A Multiprotein Complex Involved In Atrx Syndrome

ATRX综合征代表α-地中海贫血、智力迟钝和多种相关发育异常的组合。ATRX缺陷基因已定位于X染色体并克隆。同一基因的突变也会导致其他几种形式的X连锁综合征性精神发育迟滞。ATRX基因编码含有SWI 2/SNF 2型DNA依赖性ATP酶结构域的基因产物。因此，有人假设ATRX可以在ATP依赖性染色质重塑复合物中发挥作用，并参与基因表达的调控。我们与Doug Higgs和Richard Gibbons博士建立了合作关系，他们克隆了ATRX基因，并发现了其功能的重要特征。通过免疫沉淀的方法，我们发现ATRX与转录辅因子Daxx形成复合物。以下证据支持ATRX和Daxx是ATP依赖性染色质重塑复合物的组分：1）Daxx和ATRX可以通过对每种蛋白质特异的抗体共免疫分离; 2）通过凝胶过滤分析，Daxx与ATRX共分级分离的比例为1 MDa的复合物; 3）在ATRX综合征患者的细胞提取物中，Daxx-ATRX复合物的水平相应降低; 4）一部分ATRX和Daxx共定位在PML核体中，其中Daxx先前已被发现;和5）ATRX复合物显示类似于其他染色质重塑复合物的ATP依赖性活性，包括三螺旋DNA置换和单核小体破坏模式的改变。但与先前描述的SWI/SNF或CIDD复合物不同，ATRX复合物不会使单核体的DNA定相随机化，这表明它可以不同地重塑染色质。两者合计，结果表明，ATRX功能与Daxx作为一种新的染色质重塑复合物。ATR-X综合征的缺陷可能是由这种复合物控制的基因的不适当表达引起的。我们现在正在研究这些因子调控的基因是否与ATR-X综合征有关。

ATRX syndrome represents a combination of alpha-thalassemia, mental retardation, and multiple associated developmental abnormalities. The gene defective in ATRX has been localized to the X chromosome and cloned. Mutations in the same gene also cause several other forms of syndromal X-linked mental retardation. The ATRX gene encodes a gene product containing a SWI2/SNF2-type DNA-dependent ATPase domain. Thus, it has been hypothesized that ATRX could function in an ATP-dependent chromatin-remodeling complex and participate in regulation of gene expression. We established a collaboration with Drs. Doug Higgs and Richard Gibbons, who had cloned the ATRX gene, and have discovered important features of its function. By immunoprecipitation from HeLa extract, we found that ATRX is in a complex with transcription cofactor Daxx. The following evidence supports that ATRX and Daxx are components of an ATP-dependent chromatin-remodeling complex: 1) Daxx and ATRX can be coimmunoisolated by antibodies specific for each protein; 2) a proportion of Daxx co-fractionates with ATRX as a complex of 1 MDa by gelfiltration analysis; 3) in extract of cells of a patient with ATRX syndrome, the level of the Daxx-ATRX complex is correspondingly reduced; 4) a proportion of ATRX and Daxx colocalize in PML nuclear bodies, where Daxx had previously been found; and 5) ATRX complex displays ATP-dependent activities that resemble those of other chromatin remodeling complexes, including triple helix DNA displacement and alteration of mononucleosome disruption patterns. But unlike the previously described SWI/SNF or NURD complexes, the ATRX complex does not randomize DNA phasing of the mononucleosomes, suggesting that it may remodel chromatin differently. Taken together, the results suggest that ATRX functions in conjunction with Daxx as a novel chromatin-remodeling complex. The defects in ATR-X syndrome may result from inappropriate expression of genes controlled by this complex.We have identified several sequence-specific transcription factors that co-purify with Daxx. We are now studying if the genes regulated by these factors could be involved in ATR-X syndrome.