De novo and rare inherited mutations implicate the transcriptional coregulator TCF20/SPBP in autism spectrum disorder.
Bottom Line: IMGSAC family screening identified a de novo missense mutation of TCF20 in a single case and significant association of a different missense mutation of TCF20 with ASD in three further families.We did not identify a significant association of TNRC6B mutations with ASD.This study provides the first evidence that mutations in TCF20 are also associated with ASD.
Affiliation: Weatherall Institute of Molecular Medicine, John Radcliffe Hospital, Oxford, UK NIHR Biomedical Research Centre, Oxford, UK.Show MeSH
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Mentions: To characterise the molecular nature of the pericentric inversion, we performed FISH using multiple BACs and fosmids (table 1). These probes were initially focused on the 22q13.1 band in which the long-arm breakpoint had been tentatively located, but several further rounds of analysis were performed as greater complexity in the rearrangement became apparent (figure 2). The observation of split signals with two fosmids localised one breakpoint (termed breakpoint A) to a ∼37 kb region (figure 2A). Further analysis by Southern blotting with single-copy probes identified breakpoint fragments, initially within a ∼15 kb EcoRI fragment, and subsequently within a 248 bp fragment bordered by StuI and AflIII restriction sites (not shown). PCR primers were designed to amplify across the breakpoint in sequentially nested amplifications with degenerate primers (see ‘Methods’). Surprisingly, DNA sequencing of this amplification product identified the sequence on the centromeric side of the break as originating from a location ∼1.9 Mb centromeric of breakpoint A (figure 2D, bottom right). These sequence data showed contiguity between nucleotides at coordinates at 40 709 620 bp (breakpoint B) and 42 634 698 bp (breakpoint A), adjacent to a short stretch of 5-nucleotide (5′-GACCT-3′) complementarity (figure 2D). Confirming the identification of breakpoint B, clones closely adjacent on either side of this location mapped to opposite arms of the der(22) (figure 2B). This result implied that a third more centromeric break on the long arm (‘breakpoint C’) must have occurred, to which the intermediate segment (B-A) had been joined. This break was localised using FISH to an ∼44.7 kb region within BAC clone W12-1570N6 (figure 2C). Analysis by Southern blotting revealed a HindIII restriction fragment that likely spanned the breakpoint (figure 2D, bottom left), locating the breakpoint to a ∼4 kb region between 39 507 139 and 39 511 083 bp. Figure 2D summarises the structure of the derivative chromosome 22 as concluded from the FISH, Southern blotting and DNA sequencing results. Breakpoint D is predicted to occur in the short arm satellite sequence of chromosome 22 and was not characterised further. Although (as demonstrated by array CGH) there has been no major gain or loss of material at the breakpoints, we found evidence of a small (∼10 kb) duplication at breakpoint A (data not shown) and this may apply to others too, most consistent with the replication-based fork stalling template switching (FoSTeS)-type mechanism for the complex chromosome rearrangement.29
Affiliation: Weatherall Institute of Molecular Medicine, John Radcliffe Hospital, Oxford, UK NIHR Biomedical Research Centre, Oxford, UK.