Super-Resolution Compatible DNA Labeling Technique Reveals Chromatin Mobility and Organization Changes During...

Area of Science:

• Cell Biology
• Neuroscience
• Genetics

Background:

• Chromatin dynamics are vital for cellular differentiation, but studying global chromatin mobility in live cells is challenging.
• Existing tools for observing chromatin behavior during differentiation are limited.

Purpose of the Study:

• To develop and validate a novel metabolic labeling probe for tracking chromatin dynamics in living cells.
• To investigate changes in chromatin mobility and organization during neural differentiation.

Main Methods:

• Metabolic labeling of chromatin using a novel silicon rhodamine-conjugated deoxycytidine triphosphate (dC^SiRTP) delivered by a synthetic transporter (SNTT1).
• Correlative confocal and STED super-resolution microscopy for quantifying chromatin domain sizes.
• Time-lapse super-resolution microscopy with single particle tracking to analyze chromatin mobility.
• Micrococcal nuclease digestion assays, chromatin compaction, and histone modification analyses.

Main Results:

• The dC^SiRTP probe was successfully delivered into human induced pluripotent stem cells (iPSCs) and neural stem cells (NSCs).
• Chromatin mobility significantly decreased during the transition from iPSCs to neurons.
• Neuronal differentiation correlated with reduced chromatin accessibility and increased heterochromatin-associated histone modifications.

Conclusions:

• The developed probe enables real-time tracking of chromatin dynamics in living cells during differentiation.
• Chromatin undergoes structural changes, becoming more compact and less accessible during neurogenesis.
• These findings provide novel insights into the regulation of chromatin organization during cell fate commitment.