Massive expansions of Dscam splicing diversity via staggered homologous recombination during arthropod evolution

Abstract

The arthropod Down syndrome cell adhesion molecule (Dscam) gene can generate tens of thousands of protein isoforms via combinatorial splicing of numerous alternative exons encoding immunoglobulin variable domains organized into three clusters referred to as the exon 4, 6, and 9 clusters. Dscam protein diversity is important for nervous system development and immune functions. We have performed extensive phylogenetic analyses of Dscam from 20 arthropods (each containing between 46 and 96 alternative exons) to reconstruct the detailed history of exon duplication and loss events that built this remarkable system over 450 million years of evolution. Whereas the structure of the exon 4 cluster is ancient, the exon 6 and 9 clusters have undergone massive, independent expansions in each insect lineage. An analysis of nearly 2000 duplicated exons enabled detailed reconstruction of the timing, location, and boundaries of these duplication events. These data clearly show that new Dscam exons have arisen continuously throughout arthropod evolution and that this process is still occurring in the exon 6 and 9 clusters. Recently duplicated regions display boundaries corresponding to a single exon and the adjacent intron. The boundaries, homology, location, clustering, and relative frequencies of these duplication events strongly suggest that staggered homologous recombination is the major mechanism by which new Dscam exons evolve. These data provide a remarkably detailed picture of how complex gene structure evolves and reveal the molecular mechanism behind this process.

The landscape of human mutually exclusive splicing

“…It contains a cluster of MXEs (exons 3b–3e) that is differentially included into a set of two exons (exon 3 and exon 4), and the two sets are themselves mutually exclusive (Fig 1F, Appendix Figs S16 and S17). The identification of large clusters with multiple MXEs and many genes with multiple clusters shows that complex mutually exclusive splicing is not restricted to arthropods (Schmucker et al , 2000; Graveley, 2005; Lee et al , 2010; Hatje & Kollmar, 2013) but might be present in all bilateria.…”

“…Mutually exclusive splicing generates alternative isoforms by retaining only one exon of a cluster of neighbouring internal exons in the mature transcript and is a sophisticated way to modulate protein function (Letunic et al , 2002; Meijers et al , 2007; Pohl et al , 2013; Tress et al , 2017a). The most extreme cases known so far are the arthropod DSCAM genes, for which up to 99 mutually exclusive exons (MXEs) spread into four clusters were identified (Schmucker et al , 2000; Lee et al , 2010; Pillmann et al , 2011). …”

Alternative splicing ofU2AF1reveals a shared repression mechanism for duplicated exons

“…The majority of invertebrate tandem exon duplications associated with mutually exclusive splicing resulted from homologous recombination (HR) events that engaged the upstream intron in each case (73). A HR-mediated duplication of the U2AF1 intron 2–exon 3 segment (Figure 5A) would also explain the mutually exclusive splicing of exons Ab and 3 as well as the existence of the two regions of vertebrate conservation, which are located at a similar distance from their 3′ss (Figure 3A) and share significant sequence identity (Supplementary Figure S8).…”

Penaeus monodon Dscam (PmDscam) has a highly diverse cytoplasmic tail and is the first membrane-bound shrimp Dscam to be reported

“…Finally, even though most L. vannamei genes (including Dscam) show a high homology to the corresponding P. monodon gene, the expression of more highly diverse PmDscam alternative sequences in P. monodon might still be due to species difference. A comparison of 20 arthropod Dscam genomic sequences found that the number of alternative exons within many exon clusters varied significantly [17]. For instance, in the exon cluster encoding N-terminal Ig3 domain, the number of alternative sequences ranges from 22 to 52.…”