An Qian, Wright Sarah L, Konn Zoë J, Matheson Elizabeth, Minto Lynne, Moorman Anthony V, Parker Helen, Griffiths Mike, Ross Fiona M, Davies Teresa, Hall Andy G, Harrison Christine J, Irving Julie A, Strefford Jon C
Cancer Genomics and Leukaemia Research Cytogenetics Groups, Cancer Sciences Division, University of Southampton, Southampton, SO16 6YD, United Kingdom.
Proc Natl Acad Sci U S A. 2008 Nov 4;105(44):17050-4. doi: 10.1073/pnas.0803494105. Epub 2008 Oct 28.
The search for target genes involved in unbalanced acquired chromosomal abnormalities has been largely unsuccessful, because the breakpoints of these rearrangements are too variable. Here, we use the example of dicentric chromosomes in B cell precursor acute lymphoblastic leukemia to show that, despite this heterogeneity, single genes are targeted through a variety of mechanisms. FISH showed that, although they were heterogeneous, breakpoints on 9p resulted in the partial or complete deletion of PAX5. Molecular copy number counting further delineated the breakpoints and facilitated cloning with long-distance inverse PCR. This approach identified 5 fusion gene partners with PAX5: LOC392027 (7p12.1), SLCO1B3 (12p12), ASXL1 (20q11.1), KIF3B (20q11.21), and C20orf112 (20q11.1). In each predicted fusion protein, the DNA-binding paired domain of PAX5 was present. Using quantitative PCR, we demonstrated that both the deletion and gene fusion events resulted in the same underexpression of PAX5, which extended to the differential expression of the PAX5 target genes, EBF1, ALDH1A1, ATP9A, and FLT3. Further molecular analysis showed deletion and mutation of the homologous PAX5 allele, providing further support for the key role of PAX5. Here, we show that specific gene loci may be the target of heterogeneous translocation breakpoints in human cancer, acting through a variety of mechanisms. This approach indicates an application for the identification of cancer genes in solid tumours, where unbalanced chromosomal rearrangements are particularly prevalent and few genes have been identified. It can be extrapolated that this strategy will reveal that the same mechanisms operate in cancer pathogenesis in general.
寻找与获得性染色体不平衡异常相关的靶基因的工作在很大程度上并不成功，因为这些重排的断点变化太大。在此，我们以B细胞前体急性淋巴细胞白血病中的双着丝粒染色体为例，表明尽管存在这种异质性，但单个基因可通过多种机制成为靶点。荧光原位杂交（FISH）显示，尽管9p上的断点具有异质性，但会导致PAX5部分或完全缺失。分子拷贝数计数进一步明确了断点，并通过长距离反向PCR促进了克隆。该方法鉴定出5个与PAX5融合的基因伙伴：LOC392027（7p12.1）、SLCO1B3（12p12）、ASXL1（20q11.1）、KIF3B（20q11.21）和C20orf112（20q11.1）。在每个预测的融合蛋白中，PAX5的DNA结合配对结构域均存在。通过定量PCR，我们证明缺失和基因融合事件均导致PAX5表达下调，这延伸至PAX5靶基因EBF1、ALDH1A1、ATP9A和FLT3的差异表达。进一步的分子分析显示同源PAX5等位基因的缺失和突变，为PAX5的关键作用提供了进一步支持。在此，我们表明特定基因位点可能是人类癌症中异质易位断点的靶点，通过多种机制发挥作用。该方法表明在实体瘤中鉴定癌症基因具有应用价值，在实体瘤中染色体不平衡重排尤为普遍，而已鉴定的基因很少。可以推断，该策略将揭示相同的机制在癌症发病机制中普遍起作用。
