A Classic Zinc Finger from Friend of GATA Mediates an Interaction with the Coiled-coil of Transforming Acidic Coiled-coil 3

Abstract

Classic zinc finger domains (cZFs) consist of a ␤-hairpin followed by an ␣-helix. They are among the most abundant of all protein domains and are often found in tandem arrays in DNA-binding proteins, with each finger contributing an ␣-helix to effect sequence-specific DNA recognition. Lone cZFs, not found in tandem arrays, have been postulated to function in protein interactions. We have studied the transcriptional co-regulator Friend of GATA (FOG), which contains nine zinc fingers. We have discovered that the third cZF of FOG contacts a coiled-coil domain in the centrosomal protein transforming acidic coiled-coil 3 (TACC3). Although FOG-ZF3 exhibited low solubility, we have used a combination of mutational mapping and protein engineering to generate a derivative that was suitable for in vitro and structural analysis. We report that the ␣-helix of FOG-ZF3 recognizes a C-terminal portion of the TACC3 coiled-coil. Remarkably, the ␣-helical surface utilized by FOG-ZF3 is the same surface responsible for the well established sequence-specific DNA-binding properties of many other cZFs. Our data demonstrate the versatility of cZFs and have implications for the analysis of many as yet uncharacterized cZF proteins.

The Protein-Binding Potential of C2H2 Zinc Finger Domains

“…As a result of their ability to interact with a variety of substrates, ZFs are involved in numerous cellular processes including chromatin remodeling, gene transcription, translation, RNA trafficking and packaging, protein folding, cellular organization, cell adhesion, and zinc sensing (Brown, 2005;Fox et al, 1999;Hall, 2005;Honma et al, 1999;Kelley et al, 1998;Laity et al, 2001;Lee et al, 2006;Lu et al, 2003;Morgan et al, 1997;Pelham and Brown, 1980;Perdomo et al, 2000;Simpson et al, 2004;Sun et al, 1996).…”

“…After conducting yeast-two hybrid and immunoprecipitation experiments to find and map the PPI between the third (classic) zinc finger of FOG1 and TACC3, Simpson et al (Simpson et al, 2004) combined NMR and alanine mutagenesis to pinpoint critical amino acid involved in the interaction. Interestingly, they demonstrated that amino acids in the α-helix of FOG1 formed the binding surface for the interaction and that residues in positions normally involved in contacting DNA, positions -1, 2, 3, and 6, were also utilized for protein interactions (Figure 1.7).…”

Coactivators necessary for transcriptional output of the hypoxia inducible factor, HIF, are directly recruited by ARNT PAS-B

“…2B). We were particularly interested in the role of the C-terminal 20 residues (CT20) in the ARNT/TACC3 interaction, because these residues are critical for TACC3 binding to the transcriptional coregulator FOG-1 (friend of GATA-1) (23). Using pulldown assays with His 6 -tagged ARNT PAS-B, we determined that a C-terminal fragment of the GST-TACC domain (residues 561-631) was sufficient to interact with ARNT PAS-B (Fig.…”

“…Using a high-resolution structure of the ARNT PAS-B/HIF-2α PAS-B heterodimer (26) and a de novo model of the TACC3 C-terminal coiled coil validated by experimental studies (23) (Fig. S4B), we used a semirigid body docking algorithm implemented within the HADDOCK program (27).…”

Zinc fingers are known as domains for binding DNA and RNA. Do they also mediate protein-protein interactions?

“…Rather, four of them have been shown to interact independently with the transcription factor GATA-1 (7), and mutations that abolish this interaction are associated with inherited blood disorders (8,9). Another ZnF of FOG-1 recognizes the coiled-coil domain of TACC3 using residues in the a-helix (10,11). It is notable that the surfaces used by FOG-1 ZnFs to recognize other proteins overlap partially with those used in interactions between other classical ZnFs and DNA, suggesting that ZnFs are likely to be an ancient structural scaffold onto which different functions have been added.…”