Transcription factors RUNX1/AML1 and RUNX2/Cbfa1 dynamically associate with stationary subnuclear domains.
The runt-related transcription factors (RUNX/Cbfa/AML) are essential for cellular differentiation and fetal development. C-terminal truncations of RUNX factors that eliminate the targeting of these factors to subnuclear foci result in lethal hematopoietic and skeletal phenotypes. Here we demonstrate that in living cells the RUNX C-terminus is necessary for the dynamic association of RUNX into stable subnuclear domains. Time-lapse fluorescence microscopy shows that RUNX1 and RUNX2 localize to punctate foci that remain stationary in the nuclear space. By fluorescence recovery after photobleaching assays, both proteins are shown to dynamically associate at these subnuclear foci, with a 10 second half-time of recovery. A truncation of RUNX2, removing its intranuclear targeting signal (NMTS), increases its mobility by an order of magnitude, resulting in a half-time of recovery equivalent to that of EGFP alone. We propose that the dynamic shuttling of RUNX factors in living cells to positionally stabilized foci, which is dependent on the C-terminus, is a component of the mechanism for gene regulation in vivo.
D021581:Active Transport, Cell Nucleus / Q000235:genetics; D002454:Cell Differentiation / Q000235:genetics; D002467:Cell Nucleus / Q000378:metabolism* / Q000648:ultrastructure; D050659:Core Binding Factor Alpha 1 Subunit; D050676:Core Binding Factor Alpha 2 Subunit; D050657:Core Binding Factor alpha Subunits; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D005057:Eukaryotic Cells / Q000166:cytology / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D018507:Gene Expression Regulation, Developmental / Q000235:genetics; D006367:HeLa Cells; D006801:Humans; D046911:Macromolecular Substances; D009363:Neoplasm Proteins; D017434:Protein Structure, Tertiary / Q000235:genetics; D011518:Proto-Oncogene Proteins; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*
D050659:Core Binding Factor Alpha 1 Subunit; D050676:Core Binding Factor Alpha 2 Subunit; D050657:Core Binding Factor alpha Subunits; D004268:DNA-Binding Proteins; D046911:Macromolecular Substances; D009363:Neoplasm Proteins; D011518:Proto-Oncogene Proteins; C493728:RUNX1 protein, human; D014157:Transcription Factors