Scientific Updates

Developmental Biology | Dynamic transcription regulation at the single-molecule level

  On Dec. 9th, 2021, the Deng Lab from Biomedical Pioneering Innovation Center (BIOPIC), Beijing Advanced Innovation Center for Genomics (ICG), School of Life Sciences in Peking University, published a review entitled “Dynamic transcription regulation at the single-molecule level” in Developmental Biology.


  Cell fate changes during development, differentiation, and reprogramming are controlled mainly at the transcription level. The DNA-binding transcription factors (TFs) often act in a combinatorial fashion to alter chromatin states and drive cell type-specific gene expression. Recent advances in fluorescent microscopy technologies have enabled direct visualization of biomolecules involved in the process of transcription and its regulatory events at the single-molecule level in living cells. Remarkably, imaging and tracking individual TF molecules at high temporal and spatial resolution revealed that they are highly dynamic in searching and cognate binding targets, rather than static and binding constantly (Liu et al., 2015; Lionnet and Wu, 2021). In combination with investigation using techniques from biochemistry, structure biology, genetics, and genomics, a more well-rounded view of transcription regulation is emerging.

  In the first part of the review, the technical aspects of live-cell single-molecule imaging are covered and general experimental pipelines of single-molecule tracking are explained briefly. (Figure 1).


  Figure 1 The experimental pipeline of single-molecule tracking

  The biological relevance and interpretation of the single-molecule dynamic features of transcription regulatory events observed in the native chromatin environment of living eukaryotic cells are next to be focused on. Specifically, the single-molecule dynamics of transcription basal machinery (Figure 2), transcription factors, histones, and chromatin regulators in living cells are covered, showcasing a general dynamic view of these proteins forming transient clusters, rapidly diffusing in the confined nucleus, searching for DNA targets (Figure 3), binding on and off the chromatin. Moreover, single-molecule studies on the heterogenous subnuclear distribution of nuclear proteins, particularly liquid-liquid phase separation is reviewed. Also, how these dynamic features might shed light on mechanistic understanding of transcription regulation are discussed.  


  Figure 2 RNA Pol II forms distinct condensates during transcription initiation, elongation, and splicing, along with Mediator, Cyclin T1, various splicing factors respectively.


  Figure 3 The facilitated diffusion model for efficient target searching mechanism of TFs through (a) 1D sliding, (b) hopping, and (c) intersegmental transfer.

  This review focused on using live-cell single-molecule fluorescence imaging and tracking technology to directly visualize nuclear protein factors diffusing and binding chromatin targets in its native environment. It systematically discussed the single-molecule characteristics of the basal transcription machinery, transcription factors, histones, and chromatin regulatory proteins in living cells, which can actively engage in dynamic clustering, rapid diffusion, target search, and chromatin association and dissociation. Moreover, this review underscored a new perspective of understanding the transcription regulation mechanisms in cell fate determination. This review would stimulate studies of cell fate control by measuring the single-molecule dynamics of determining protein factors.

  Ph.D. candidate Zuhui Wang is the first author of the paper and Prof. Wulan Deng is the co-corresponding author of the paper. The project was funded by the Beijing Advanced Innovation Center for Genomics (ICG) at Peking University, Peking-Tsinghua Center for Life Sciences (CLS), the National Key R&D Program of China, and the National Natural Science Foundation of China.