Jackson Stephen M, Whitworth Alex J, Greene Jessica C, Libby Randell T, Baccam Sandy L, Pallanck Leo J, La Spada Albert R
Department of Genome Sciences, University of Washington, Seattle, WA 98195, USA.
Gene. 2005 Feb 28;347(1):35-41. doi: 10.1016/j.gene.2004.12.008.
CAG and CTG repeat expansions are the cause of at least a dozen inherited neurological disorders. In these so-called "dynamic mutation" diseases, the expanded repeats display dramatic genetic instability, changing in size when transmitted through the germline and within somatic tissues. As the molecular basis of the repeat instability process remains poorly understood, modeling of repeat instability in model organisms has provided some insights into potentially involved factors, implicating especially replication and repair pathways. Studies in mice have also shown that the genomic context of the repeat sequence is required for CAG/CTG repeat instability in the case of spinocerebellar ataxia type 7 (SCA7), one of the most unstable of all CAG/CTG repeat disease loci. While most studies of repeat instability have taken a candidate gene approach, unbiased screens for factors involved in trinucleotide repeat instability have been lacking. We therefore attempted to use Drosophila melanogaster to model expanded CAG repeat instability by creating transgenic flies carrying trinucleotide repeat expansions, deriving flies with SCA7 CAG90 repeats in cDNA and genomic context. We found that SCA7 CAG90 repeats are stable in Drosophila, regardless of context. To screen for genes whose reduced function might destabilize expanded CAG repeat tracts in Drosophila, we crossed the SCA7 CAG90 repeat flies with various deficiency stocks, including lines lacking genes encoding the orthologues of flap endonuclease-1, PCNA, and MutS. In all cases, perfect repeat stability was preserved, suggesting that Drosophila may not be a suitable system for determining the molecular basis of SCA7 CAG repeat instability.
CAG和CTG重复序列的扩增是至少十几种遗传性神经疾病的病因。在这些所谓的“动态突变”疾病中，扩增的重复序列表现出显著的遗传不稳定性，在通过种系传递以及在体细胞组织中时大小会发生变化。由于重复序列不稳定性过程的分子基础仍知之甚少，在模式生物中对重复序列不稳定性进行建模为可能涉及的因素提供了一些见解，尤其涉及复制和修复途径。对小鼠的研究还表明，对于7型脊髓小脑共济失调（SCA7），即所有CAG/CTG重复疾病位点中最不稳定的位点之一，重复序列的基因组背景是CAG/CTG重复序列不稳定性所必需的。虽然大多数关于重复序列不稳定性的研究都采用了候选基因方法，但缺乏对参与三核苷酸重复序列不稳定性的因素进行无偏筛选。因此，我们试图通过创建携带三核苷酸重复序列扩增的转基因果蝇，在cDNA和基因组背景中获得具有SCA7 CAG90重复序列的果蝇，利用黑腹果蝇对扩增的CAG重复序列不稳定性进行建模。我们发现，无论背景如何，SCA7 CAG90重复序列在果蝇中都是稳定的。为了筛选其功能降低可能会使果蝇中扩增的CAG重复序列不稳定的基因，我们将SCA7 CAG90重复序列果蝇与各种缺失品系杂交，包括缺乏编码瓣状核酸内切酶-1、增殖细胞核抗原和MutS同源物的基因的品系。在所有情况下，重复序列的完美稳定性都得以保留，这表明果蝇可能不是确定SCA7 CAG重复序列不稳定性分子基础的合适系统。
