Evolutionary scenario after exonization of sequences, from the ancestral constitutive to alternative and, occasionally, to novel constitutive splicing. Intensities of selection are indicated at the left and right bars. For evolution of alternative splice sites, the intensities of selection vary over time. Initially, inclusion of the novel exon might be slightly deleterious, neutral, or slightly advantageous. Subsequently, selection changes, perhaps during a period of positive selection to increasing levels of negative selection (bar at the left with a gradient from white to red). The numeration refers to intronic insertion of a transposed element (red box) (1), followed by acquisition of necessary components for alternative splicing (2). Under relaxed selective pressure, the exonized condition might revert or become fixed in different lineages (vertical double arrow). In an extreme case, (3), as shown for ZNF639, the novel alternative splice form might, over time, completely replace the ancestral constitutive splice form, thus representing a different constitutive form (bar at the right side, with gradients from black via white to red). However, the constitutive inclusion of the exonized sequence in ZNF639 could also have been acquired directly after acquisition of the necessary splice sites. Examples corresponding to different stages of exonization taken from previous works (Singer et al. 2004; Krull et al. 2005; Mola et al. 2007) or from the present work (bold letters) are presented in ovals.
