Nance-Horan syndrome (NHS) is an X-linked developmental disorder characterized by congenital cataract, dental anomalies, facial dysmorphism and, in some cases, mental retardation. Protein truncation mutations in a novel gene (NHS) have been identified in patients with this syndrome. We previously mapped X-linked congenital cataract (CXN) in one family to an interval on chromosome Xp22.13 which encompasses the NHS locus; however, no mutations were identified in the NHS gene. In this study, we show that NHS and X-linked cataract are allelic diseases. Two CXN families, which were negative for mutations in the NHS gene, were further analysed using array comparative genomic hybridization. CXN was found to be caused by novel copy number variations: a complex duplication–triplication re-arrangement and an intragenic deletion, predicted to result in altered transcriptional regulation of the NHS gene. Furthermore, we also describe the clinical and molecular analysis of seven families diagnosed with NHS, identifying four novel protein truncation mutations and a novel large deletion encompassing the majority of the NHS gene, all leading to no functional protein. We therefore show that different mechanisms, aberrant transcription of the NHS gene or no functional NHS protein, lead to different diseases. Our data highlight the importance of copy number variation and non-recurrent re-arrangements leading to different severity of disease and describe the potential mechanisms involved.

No abstract

The distribution of d(CT)-rich pyrimidine tracts in the karyotypes of a variety of vertebrates was studied by in situ hybridization. The probe for these studies was a 56bp homopyrimidine/homopurine sequence obtained from a mouse genomic library constructed with DNA prepared from a restriction enzyme digestion of metaphase chromosomes. Single-stranded DNA nuclease digestions and two-dimensional gel analysis of topoisomers of this sequence indicated that it is capable of adopting a triplex conformation in vitro. In situ hybridization with this probe to the karyotypes of ten different vertebrate species revealed a highly conserved chromosomal distribution of d(CT)-rich tracts. These tracts are found throughout the chromosomal arms and in some karyotypes they are clustered, producing a banding pattern. However, at the resolution of the light microscope these tracts appeared to be absent from the centromeric regions of all chromosomes examined except those of chicken. The non-random distribution of these tracts to the chromosomal arm regions implies an organizational or functional role for this repeat class. It is unlikely that the 56 bp sequence type contributed to the formation of the triplex DNA structure previously detected in centromeric domains of mouse.

pBR322-derived plasmids have been constructed carrying d(GTAC)n.d(GTAC)n inserts of different lengths, in order to investigate the effect of insert size on cruciform extrusion and/or the B-Z transition. Plasmids with n ranging from 4 to 12 are hypersensitive to cleavage by the single-strand specific nucleases, S1 nuclease and Bal31 nuclease. Hypersensitive sites associated with the smaller alternating purine-pyrimidine tracts, however, coexist with the major pBR322 sites. Site-selective cleavage of these plasmids with the resolvase, T7 endonuclease I, demonstrates that all the inserts form cruciform structures when stably integrated into negatively supercoiled plasmids. An increase in the negative superhelical density of the DNA's induces cruciform formation within the insert region, resulting in a reduction in torsional stress consistent with the size of the insert. Moreover, as n decreases, the superhelical density required to stabilise the cruciform state increases. Therefore, the cruciform geometry is the favoured conformation of these d(GTAC)n.d(GTAC)n sequences under torsional stress. The stability of these cruciforms increases as n increases, with cruciformation occurring at lower superhelical densities and to the exclusion of the other pBR322 cruciforms.

No abstract