美国斯坦福大学Paul A. Khavari研究组发现通过附加型募集的近端生物素化(PROBER)，识别与活细胞中可编程序列特异性 DNA 相关的蛋白质。这一研究成果发表在2022年8月4日出版的国际学术期刊《自然—方法学》上。

为了提高识别和量化与活细胞中特定 DNA 序列相关的蛋白质的速度和信噪比 (SNR)，他们开发了通过附加型募集的近端生物素化(PROBER)。 PROBER 使用高拷贝附加体来放大 SNR，并使用邻近蛋白质组学 (BioID) 来识别与感兴趣的短 DNA 序列相关的转录因子和其他基因调节因子。PROBER 量化了转录因子及相应染色质调节因子，与靶 DNA 序列和结合数量性状基因座的组成型和诱导型关联。

PROBER 确定了由于 hTERT 启动子中的癌症热点突变引起的调节器关联的改变，表明这些突变增加了启动子与特定基因激活剂的关联。PROBER 提供了一种快速识别与活细胞中特定 DNA 序列及其变体相关的蛋白质的方法。

据介绍，DNA-蛋白质相互作用介导生理基因调控，并可能被与多基因疾病相关的 DNA 变体改变。

附：英文原文

Title: PROBER identifies proteins associated with programmable sequence-specific DNA in living cells

Author: Mondal, Smarajit, Ramanathan, Muthukumar, Miao, Weili, Meyers, Robin M., Rao, Deepti, Lopez-Pajares, Vanessa, Siprashvili, Zurab, Reynolds, David L., Porter, Douglas F., Ferguson, Ian, Neela, Poornima, Zhao, Yang, Meservey, Lindsey M., Guo, Margaret, Yang, Yen-Yu, Li, Lin, Wang, Yinsheng, Khavari, Paul A.

Issue&Volume: 2022-08-04

Abstract: DNA–protein interactions mediate physiologic gene regulation and may be altered by DNA variants linked to polygenic disease. To enhance the speed and signal-to-noise ratio (SNR) in the identification and quantification of proteins associated with specific DNA sequences in living cells, we developed proximal biotinylation by episomal recruitment (PROBER). PROBER uses high-copy episomes to amplify SNR, and proximity proteomics (BioID) to identify the transcription factors and additional gene regulators associated with short DNA sequences of interest. PROBER quantified both constitutive and inducible association of transcription factors and corresponding chromatin regulators to target DNA sequences and binding quantitative trait loci due to single-nucleotide variants. PROBER identified alterations in regulator associations due to cancer hotspot mutations in the hTERT promoter, indicating that these mutations increase promoter association with specific gene activators. PROBER provides an approach to rapidly identify proteins associated with specific DNA sequences and their variants in living cells. PROBER offers a method to identify proteins associated with the DNA sequence of interest and quantify differential binding caused by sequence variants in living cells.

DOI: 10.1038/s41592-022-01552-w

Source: https://www.nature.com/articles/s41592-022-01552-w