Alternative splicing in concert with protein intrinsic disorder enables increased functional diversity in multicellular organisms

Alternative splicing of pre-mRNA generates two or more protein isoforms from a single gene, thereby contributing to protein diversity. Despite intensive efforts, an understanding of the protein structure-function implications of alternative splicing is still lacking. Intrinsic disorder, which is a lack of equilibrium 3D structure under physiological conditions, may provide this understanding. Intrinsic disorder is a common phenomenon, particularly in multicellular eukaryotes, and is responsible for important protein functions including regulation and signaling. We hypothesize that polypeptide segments affected by alternative splicing are most often intrinsically disordered such that alternative splicing enables functional and regulatory diversity while avoiding structural complications. We analyzed a set of 46 differentially spliced genes encoding experimentally characterized human proteins containing both structured and intrinsically disordered amino acid segments. We show that 81% of 75 alternatively spliced fragments in these proteins were associated with fully (57%) or partially (24%) disordered protein regions. Regions affected by alternative splicing were significantly biased toward encoding disordered residues, with a vanishingly small P value. A larger data set composed of 558 SwissProt proteins with known isoforms produced by 1,266 alternatively spliced fragments was characterized by applying the PONDR VSL1 disorder predictor. Results from prediction data are consistent with those obtained from experimental data, further supporting the proposed hypothesis. Associating alternative splicing with protein disorder enables the time- and tissue-specific modulation of protein function needed for cell differentiation and the evolution of multicellular organisms.