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Updated: May 19, 2026

High Sensitivity Measurement of Transcription Factor-DNA Binding Affinities by Competitive Titration Using Fluorescence Microscopy
Published on: February 7, 2019
Characterization of the DNA-binding properties of the Mohawk homeobox transcription factor
Douglas M Anderson1, Rajani George1, Marcus B Noyes2
1School of Life Sciences, Biodesign Institute, Arizona State University, Tempe, Arizona 85287-4501; Molecular and Cellular Biology Graduate Program, Biodesign Institute, Arizona State University, Tempe, Arizona 85287-4501.
Abstract:
The homeobox transcription factor Mohawk (Mkx) is a potent transcriptional repressor expressed in the embryonic precursors of skeletal muscle, cartilage, and bone. MKX has recently been shown to be a critical regulator of musculoskeletal tissue differentiation and gene expression; however, the genetic pathways through which MKX functions and its DNA-binding properties are currently unknown. Using a modified bacterial one-hybrid site selection assay, we determined the core DNA-recognition motif of the mouse monomeric Mkx homeodomain to be A-C-A. Using cell-based assays, we have identified a minimal Mkx-responsive element (MRE) located within the Mkx promoter, which is composed of a highly conserved inverted repeat of the core Mkx recognition motif. Using the minimal MRE sequence, we have further identified conserved MREs within the locus of Sox6, a transcription factor that represses slow fiber gene expression during skeletal muscle differentiation. Real-time PCR and immunostaining of in vitro differentiated muscle satellite cells isolated from Mkx-null mice revealed an increase in the expression of Sox6 and down-regulation of slow fiber structural genes. Together, these data identify the unique DNA-recognition properties of MKX and reveal a novel role for Mkx in promoting slow fiber type specification during skeletal muscle differentiation.
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