The cleidocranial dysplasia-related R131G mutation in the Runt-related transcription factor RUNX2 disrupts binding to DNA but not CBF-beta
Department of Cell Biology
Amino Acid Motifs; Amino Acid Substitution; Animals; CHO Cells; Cell Nucleus; Cleidocranial Dysplasia; Core Binding Factor Alpha 1 Subunit; Core Binding Factor Alpha 2 Subunit; Core Binding Factor beta Subunit; Cricetinae; Cricetulus; DNA; Hela Cells; Humans; Mutant Proteins; Mutation; Protein Binding; Protein Multimerization; Protein Structure, Tertiary; Protein Transport; Transcriptional Activation
Cleidocranial dysplasia (CCD) is caused by haploinsufficiency in RUNX2 function. We have previously identified a series of RUNX2 mutations in Korean CCD patients, including a novel R131G missense mutation in the Runt-homology domain. Here, we examine the functional consequences of the RUNX2(R131G) mutation, which could potentially affect DNA binding, nuclear localization signal, and/or heterodimerization with core-binding factor-beta (CBF-beta). Immunofluorescence microscopy and western blot analysis with subcellular fractions show that RUNX2(R131G) is localized in the nucleus. Immunoprecipitation analysis reveals that heterodimerization with CBF-beta is retained. However, precipitation assays with biotinylated oligonucleotides and reporter gene assays with RUNX2 responsive promoters together reveal that DNA-binding activity and consequently the transactivation of potential of RUNX2(R131G) is abrogated. We conclude that loss of DNA binding, but not nuclear localization or CBF-beta heterodimerization, causes RUNX2 haploinsufficiency in patients with the RUNX2(R131G) mutation. Retention of specific functions including nuclear localization and binding to CBF-beta of the RUNX2(R131G) mutation may render the mutant protein an effective competitor that interferes with wild-type function.
DOI of Published Version
J Cell Biochem. 2010 May;110(1):97-103. Link to article on publisher's site
Journal of cellular biochemistry
Han M, Kim H, Wee H, Lim K, Park N, Bae S, Van Wijnen AJ, Stein JL, Lian JB, Stein GS, Choi J. (2010). The cleidocranial dysplasia-related R131G mutation in the Runt-related transcription factor RUNX2 disrupts binding to DNA but not CBF-beta. Stein, Stein, Lian, vanWijnen Lab Publications. https://doi.org/10.1002/jcb.22516. Retrieved from https://escholarship.umassmed.edu/stein/16