(786.4) Structural characterization of the cone-rod homeobox protein binding to DNA

Poster Board Number: A20

Christine Buchholz (James Madison University), Abigail Sholes (James Madison University), Raymond Enke (James Madison University), Christopher Berndsen (James Madison University)

Roses are red, some eyes are blue, you wouldn’t see this without CRX too. The Cone-Rod Homeobox protein or CRX is an essential transcription factor for the development of cells in the eye called photoreceptors. Photoreceptor cells (PRs) convert light into signals that the brain can understand. CRX plays a role in the differentiation of these photoreceptors into rods and cones, which detect achromatic and colored light, respectively. While there are details known about the molecular function of CRX, there is little known about the structural information of the protein. It is known that CRX contains a DNA binding domain (DBD) and two activation domains (ADs). Changes to the amino acid sequence in any of these regions are linked to a number of diseases associated with congenital blindness. We would like to better understand the structure and function of the AD and DBD to describe the regulation of photoreceptor development by CRX. We used Small-Angle X-Ray Scattering (SAXS) and DNA binding assays on the DBD showing that the DBD is globular and can bind to DNA in the absence of the activation domain. We crosslinked the DBD peptide to DNA with glutaraldehyde in an attempt to get a bound structure of the peptide-DNA complex. The peptide-DNA complex will show more accurately how the DBD binds to DNA and the stoichiometry of the complex. We are working to confirm preliminary findings suggesting a 2:1 DBD:DNA complex.  These experiments will help us to understand the role that CRX plays in regulating transcriptional networks in PR neurons.

This work was supported in part by award NIH NEI R15EY028725 (to R.E.), the 4-VA organization, and equipment purchased by an award from the Thomas and Kate F. Jeffress Memorial Trust (to C.E.B.).