A Multiprotein Complex Involved In Atrx Syndrome

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Abstract

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. Using our techniques, ATRX fractionated as a complex of 1.5 Mda by gel-filtration chromatography. The ATRX complex was immunoisolated using 4 antibodies against 4 different regions of the protein. Mass spectrometry and immunoblotting analysis revealed that the complex comprises ATRX and at least 7 other components (referred to as AXAPs for ATRX-associated proteins). Several AXAPs have been previously identified as transcription factors. For example, AXAP110 has been shown to act as a transcriptional corepressor. In a critical control, antibody against AXAP100 could immunoprecipitate ATRX. Immunofluorescence studies showed that these two proteins colocalize in vivo. Two other AXAPs have been previously identified as chromatin-dependent transcription factors. We have tested the ATRX complex for conventional mononucleosomes disruption activity. ATRX complex has an activity rather different from that of the SWI/SNF or NURD families of remodeling complexes, and we are currently extending assays to study biochemical activities of the ATRX complex.