Elucidation of the Regulatory Mechanisms of Transcriptional Bursting

Introduction

When genes are transcribed within a cell, their activity is not continuous but instead alternates between ON (active) and OFF (inactive) states. For instance, a gene may randomly switch to an ON state for several minutes, producing a burst of RNA, and then abruptly turn OFF. This irregular, dynamic transcription behavior is known as transcriptional bursting and serves as a critical mechanism for fine-tuning gene activity in individual cells (see Figure 1).


Transcriptional bursting leads to variability in gene expression levels even among cells in the same tissue or culture environment. This phenomenon is thought to contribute to diverse cellular behaviors observed during early embryogenesis and cancer evolution, making it extremely important in the context of biological processes and diseases. However, the precise mechanisms that control transcriptional bursting remain insufficiently understood (see Figure 2). Why do some cells exhibit frequent bursts of a particular gene while others remain quiet? What factors determine the timing and duration of these bursts? Answering these questions will open up new avenues in cell biology and gene regulation.


Objective of the Research

This research project aims to elucidate the regulatory mechanisms governing transcriptional bursting. By uncovering the molecular basis behind transcriptional bursting, we can better understand how cells modulate gene expression levels and the inherent fluctuations (noise) in transcription. This knowledge not only deepens our grasp of the fundamental principles of gene expression but also provides clues for explaining the variability observed among cells with identical genetic backgrounds.

Variations in gene expression among cells significantly influence cell fate decisions and stress responses. For example, within a population of undifferentiated stem cells, differences in transcriptional bursting may cause only some cells to express key genes that trigger differentiation, while others remain undifferentiated. Understanding the regulatory mechanisms of transcriptional bursting is therefore crucial for predicting and controlling cell state transitions and fate decisions—an insight that has implications from developmental biology to regenerative medicine. Moreover, the inherent fluctuations resulting from transcriptional bursting may enhance the adaptability of multicellular organisms to environmental changes, thereby maintaining overall organismal health. Conversely, excessive noise might lead to tissue dysfunction or disease; hence, deciphering how cells balance these fluctuations is a central goal of this research.

Research Approach and Methods

To address the complex phenomenon of transcriptional bursting, our laboratory employs a multiscale and multimodal approach by integrating several advanced techniques:

Live Imaging:
We observe living cells in real time to directly visualize the switching between transcription ON and OFF states (i.e., the occurrence of bursts). By using fluorescent protein reporters and in vivo labeling techniques, we can track transcription dynamics on a scale of seconds to minutes and quantitatively assess burst frequency and duration.

Single-Molecule Fluorescence In Situ Hybridization (smFISH):
This method detects individual mRNA molecules in fixed cells. By counting each transcript produced from a gene, smFISH provides a snapshot of the distribution of mRNA numbers in individual cells, allowing us to visualize the variability resulting from transcriptional bursts.

Multimodal seqFISH (DNA/RNA/IF-FISH):
This advanced sequential FISH technique simultaneously visualizes DNA, RNA, and proteins within the same cell. Specifically, it involves sequentially labeling specific genomic regions (e.g., enhancers and promoters), corresponding RNA transcripts, and transcription factors (via immunofluorescence). This approach enables us to observe changes in DNA structure (such as alterations in inter-domain distances) and local accumulation of regulatory factors near genes undergoing transcriptional bursts, thereby revealing the spatial context of burst regulation.

Data from these complementary techniques are integrated to develop mathematical models of transcriptional bursting. For instance, we model burst occurrence using a two-state (ON/OFF) or probabilistic framework and fit these models to our experimental data to estimate key regulatory parameters. This combined experimental and computational approach allows us to comprehensively dissect the phenomenon of transcriptional bursting.

Related Research and Achievements

Our laboratory has been at the forefront of transcriptional bursting research. Recent key publications include:

Future Perspectives

Moving forward, we will expand our approach to examine whether the regulatory factors and mechanisms identified in mouse ES cells are universally applicable across different cell types and developmental stages. For example, we plan to compare the control of transcriptional bursting in various tissues and during distinct phases of development to understand both universal principles and cell type–specific differences.

Moreover, we aim to investigate whether aberrations in transcriptional bursting contribute to disease—such as the increased gene expression variability observed in cancer—and explore interventions that restore normal burst dynamics. Ultimately, insights gained from deciphering the mechanisms controlling transcriptional bursts may lead to innovative strategies for stabilizing or modulating gene expression, with implications for drug discovery and regenerative medicine.

Our goal is to translate these foundational insights into applications that improve early diagnosis, preventive strategies, and treatments for age-related and other diseases.