The lamin A/C Ig-fold undergoes cell density-dependent changes that alter epitope binding.

Area of Science:

• Cell Biology
• Biochemistry
• Biophysics

Background:

• Lamins A/C are nuclear intermediate filament proteins crucial for cellular mechanics and biochemistry.
• Antibody-based detection is vital for studying Lamin A/C localization and function.

Purpose of the Study:

• To investigate the cell density-dependent recognition of Lamins A/C by antibodies.
• To understand the underlying mechanisms and implications for immunofluorescence studies.

Main Methods:

• Immunofluorescence using JOL-2 antibody targeting Lamin A/C Ig-fold.
• Cell density manipulation and cell spreading assays.
• Cytoskeletal disruption and Linker of Nucleoskeleton and Cytoskeleton (LINC) complex perturbation.
• Nuclear stiffness and force transmission measurements.

Main Results:

• Antibody JOL-2 recognition of Lamin A/C is highly dependent on cell density, despite unchanged Lamin A/C levels.
• Proposed mechanism involves conformational changes (unfolding/masking) of Ig-fold loops (C'E, EF) upon cell spreading.
• Labeling was unaffected by cytoskeletal or LINC complex disruption.
• Nuclear stiffness and force transmission remained constant across cell densities.

Conclusions:

• Cell density influences Lamin A/C antibody accessibility, likely via conformational changes.
• Findings necessitate careful interpretation of immunofluorescence data for Lamins A/C.
• Conformational plasticity of Lamins A/C may be functionally relevant in cellular processes.