A similar principle may be applicable to ASD caused by defects in other genes. Consistent with this notion, many known ASD genes, such as neuroligins and neurexins, display GPCR & G Protein inhibitor a complex pattern of isoform-specific expression in brain ( Boucard et al., 2005; Südhof, 2008), with different isoforms having very distinct functions ( Chih et al., 2006). In the cases of neurexins, more than 1,000 isoforms have been reported ( Missler and Südhof, 1998). The expression of Shank3 isoforms is cell type-specific and developmentally regulated ( Lim et al., 1999; Maunakea et al., 2010). RNA in situ hybridization in rat brain using a single probe from exon 21 encoding the proline-rich domain showed

that Shank3 is widely expressed in all brain regions at a low level at birth but increases after 2 weeks of age in the striatum, hippocampus, cerebellum, and in layers 1 and 2 of the neocortex ( Böckers et al., 2001, 2004). Similar findings were reported in mouse brain using a probe from exon 21 encoding the proline-rich domain of mouse Shank3 ( Peça et al., 2011). Peak expression of Shank3 occurs at an important developmental stage of synaptic plasticity and experience-dependent circuit maturation ( Böckers et al.,

2004). These studies, however, have not defined the isoform-specific expression of Shank3, and thus the expression profile for different Shank3 isoforms and regulation of isoform-specific expression remain to be elucidated. To add further complexity, SHANK3 has five CpG islands across the gene and these CpG islands display brain-specific and cell-type-specific DNA methylation BMN 673 supplier ( Figure 1A; Beri et al., 2007; Ching et al.,

2005; Maunakea et al., 2010). Both DNA methylation and histone deacetylase inhibitors have been shown to modulate the isoform specific gene expression of Shank3 in cultured neurons ( Beri et al., 2007; Maunakea et al., 2010). Thus, in addition to alternate promoter use and mRNA splicing, epigenetic mechanisms such as DNA methylation and histone acetylation regulate the expression of the Shank3 gene in an isoform-specific manner. Multiple intragenic CpG islands are Tryptophan synthase also associated with SHANK1 and SHANK2 ( Figures 1B and 1C), but the role of these CpG islands in transcriptional regulation remains to be investigated. SHANK2 exhibits transcriptional regulation similar to SHANK3 ( Leblond et al., 2012). Specifically, SHANK2 has several isoforms driven by multiple promoters and alternative splicing of coding exons ( Figure 1B). The longest Shank2e isoform containing all five protein domains was initially reported as an epithelia-specific isoform in rat ( McWilliams et al., 2004). However, a recent report indicates that SHANK2E is also expressed in brain tissues in humans ( Leblond et al., 2012). Several short isoforms (SHANK2A, SHANK2B, and SHANK2C) are transcribed from downstream promoters (SHANK2A, SHANK2B) or result from alternative splicing (SHANK2C) that contain distinct combinations of protein domains ( Figure 2C).