Govani Fatima S, Shovlin Claire L
NHLI Cardiovascular Sciences, Imperial College London, UK, Hammersmith Campus, Du Cane Rd, London W12 0NN, UK.
J Angiogenes Res. 2010 Aug 11;2:15. doi: 10.1186/2040-2384-2-15.
There is significant interest in new loci for the inherited condition hereditary haemorrhagic telangiectasia (HHT) because the known disease genes encode proteins involved in vascular transforming growth factor (TGF)-beta signalling pathways, and the disease phenotype appears to be unmasked or provoked by angiogenesis in man and animal models. In a previous study, we mapped a new locus for HHT (HHT3) to a 5.7 Mb region of chromosome 5. Some of the polymorphic markers used had been uninformative in key recombinant individuals, leaving two potentially excludable regions, one of which contained loci for attractive candidate genes encoding VE Cadherin-2, Sprouty4 and FGF1, proteins involved in angiogenesis.
Extended analyses in the interval-defining pedigree were performed using informative genomic sequence variants identified during candidate gene sequencing. These variants were amplified by polymerase chain reaction; sequenced on an ABI 3730xl, and analysed using FinchTV V1.4.0 software.
Informative genomic sequence variants were used to construct haplotypes permitting more precise citing of recombination breakpoints. These reduced the uninformative centromeric region from 141.2-144 Mb to between 141.9-142.6 Mb, and the uninformative telomeric region from 145.2-146.9 Mb to between 146.1-146.4 Mb.
The HHT3 interval on chromosome 5 was reduced to 4.5 Mb excluding 30% of the coding genes in the original HHT3 interval. Strong candidates VE-cadherin-2 and Sprouty4 cannot be HHT3.
遗传性出血性毛细血管扩张症(HHT)新基因座备受关注,因为已知的疾病基因编码参与血管转化生长因子(TGF)-β信号通路的蛋白质,并且在人和动物模型中,疾病表型似乎因血管生成而显现或被激发。在先前的一项研究中,我们将HHT的一个新基因座(HHT3)定位到5号染色体上一个5.7 Mb的区域。所使用的一些多态性标记在关键重组个体中无信息,留下两个可能可排除的区域,其中一个区域包含编码血管内皮钙黏蛋白-2、Sprouty4和FGF1的有吸引力的候选基因座,这些蛋白质参与血管生成。
在界定区间的家系中进行扩展分析,使用候选基因测序过程中鉴定的信息丰富的基因组序列变异。这些变异通过聚合酶链反应进行扩增;在ABI 3730xl上进行测序,并使用FinchTV V1.4.0软件进行分析。
信息丰富的基因组序列变异用于构建单倍型,从而更精确地确定重组断点。这将无信息的着丝粒区域从141.2 - 144 Mb缩小到141.9 - 142.6 Mb之间,将无信息的端粒区域从145.2 - 146.9 Mb缩小到146.1 - 146.4 Mb之间。
5号染色体上的HHT3区间缩小到4.5 Mb,排除了原始HHT3区间中30%的编码基因。强有力的候选基因血管内皮钙黏蛋白-2和Sprouty4不可能是HHT3。