Tassone F, Hagerman R J, Chamberlain W D, Hagerman P J
Department of Biochemistry and Molecular Genetics B-121, University of Colorado School of Medicine, 4200 East 9th Avenue, Denver, CO 80262, USA.
Am J Med Genet. 2000 Fall;97(3):195-203. doi: 10.1002/1096-8628(200023)97:3<195::AID-AJMG1037>3.0.CO;2-R.
Fragile X syndrome generally arises as a consequence of a large expansion of a CGG trinucleotide repeat element that is located in the GC-rich promoter region of the fragile X mental retardation gene (FMR1). In the conventional model for fragile X, clinical involvement arises as a consequence of silencing of the FMR1 gene, with the attendant loss of FMR1 protein (FMRP). However, it has recently been demonstrated that most males with large premutation alleles (100-200 repeats), or with unmethylated full mutation alleles, have FMR1 mRNA levels that are higher than normal, despite reduced levels of FMRP. In the current work, we extend and confirm these observations using quantitative (fluorescent) reverse transcription polymerase chain reaction on larger sample populations, establishing that even for smaller premutation alleles (55-100 repeats) the mRNA levels are significantly elevated (mean 2.1-fold elevation; P = 3.9 x 10(-3)), relative to normal controls. Thus, an abnormal molecular phenotype is established close to the upper end of the normal range. We also demonstrate that the levels of FMR1 mRNA are elevated in females with premutation alleles; however, the mRNA levels are more varied than in the males, and are attenuated in a manner that is consistent with the fraction of normal alleles that are active in any given individual. Finally, we demonstrate that in lymphoblastoid cells derived from a patient with a severe form of fragile X caused by a point mutation in the second KH domain of the gene, but with a normal CGG element (25 repeats), the FMR1 mRNA level is normal. Thus, although models in which FMRP level (or level of function) modulates transcriptional activity remain viable, other explanations for the elevated message levels, including direct (cis) effects of the CGG element on transcription, must also be considered.
脆性X综合征通常是由于位于脆性X智力低下基因（FMR1）富含GC的启动子区域的CGG三核苷酸重复元件大量扩增所致。在脆性X的传统模型中，临床受累是FMR1基因沉默的结果，随之FMR1蛋白（FMRP）缺失。然而，最近已证明，大多数具有大的前突变等位基因（100 - 200个重复）或未甲基化的全突变等位基因的男性，尽管FMRP水平降低，但FMR1 mRNA水平高于正常。在当前研究中，我们通过对更大样本群体进行定量（荧光）逆转录聚合酶链反应扩展并证实了这些观察结果，确定即使对于较小的前突变等位基因（55 - 100个重复），相对于正常对照，mRNA水平也显著升高（平均升高2.1倍；P = 3.9×10⁻³）。因此，在正常范围上限附近建立了一种异常分子表型。我们还证明，具有前突变等位基因的女性中FMR1 mRNA水平升高；然而，mRNA水平比男性更具变异性，并且以与任何给定个体中活跃的正常等位基因比例一致的方式减弱。最后，我们证明，在由该基因第二个KH结构域中的点突变引起但具有正常CGG元件（25个重复）的严重脆性X患者来源的淋巴母细胞中，FMR1 mRNA水平正常。因此，尽管FMRP水平（或功能水平）调节转录活性的模型仍然可行，但也必须考虑对升高的信使水平的其他解释，包括CGG元件对转录的直接（顺式）影响。
