Cellular response to heat shock studied by multiconfocal fluorescence correlation spectroscopy

Area of Science:

• Cellular Biology
• Molecular Biology
• Biophysics

Background:

• Heat shock induces a protective cellular response mediated by heat shock factor 1 (HSF1).
• Understanding HSF1 dynamics is crucial for comprehending cellular stress adaptation.
• Existing methods have limitations in precisely measuring HSF1 dynamics in living cells.

Purpose of the Study:

• To develop and utilize a novel multiconfocal fluorescence correlation spectroscopy (FCS) system.
• To quantitatively measure the dynamic behavior of HSF1 in response to heat shock.
• To investigate the intracellular variability of HSF1 dynamics within the cell nucleus.

Main Methods:

• Development of a multiconfocal FCS setup incorporating a spatial light modulator and an electron-multiplying CCD camera.
• Analysis of autocorrelation curves from eGFP and HSF1-eGFP expressing cells with high temporal resolution (14 μs).
• Application of a diffusion-and-binding model to evaluate dynamic parameters of HSF1.

Main Results:

• Heat shock significantly reduces the diffusion rate of HSF1 within the cell nucleus.
• HSF1 dissociation rate from binding partners decreases post-heat shock, while association rate remains unchanged.
• Intracellular variability of HSF1 diffusion increases after heat shock, exceeding that of inert eGFP.

Conclusions:

• The observed changes in HSF1 dynamics, including slower diffusion and altered binding kinetics, support increased interactions with DNA during heat shock.
• The findings cannot be attributed to heat-induced alterations in nuclear organization alone.
• Multiconfocal FCS provides a powerful tool for dissecting the complex dynamics of transcription factors in response to cellular stimuli.