Mapping the nuclear landscape with multiplexed super-resolution fluorescence microscopy.

Area of Science:

• Cell Biology
• Molecular Biology
• Genomics

Background:

• The cell nucleus orchestrates vital cellular functions.
• Understanding spatial organization of nuclear components like DNA, protein complexes, and epigenetic marks is crucial.
• Existing imaging techniques have limitations in resolving structures across multiple length scales.

Purpose of the Study:

• To develop a multiplexed imaging protocol for high-precision localization of multiple nuclear targets.
• To investigate the spatial organization of active and repressive nuclear states.
• To explore the relationship between molecular organization and nuclear function.

Main Methods:

• Developed a multiplexed imaging protocol.
• Localized 13 different nuclear targets with nanometer precision in single cells.
• Analyzed spatial correlations of nuclear components at micro- and nanoscale.

Main Results:

• Nuclear states exist on a spectrum of length scales, with distinct nanoscale organizational principles.
• Heterochromatin and euchromatin exhibit unique nanoscale organization.
• Correlation of Heterochromatin protein 1-alpha (HP1-α) with DNA varied by scale.
• RNA Polymerase II, p300, and CDK9 showed scale-dependent partitioning.
• Perturbations in histone acetylation or transcription affected nanoscale organization.

Conclusions:

• The developed imaging and analysis pipeline provides high-resolution insights into nuclear organization.
• Nuclear organization principles differ significantly at the micro- and nanoscale.
• This approach can be applied to study other cellular compartments.