使用纳米流控装置定量测定两种 5-甲基胞嘧啶结合蛋白的 DNA 环化。
DNA looping by two 5-methylcytosine-binding proteins quantified using nanofluidic devices.
机构信息
Department of Physics, North Carolina State University, Raleigh, NC, 27695-8202, USA.
Department of Pathology and Laboratory Medicine, University of North Carolina, Chapel Hill, NC, 27599-7525, USA.
出版信息
Epigenetics Chromatin. 2020 Mar 16;13(1):18. doi: 10.1186/s13072-020-00339-7.
BACKGROUND
MeCP2 and MBD2 are members of a family of proteins that possess a domain that selectively binds 5-methylcytosine in a CpG context. Members of the family interact with other proteins to modulate DNA packing. Stretching of DNA-protein complexes in nanofluidic channels with a cross-section of a few persistence lengths allows us to probe the degree of compaction by proteins.
RESULTS
We demonstrate DNA compaction by MeCP2 while MBD2 does not affect DNA configuration. By using atomic force microscopy (AFM), we determined that the mechanism for compaction by MeCP2 is the formation of bridges between distant DNA stretches and the formation of loops.
CONCLUSIONS
Despite sharing a similar specific DNA-binding domain, the impact of full-length 5-methylcytosine-binding proteins can vary drastically between strong compaction of DNA and no discernable large-scale impact of protein binding. We demonstrate that ATTO 565-labeled MBD2 is a good candidate as a staining agent for epigenetic mapping.
背景
MeCP2 和 MBD2 是一类具有选择性结合 CpG 环境中 5-甲基胞嘧啶的结构域的蛋白家族成员。该家族成员与其他蛋白相互作用以调节 DNA 包装。在几纳牛力学长度的纳米流道中拉伸 DNA-蛋白复合物，使我们能够探测蛋白的紧凑程度。
结果
我们证明了 MeCP2 对 DNA 的压缩作用，而 MBD2 不影响 DNA 构象。通过原子力显微镜（AFM），我们确定了 MeCP2 压缩的机制是在远距离 DNA 伸展之间形成桥，并形成环。
结论
尽管具有相似的特定 DNA 结合结构域，但全长 5-甲基胞嘧啶结合蛋白的影响可能在 DNA 的强烈压缩和蛋白结合的无明显大规模影响之间有很大差异。我们证明，ATTO 565 标记的 MBD2 是用于表观遗传学图谱的良好染色剂候选物。
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