Single-cell dynamics of genome-nucleolus interactions captured by nucleolar laser microdissection (NoLMseq)

Area of Science:

• * Molecular Biology
• * Genomics
• * Cell Biology

Background:

• * Gene positioning within the nucleus is crucial for gene regulation, with repressed chromatin often associating with the nuclear lamina or nucleoli.
• * The nucleolus changes structure in different cellular states, potentially affecting genome organization, but its influence on 3D genome architecture is poorly understood.
• * Existing methods struggle to map nucleolar-associated domains (NADs) in single cells or under nucleolar stress conditions.

Purpose of the Study:

• * To develop a novel method for mapping NADs in single cells.
• * To investigate the impact of nucleolar states and stress on 3D genome architecture.
• * To explore the relationship between NADs, gene expression, and chromatin states.

Main Methods:

• * Development of NoLMseq (Nucleolar-associated domain Mapping by sequencing), a technique combining laser-capture microdissection and DNA sequencing.
• * Application of NoLMseq to map NADs in single mouse embryonic stem cells.
• * Analysis of chromosome organization and NAD heterogeneity under normal and nucleolar stress conditions.

Main Results:

• * NoLMseq successfully mapped NADs in single cells, revealing significant heterogeneity in mouse embryonic stem cells with two distinct populations.
• * NADs were found to predominantly contact nucleoli monoallelically, with contact frequency linked to gene expression and chromatin states.
• * Nucleolar stress induced extensive chromosome reorganization, underscoring the nucleolus's role in maintaining genome organization.

Conclusions:

• * NoLMseq is a powerful new tool for studying single-cell NADs and 3D genome organization.
• * Nucleolar states and integrity are critical for maintaining genome architecture.
• * The findings provide insights into how cellular states and stress influence genome organization, with implications for health and disease.