with the polyadenylation machineryAlternative polyadenylation (APA) is a key RNA-processing mechanism that generates distinct 3' termini on mRNAs and other RNA polymerase II transcripts. It is widespread in eukaryotic species and plays a major role in gene regulation. APA is tissue-specific and important for cell proliferation and differentiation. This review discusses the roles of APA in various cellular processes, including mRNA metabolism, protein diversification, and protein localization. It also covers the molecular mechanisms underlying APA, such as variations in core processing factors and RNA-binding proteins, as well as transcription-based regulation. APA occurs most frequently in the 3' untranslated region (3' UTR) of mRNAs and is used in nearly all eukaryotic species. For example, at least 70% of mammalian mRNA-encoding genes express APA isoforms. APA can significantly affect post-transcriptional gene regulation through modulation of mRNA stability, translation, nuclear export, and protein localization. One remarkable feature of 3' UTR-APA is its global regulation, involving numerous transcripts in a cell. APA can influence microRNA (miRNA) functions by affecting the stability and/or translation of target mRNAs. miRNA target sites are often located in 3' UTRs. Differential targeting of 3' UTR-APA isoforms was first demonstrated in activated T cells and cancer cells. APA can also affect mRNA stability through destabilizing elements, such as AU-rich elements (AREs), GU-rich elements (GREs), and PUF protein-binding elements. Long 3' UTRs are generally less stable than short ones, but this view has been challenged by genome-wide studies. APA can also affect mRNA translation, with long isoforms associated with more ribosomes than short ones. However, another study using human cells reported that 3' UTR length can suppress translation. APA can influence mRNA nuclear export and localization, with long 3' UTRs more abundant in the nucleus than in the cytoplasm. APA can also affect protein localization, with sequences in 3' UTRs regulating protein localization independently of mRNA localization. APA upstream of the last exon (UR-APA) can lead to the expression of alternative terminal exons and can result in changes to both the coding sequence and 3' UTR of an mRNA. UR-APA is generally upregulated in proliferating cells and suppressed during cell differentiation, mirroring the use of proximal PASs in 3' UTRs. UR-APA can affect gene expression in various ways, including the regulation of protein isoform switching. APA can lead to the expression of truncated proteins with dominant negative functions. For example, retinoblastoma-binding protein 6 (RBBP6) produces several isoforms through differential RNA processing. One of these isoforms, Iso3, is severely truncated and can compete with full-length RBBP6 for association with the polyadenylation machinery