CREST Center for Cellular and Biomolecular Machines, University of California Merced, Merced, CA 95343, USA.
Department of Bioengineering, University of California Merced, Merced, CA 95343, USA.
Nucleic Acids Res. 2024 Oct 14;52(18):11098-11114. doi: 10.1093/nar/gkae790.
Most DNA scanning proteins uniquely recognize their cognate sequence motif and slide on DNA assisted by some sort of clamping interface. The pioneer transcription factors that control cell fate in eukaryotes must forgo both elements to gain access to DNA in naked and chromatin forms; thus, whether or how these factors scan naked DNA is unknown. Here, we use single-molecule techniques to investigate naked DNA scanning by the Engrailed homeodomain (enHD) as paradigm of highly promiscuous recognition and open DNA binding interface. We find that enHD scans naked DNA quite effectively, and about 200000-fold faster than expected for a continuous promiscuous slide. To do so, enHD scans about 675 bp of DNA in 100 ms and then redeploys stochastically to another location 530 bp afar in just 10 ms. During the scanning phase enHD alternates between slow- and medium-paced modes every 3 and 40 ms, respectively. We also find that enHD binds nucleosomes and does so with enhanced affinity relative to naked DNA. Our results demonstrate that pioneer-like transcription factors can in principle do both, target nucleosomes and scan active DNA efficiently. The hybrid scanning mechanism used by enHD appears particularly well suited for the highly complex genomic signals of eukaryotic cells.
大多数 DNA 扫描蛋白特异性地识别其同源序列基序，并在某种夹钳界面的辅助下在 DNA 上滑动。在真核生物中控制细胞命运的先驱转录因子必须放弃这两个元素才能进入裸露和染色质形式的 DNA；因此，这些因子是否以及如何扫描裸露的 DNA是未知的。在这里，我们使用单分子技术研究了 Engrailed 同源域（enHD）作为高度混杂识别和开放 DNA 结合界面范例的裸露 DNA 扫描。我们发现 enHD 非常有效地扫描裸露的 DNA，其速度比连续混杂滑动的预期速度快约 200000 倍。为了做到这一点，enHD 在 100 毫秒内扫描约 675 bp 的 DNA，然后在 10 毫秒内随机重新部署到 530 bp 远的另一个位置。在扫描阶段，enHD 分别以慢和中速模式交替，每 3 和 40 毫秒一次。我们还发现 enHD 结合核小体，并且与裸露 DNA 相比，结合核小体的亲和力增强。我们的结果表明，先驱转录因子原则上可以同时做到这两点，即靶向核小体和有效地扫描活性 DNA。enHD 所使用的混合扫描机制似乎特别适合真核细胞高度复杂的基因组信号。
