Structural basis for DNA recognition by STAT6
Saturday, 2016/11/19 | 08:17:29
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Jing Li, Jose Pindado Rodriguez, Fengfeng Niu, Mengchen Pu, Jinan Wang, Li-Wei Hung, Qiang Shao, Yanping Zhu, Wei Ding, Yanqing Liu, Yurong Da, Zhi Yao, Jie Yang, Yongfang Zhao, Gong-Hong Wei, Genhong Cheng, Zhi-Jie Liu, and Songying Ouyang
Significance
STAT6 is a transcription factor and plays a predominant role in IL-4/IL-13 and virus-mediated signaling pathways. Extensive studies have linked malfunctions of STAT6 to pathological features of asthma and cancer. Targeting the function of STAT6 has become an attractive therapy. Understanding the molecular mechanisms of STAT6 transcriptional regulation is still scarce. Here, we report the atomic-level structures of the phosphorylated STAT6 core fragment homodimer, both in DNA-free and complexed with N4 or N3 site DNA, uncovering both a larger dimer interface intersection angle and the unique residue H415 of STAT6 as important factors for discrimination of N4 from N3 site DNA. This study uncovers a dramatic conformational change in STAT6 dimer for recognizing and preferring N4 site DNA.
Abstract
STAT6 participates in classical IL-4/IL-13 signaling and stimulator of interferon genes-mediated antiviral innate immune responses. Aberrations in STAT6-mediated signaling are linked to development of asthma and diseases of the immune system. In addition, STAT6 remains constitutively active in multiple types of cancer. Therefore, targeting STAT6 is an attractive proposition for treating related diseases. Although a lot is known about the role of STAT6 in transcriptional regulation, molecular details on how STAT6 recognizes and binds specific segments of DNA to exert its function are not clearly understood. Here, we report the crystal structures of a homodimer of phosphorylated STAT6 core fragment (STAT6CF) alone and bound with the N3 and N4 DNA binding site. Analysis of the structures reveals that STAT6 undergoes a dramatic conformational change on DNA binding, which was further validated by performing molecular dynamics simulation studies and small angle X-ray scattering analysis. Our data show that a larger angle at the intersection where the two protomers of STAT meet and the presence of a unique residue, H415, in the DNA-binding domain play important roles in discrimination of the N4 site DNA from the N3 site by STAT6. H415N mutation of STAT6CF decreased affinity of the protein for the N4 site DNA, but increased its affinity for N3 site DNA, both in vitro and in vivo. Results of our structure–function studies on STAT6 shed light on mechanism of DNA recognition by STATs in general and explain the reasons underlying STAT6’s preference for N4 site DNA over N3. See http://www.pnas.org/content/113/46/13015.abstract.html?etoc PNAS November 15 2016; vol.113; no.46: 13015–13020
Figure 1: Structure of STAT6CF and N4 site DNA complex. (A) Schematic diagram showing the domain organization of human STAT6, including N-terminal domain (gray), coiled coil domain (yellow), DNA-binding domain (red), linker domain (orange), SH2 domain (cyan), and TAD domain (gray). The core fragment and phosphorylation site Y641 are indicated. (B) Cartoon diagram of the STAT6CF-N4 complex (in front view). Colors of each domain are the same as in A. (C) Drawing depicting details of the STAT6CF–DNA interface. The side chains of residues (A chain) donating hydrogen bonds are shown as magenta colored sticks, and the hydrogen bonds are shown as yellow dashed lines. The conserved palindromic bases (TTC/GAA) in both sides of the DNA molecule are shown in orange. (D) A schematic drawing highlighting STAT6CF–DNA interactions. Residues forming hydrogen bonds are colored in magenta (D chain) and black (B chain). |
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