Cys2His2 zinc fingers, the most common structural motif found in eukaryotic transcription factors, can mediate both protein-DNA and protein-protein interactions.  The widespread prevalence of zinc fingers in genomes ranging from yeast to humans emphasizes the importance of understanding how zinc finger proteins function in normal and disease-related cell processes. 

Extended Zinc Finger Protein-DNA Interfaces

We are studying the Neuron Restrictive Silencer Factor/RE-1 Silencing Transcription Factor (NRSF/REST), a transcriptional regulator that represses or silences the expression of neuron-specific genes in non-neuronal cells by binding to an extended 21-base-pair DNA sequence using a cluster of eight Cys2His2 zinc fingers.  We are using a combination of genetic, biochemical, and structural studies to define the complex series of amino acid-DNA base interactions at the NRSF/REST-DNA interface. 

Protein-Protein Interactions by Zinc Fingers

We are studying protein-protein interactions mediated by the human Ikaros and the Drosophila melanogaster Hunchback proteins.  Each of these transcription factors uses a pair of zinc fingers to specifically mediate self-oligomerization.  We have created genetic selection systems based in E. coli that have enabled us to map the molecular level interactions that occur at these finger-finger interfaces.  In addition, the insights we have gained from these studies combined with the ability to perform genetic selections have permitted us to create novel zinc finger domains with altered interaction specificities (Giesecke et al., Molecular Systems Biology 2006).

Design and Evolution of Proteins for Targeted Recombination

Proteins that could direct high efficiency gene targeting events to a specific desired endogenous locus would have numerous applications in biological research, and potentially in gene therapy.  Recent studies have demonstrated that engineered “zinc finger nucleases” can be used to promote high-efficiency gene targeting to a desired genomic locus in plant, fly, worm, and human cells.  However, significant protein design challenges remain to be addressed before targeted single-gene recombination can become a reality:  1) development of rapid methods for creating customized zinc finger domains that function well in a cellular context, and 2) improving the currently promiscuous specificity of restriction enzyme fusions described to date.  We have previously described a method for producing highly specific multi-finger proteins using a directed domain shuffling approach (Hurt et al, PNAS 2003) and are developing a modified version of this approach designed for higher-throughput use. 

The Joung lab co-founded and leads the Zinc Finger Consortium, an international group of academic investigators committed to the development of engineered zinc finger technology (see http://www.zincfingers.org for additional information).  The Zinc Finger Consortium has recently published their initial work describing a single consistent platform for assembling engineered zinc fingers using modular assembly of pre-selected domains (Wright et al., Nature Protocols 2006).