Novel TBX5 mutations and molecular mechanism for Holt-Oram syndrome.

The Holt-Oram syndrome (OMIM 142900) is an autosomal dominant disorder with clinical features characterised by a variety of skeletal malformations and congenital heart defects. The gene for Holt-Oram syndrome has been identified as TBX5 on chromosome 12q24. TBX5 encodes a protein of 518 amino acids that belongs to the family of the T box transcriptional factors, 11 and is expressed in embryonic heart and limb tissues, consistent with its involvement in development of the heart and skeletal structures. TBX5 contains a highly conserved DNA binding domain, the T box domain. Recently, three groups have provided direct evidence that TBX5 can bind to DNA and activate transcription of its target genes including the gene for atrial natriuretic factor (ANF). Furthermore, TBX5 can interact directly with the cardiac homeobox protein NKX2.5 synergistically to activate transcription of ANF. 18 To date, 22 different non-translocation TBX5 mutations have been reported. These include five truncations (nonsense mutations), three splicing changes, six frameshift mutations, seven missense mutations, and one large deletion involving exons 3-9. Frameshift, splicing, and nonsense mutations are expected to produce truncated TBX5 or no TBX5 at all (for example, via nonsense mediated mRNA decay), which causes Holt-Oram syndrome by haploinsufficiency. However, the mechanisms by which the missense mutations cause HoltOram syndrome are not well understood. Ghosh et al found that missense mutations G80R and R237Q eliminated DNA binding, whereas mutations G169R and S252I did not affect DNA binding. Hiroi et al analysed the effect of G80R and R237Q on transcription activity of the ANF promoter. Mutation G80R caused significant reduction of transcriptional activation activity of TBX5 and loss of synergistic activation with NKX2.5. Surprisingly, mutation R237Q, which eliminated DNA binding, had minor effects on transcriptional activation. Continued identification and functional analysis of TBX5 mutations will be invaluable in identifying new mechanisms underlying the pathogenesis of Holt-Oram syndrome. In this study, we have identified two novel mutations in TBX5, one microdeletion (381-408del27bp) and one nonsense mutation 192G>A (W64X), and report the detailed molecular mechanisms by which the 381-408del27bp deletion causes Holt-Oram syndrome.