Special Issue: 40 Years of TiBS Review The Spliceosome: The Ultimate RNA Chaperone and Sculptor Panagiotis Papasaikas 1,2 and Juan Valcárcel 1,2,3, * The spliceosome, one of the most complex machineries of eukaryotic cells, removes intronic sequences from primary transcripts to generate functional messenger and long noncoding RNAs (lncRNA). Genetic, biochemical, and struc- tural data reveal that the spliceosome is an RNA-based enzyme. Striking mecha- nistic and structural similarities strongly argue that pre-mRNA introns originated from self-catalytic group II ribozymes. However, in the spliceosome, protein components organize and activate the catalytic-site RNAs, and recognize and pair together splice sites at intron boundaries. The spliceosome is a dynamic, reversible, and flexible machine that chaperones small nuclear (sn) RNAs and a variety of pre-mRNA sequences into conformations that enable intron removal. This malleability likely contributes to the regulation of alternative splicing, a prevalent process contributing to cell differentiation, homeostasis, and disease. ‘and deem not profitless those fleeting moods’ W. Wordsworth, The Prelude In the Beginning Were Introns Introns are intervening sequences that are removed from primary transcripts by the process of RNA splicing, which links together the flanking pieces (exons) to generate functional mature RNAs (Figure 1A) [1]. Out of the 16 h required to transcribe the human dystrophin gene, 15 h and 54 min are devoted to the seemingly useless synthesis of intronic RNA. Therefore, efficient removal of introns, with single nucleotide precision, is essential for eukaryotic cells to produce the correct complement of mRNA and lncRNA transcripts necessary to deploy genetic pro- grams. Furthermore, ambiguity in defining certain introns, exons, and their precise boundaries allows alternative patterns of intron removal that greatly expand the diversity of transcripts generated per gene in multicellular organisms [2]. More than 90% of human genes are alterna- tively spliced, providing extensive opportunities for gene regulation during development, cell differentiation, and homeostasis [3]. For instance, alternative-splicing regulation is instrumental to control cellular and organ functions ranging from switches in glycolysis or cell division to building neural connectivity or modulating pain. Alterations in pre-mRNA splicing are also at the basis of a substantial number of genetic diseases, from spinal muscular atrophy, myotonic dystrophy, or familial dysautonomia to premature aging [4]. Alternatively spliced transcripts often differ in their coding capacity, stability, or translational efficiency [5]. A striking example is provided by the recent discovery of microexons encoding as few as one to five amino acids, whose inclusion is precisely regulated during neural differentiation and can modulate protein–protein interactions [6]. Alternatively spliced transcripts can also act Trends The spliceosome is a ribonucleoprotein complex that chaperones pre-mRNA and small nuclear (sn)RNAs into con- formations resembling the RNA-based catalytic core of group II introns. Proteins such as pre-mRNA-proces- sing-splicing factor 8 (Prp8) organize and regulate the activation of the RNA-based catalytic core of the spliceosome. The catalytic site is open to accommo- date different intron lengths and sequences, and to interact with other cellular machineries. Spliceosomal subcomplexes recog- nize and pair splice-site sequences in a highly dynamic and reversible fashion. This architecture, together with iterative substrate recognition and proofread- ing, provide opportunities for alterna- tive splicing regulation. 1 Centre de Regulació Genòmica, The Barcelona Institute of Science and Technology, Dr. Aiguader 88, 08003 Barcelona, Spain 2 Universitat Pompeu-Fabra, Dr. Aiguader 88, 08003 Barcelona, Spain 3 ICREA, Passeig Lluis Companys 23, 08010 Barcelona, Spain *Correspondence: juan.valcarcel@crg.eu (J. Valcárcel). Trends in Biochemical Sciences, January 2016, Vol. 41, No. 1 http://dx.doi.org/10.1016/j.tibs.2015.11.003 33 © 2015 Elsevier Ltd. All rights reserved.