Division of Biology
Kansas State University
Ligand-binding and structural studies of the START domain from HD-Zip proteins (2016-2017)
Plant-derived lipid metabolites such as flavonoids act via multiple mechanisms that are not understood at the molecular level, including roles in cancer progression in the inhibition of cell proliferation. In addition, misexpression of several START domain family members from humans is observed in human tumors. This project will elucidate novel interactions between lipids and transcriptional regulators and how these are related to the control of cell division with relevance to our understanding of cancer and other diseases.
The START (STeroidogenic Acute Regulatory (StAR)-related lipid Transfer) domain is a ligand binding motif that was first discovered in StAR cholesterol transporters. Fifteen human proteins contain START domains, about half of which are known to bind to specific ligands such as sterols and phospholipids. Over the past decades, START domain-containing proteins have been shown to be integral components of intracellular signaling pathways. In plants, START domains occur in homeodomain leucine-zipper (HD-ZIP) transcription factors that are master regulators of gene expression during cell-type differentiation.
Yet, the molecular mechanisms defining the function of this putative lipid-binding domain are not understood, nor has a ligand been identified for any plant-derived START domain to date. The overall goal of this project is to obtain ligand-binding and structural information for the START domain of PROTODERMAL FACTOR 2 (PDF2), a HD-ZIP transcription factor required for proper differentiation of the epidermis in Arabidopsis. Preliminary data implicate flavonoids as candidate ligands. The specific aims are: (1) Characterize ligand-binding properties of the START domain from PDF2, a representative HD-ZIP transcription factor, and (2) Determine the 3D structure of the ligand-bound complex. 6xHis-tagged proteins containing the START domain of HD-ZIP transcription factor PDF2, and mutants thereof, will be expressed in E. coli and isolated by affinity purification.
Ligand-binding studies with flavonoids and fragment libraries will be conducted using a surface plasmon resonance (SPR) based method in conjunction with the Protein Production Group at the COBRE-PSF Center. To solve the structure of the protein in the apo and co-crystal form, X-ray crystallography will be applied in collaboration with the Protein Structure Core Laboratory. The results obtained from this work are expected to lead to an improved understanding of START domain structure and function, and may reveal how START domains are linked to cell division versus cell differentiation in humans and in plants. This project will impact public health by providing molecular information on the interplay between lipid ligands and START domain proteins, which are implicated as metabolic sensors and regulators of cell cycle progression in human cancers.