The p65/RelA subunit of NF-kappaB interacts with actin-containing structures

Area of Science:

• Cell Biology
• Molecular Biology
• Cytoskeletal Dynamics

Background:

• Nuclear factor-kappa B (NF-kappaB) is a key transcription factor regulating gene expression.
• NF-kappaB is typically sequestered in the cytoplasm in unstimulated cells.
• The precise cytoplasmic localization and potential non-transcriptional roles of NF-kappaB are not well understood.

Purpose of the Study:

• To investigate the effect of actin cytoskeleton organization on NF-kappaB distribution within cells.
• To explore the interaction between the p65/RelA subunit of NF-kappaB and actin filaments.
• To identify potential novel cytoplasmic functions of NF-kappaB.

Main Methods:

• Immunofluorescence microscopy to visualize p65 localization in normal and transformed fibroblasts.
• Treatment with Cytochalasin D to disrupt actin filaments.
• Affinity chromatography using F-actin to assess direct binding of p65 to actin.

Main Results:

• The p65 subunit of NF-kappaB localizes to focal contacts and stress fibers in normal fibroblasts.
• Actin reorganization in transformed cells leads to p65 redistribution to focal contacts and stress fibers.
• p65 accumulates at leading lamellae edges and cell-to-cell contacts under specific conditions.
• Cytochalasin D treatment causes p65 to aggregate within actin-containing structures.
• Direct binding of p65 to filamentous actin was confirmed via affinity chromatography.

Conclusions:

• Cytoplasmic distribution of the p65 subunit of NF-kappaB is dependent on the state of the actin cytoskeleton.
• NF-kappaB (p65) directly interacts with filamentous actin.
• These findings suggest a previously unrecognized cytoplasmic role for NF-kappaB, linked to cytoskeletal dynamics.