Hou Caixia, McCown Claudia, Ivanov Dmitri N, Tsodikov Oleg V
Department of Pharmaceutical Sciences, College of Pharmacy, University of Kentucky, Lexington, Kentucky 40536, United States.
Department of Biochemistry, University of Texas Health Science Center at San Antonio, San Antonio, Texas 78229, United States.
Biochemistry. 2020 Nov 11. doi: 10.1021/acs.biochem.0c00774.
ETS family transcription factors control development of different cell types in humans, whereas deregulation of these proteins leads to severe developmental syndromes and cancers. One of a few members of the ETS family that are known to act solely as repressors, ERF, is required for normal osteogenesis and hematopoiesis. Another important function of ERF is acting as a tumor suppressor by antagonizing oncogenic fusions involving other ETS family factors. The structure of ERF and the DNA binding properties specific to this protein have not been elucidated. In this study, we determined two crystal structures of the complexes of the DNA binding domain of ERF with DNA. In one, ERF is in a distinct dimeric form, with Cys72 in a reduced state. In the other, two dimers of ERF are assembled into a tetramer that is additionally locked by two Cys72-Cys72 disulfide bonds across the dimers. In the tetramer, the ERF molecules are bound to a pseudocontinuous DNA on the same DNA face at two GGAA binding sites on opposite strands. Sedimentation velocity analysis showed that this tetrameric assembly forms on continuous DNA containing such tandem sites spaced by 7 bp. Our bioinformatic analysis of three previously reported sets of ERF binding loci across entire genomes showed that these loci were enriched in such 7 bp spaced tandem sites. Taken together, these results strongly suggest that the observed tetrameric assembly is a functional state of ERF in the human cell.
ETS家族转录因子控制着人类不同细胞类型的发育，而这些蛋白质的失调会导致严重的发育综合征和癌症。ERF是ETS家族中少数已知仅作为阻遏物发挥作用的成员之一，正常的骨生成和造血过程需要它。ERF的另一个重要功能是通过拮抗涉及其他ETS家族因子的致癌融合来充当肿瘤抑制因子。ERF的结构及其特有的DNA结合特性尚未阐明。在本研究中，我们确定了ERF的DNA结合结构域与DNA形成的复合物的两种晶体结构。在一种结构中，ERF呈独特的二聚体形式，半胱氨酸72处于还原状态。在另一种结构中，两个ERF二聚体组装成一个四聚体，该四聚体还通过跨二聚体的两个半胱氨酸72-半胱氨酸72二硫键锁定。在四聚体中，ERF分子在相对链上的两个GGAA结合位点处结合于同一DNA面上的假连续DNA。沉降速度分析表明，这种四聚体组装在含有由7个碱基对隔开的此类串联位点的连续DNA上形成。我们对先前报道的三组跨全基因组的ERF结合位点进行的生物信息学分析表明，这些位点富含此类间隔7个碱基对的串联位点。综上所述，这些结果强烈表明，观察到的四聚体组装是ERF在人类细胞中的一种功能状态。
