"Science Advances" magazine published the latest results of Fudan University's Ding Chen and He Fuchu team to develop a "toolkit" for precise analysis of protein-DNA transcription machinery in the open region of chromatin

Beijing 2021 Nian 10 Yue 23 days, "Science" ( Science ) magazine child journal "Science Progress" ( Science in Advances ), published online the State Key Laboratory of Genetic Engineering, Fudan University, Biomedical Research Institute, Institute of Human phenotype group, College of life sciences Ding Chen / He Fuchu team related protein -DNA latest research transcription machinery " transcriptional regulatory protein machines on the transposon biotinylated accessibility of chromatin ( Proteome-Wide Profiling of transcriptional machinery oN chromatin with Accessible biotinylated transposons ) "
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The results show that the Fudan University team has realized the identification and functional research of the transcriptional protein machinery of the open chromatin region for the first time in the world
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The transcriptional regulatory protein machinery is composed of transcription factors ( transcirption factor , TF ), transcription cofactors ( transcirptional coregulator ,TC ), etc.

At the genome level, in 2013 , Professor Howard Chang of Stanford University developed the ATAC-seq technology ( Assay for Transposase-Accessible Chromatin with highthroughput sequencing ), which can obtain chromatin open regions and active transcriptional regulatory sequences in the genome through high-throughput sequencing.

However, at the proteome level, due to the low expression abundance of transcription factors and transcription co-regulators and the lack of relevant enrichment techniques, the scientific community is still difficult to make a panoramic and fine analysis of the binding and pairing relationship between the above-mentioned protein molecules and the open regions of chromatin

For many years, the Ding Chen / He Fuchu team of the Institute of Biomedical Research of Fudan University and the Institute of Human Phenotype Group has focused on this problem and continued to carry out scientific research

In the latest results published this time, based on ATAC-seq technology , researchers have developed a new method ATAC-MS that directly enriches and quantifies the transcriptional protein machinery of the open region of endogenous chromatin in real-time in situ transcription (Figure 1 )

Figure 1.

This method has good sensitivity and specificity for the identification of transcription factors and transcription co-regulators when using cells as low as 5×105 (Figure 2 )

Figure 2.

After stimulating HeLa cells with tumor necrosis factor ( TNF-α ) , ATAC-MS successfully identified the changes in the transcription factor NFKB family and the TFs that interact with it (Figure 3 ) .

Figure 3.

Figure 4.

Furthermore, the team combined ATAC-MS technology with anion exchange technology to develop high - resolution ATAC-MS analysis technology ( fractionation ATAC-MS , fATAC-MS )

Figure 5.

Furthermore, the author designed and expressed a new type of Tn5-dCas9 fusion protein, which upgraded ATAC-MS to ctATAC-MS (dCas9 targeted ATAC-MS)

Figure 6.

In summary, Fudan Chen D / research team He Fuchu provides this publication includes .
3 Species new method --ATAC-the MS , fATAC-the MS and ctATAC-MS , including the transcriptional regulatory complex analytic " kits " such that We can not only map the active transcription regulation protein machinery at the whole genome level, but also explore protein -DNA transcription regulation complexes at specific genomic sites
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This toolkit will enable researchers in this field to dig deeper into different levels of transregulation mechanisms, and has the characteristics of taking into account in vivo / in vitro reactions.
It has a wide range of potential applications in clinical biopsy, detection of various tissue specimens, and testing of different species.

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The research was supported by the Shanghai Municipal Science and Technology Major Project "International Human Phenotype Group Project (Phase I)"
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Zhang Haizhu, a 2017 doctoral student at the Institute of Biomedical Sciences , Fudan University, Qin Zhaoyu, an associate researcher at the Institute of Human Phenotype Group, Fudan University, and Yue Xuetong, a 2018 doctoral student at the Institute of Biomedical Sciences, Fudan University, are the co-first authors of this article
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Researcher Yang Wenjun, Xinhua Hospital Affiliated to Shanghai Jiaotong University School of Medicine, Academician of the Chinese Academy of Sciences, State Key Laboratory of Genetic Engineering, and Fudan University Biomedical Research Institute Specially-appointed PI and Specially-appointed Human Phenotypic Research Institute Professor He Fuchu, Fudan University Human Phenotypic Research Institute Researcher Ding Chen, associate dean, State Key Laboratory of Genetic Engineering, School of Life Sciences, and Institute of Biomedical Sciences, is the co-corresponding author of the paper
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Link to the paper: https://doi.
org/10.
1126/sciadv.
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