Method for analyzing nucleoskeletal components from the isolated nucleus

Multiple skeletal components, including actin and associated proteins, are located in both the cytoplasm and the nucleus. Here, we describe a methodology to study nuclear actin and its associated proteins through a comprehensive approach. First, we focus on nuclear isolation as a singular step to separate and distinguish skeletal components within the nucleus. Nuclear actin organization can be regulated by using pharmacological agents (e.g., latrunculin, jasplakinolide) or genetic tools, such as mutants of nuclear actin (NLS-actinR62D). These approaches allow us to analyze the nuclear morphology, protein localization, and structural changes. Together, the development of advanced super-resolution microscopy, live-cell imaging, and biochemical approaches continues to refine our understanding of nuclear actin structure and dynamics.