GAA TTC STRUCTURES--FUNCTIONS AND FRIEDREICHS ATAXIA

弗里德赖希共济失调（FRDA）是一种常染色体隐性遗传疾病，由内含子GAA.TTC三联重复序列扩增引起。FRDA是最常见的遗传性共济失调。FRDA基因分布在40 kb以上，包含5个外显子，编码一种210个氨基酸的蛋白质，称为frataxin。正常个体含有7-22个GAA.TTC单位，而FRDA患者具有100-1700个单位的扩大和多态性等位基因。共济失调蛋白基因第一内含子中GAA.TTC重复序列的扩增导致共济失调蛋白mRNA和蛋白水平的降低。较短长度的GAA.TTC重复采用分子内和分子间三链体，长重复（>60个单元）形成几种可能的不寻常构象之一。我们最近发现，150和270重复的GAA.TTC采用了一种引人注目的新形式的“弯曲”DNA，提高了对DNA构象在这种疾病的分子病因学中的作用的兴趣。目的是阐明负责FRDA的核酸分子机制。目的1研究正常人和扩增等位基因中不同长度的GAA、TTC的构象。表征将通过流体力学性质、化学和酶探针分析、EM、环化动力学和螺旋重复研究进行。此外，我们还将研究DNA GAA、TTC和内含子rGAA之间形成的构象。目的二是通过对FRDA TRS结构的体外转录延伸率的测定，以及对RNA和mRNA、DNA结构的研究，研究FRDA TRS结构的转录特性。此外，将沿着监测转录的保真度以及转录对重复稳定性的影响。我们的新数据揭示了FRDA三联体重复序列（TRS）在体内和体外扩展的内含子遗传不稳定性的抑制作用。总之，我们将通过阐明不同长度的GAA、TTC的DNA构象及其对转录和遗传不稳定性（复制和重组）的影响来研究FRDA的分子病因学。

Friedreich's ataxia (FRDA) is an autosomal recessive disease caused by an intronic GAA.TTC triplet repeat expansion. FRDA is the most common hereditary ataxia. The FRDA gene is spread over 40 kb and contains five exons which encode a 210 amino acid protein named frataxin. Normal individuals contain 7-22 GAA.TTC units whereas FRDA patients have enlarge and polymorphic alleles with 100-1700 units. The expansion of the GAA.TTC repeat in the first intron of the frataxin gene results in reduced levels of frataxin mRNA and protein. Shorter lengths of GAA.TTC repeats adopt intra- and inter-molecular triplexes and long repeats (>60 units) form one of several possible unusual conformations. Our recent discovery that 150 and 270 repeats of GAA.TTC adopt a dramatically new form of "bent" DNA heightens the interest in the role of DNA conformation in the molecular etiology of this disease. The goal is to elucidate the nucleic acid molecular mechanisms responsible for FRDA. Aim 1 is to study the conformations of different lengths of GAA.TTC from normal individuals to the enlarged alleles. Characterization will be by hydrodynamic properties, chemical and enzymatic probe analysis, EM, circularization kinetics and helical repeat studies. Also, the conformations formed between DNA GAA.TTC and intronic rGAA will be investigated. Aim 2 is to study the transcription properties of the FRDA TRS structures by nuclear runoff experiments from patient materials, determinations of the transcription elongation rate in vitro, and studies on the RNA as well as mRNA.DNA structures. Also, the fidelity of transcription will be monitored along with the effects of transcription on repeat stability. Our new data revealed the inhibitory effects of expanded intronic genetic instabilities of the FRDA triplet repeat sequences (TRS) in vivo and vitro. In summary, we shall study the molecular etiology of FRDA by elucidating the DNA conformations of different lengths of GAA.TTC and their effect on transcription and on genetic instabilities (replication and recombination).