The human genome contains approx. 25'000 protein-coding transcription units, but the number of specified proteins is considerably higher. This increase in protein diversity is mainly due to alternative splicing which may occur in as many as 95% of all human genes (1,2). Considering its importance, it is not surprising that defects in splicing can generate aberrant transcripts with diverse pathologic consequences.
Whether or not an exon is included in the final mRNA partly depends on the 5' and 3' splice sites (SS) and the branch points (BP). However, as these cis-acting sequences are highly degenerate (Fig. 1A), many sequences within a transcription unit resemble SS, but only a minority of them are true ones. The distinction between true and false (or cryptic) signals is due to additional regulatory sequences (Fig. 1B). In a simplified scheme, positive regulatory sequences are called exonic or intronic splicing enhancers (ESE or ISE) and are recognised by a class of proteins termed SR-proteins (containing a domain rich in serine and arginine residues). In contrast negative elements, so-called exonic and intronic splicing silencers (ESS and ISS), are recognised by members of the inhibitory hnRNP protein family (3).
Alternative splicing results when the interplay of all the cis-acting sequences and trans-acting factors can be interpreted in different ways. For many transcription units, multiple splicing isoforms can coexist as mature mRNAs in a given cell. However, SS and exon choices can also vary in a given cell depending on physiological conditions. Most important are changes in splicing patterns that occur during development while cells differentiate. Some of these differences may persist and establish long-lasting tissue- or cell type-specific patterns of splicing. Alternative splicing is particularly important for the development and differentiation of complex tissues such as the nervous system.
Changes in the RNA processing machinery may lead to mis-splicing of multiple transcripts, while single-nucleotide alterations in splice sites or cis-acting splicing regulatory sites may lead to differences in the splicing of a single gene. A probabilistic analysis indicates that over 60% of human disease-causing mutations may affect splicing rather than directly affecting coding sequences (4). Abnormally spliced mRNAs are also found in a high proportion of cancer cells (5). These differences may result from changes in the relative amounts or the phosphorylation of trans-acting splicing factors.