Nuclear lipids in chromatin regulation: Biological roles, experimental approaches and existing challenges

Abstract

Lipids are crucial for various cellular functions. Besides the storage of energy equivalents, these include forming membrane bilayers and serving as signaling molecules. While significant progress has been made in the comprehension of the molecular and cellular biology of lipids, their functions in the cell nucleus remain poorly understood. The main role of the eukaryotic cell nucleus is to provide an environment for the storage and regulation of chromatin which is a complex of DNA, histones, and associated proteins. Recent studies suggest that nuclear lipids play a role in chromatin regulation and epigenetics. Here, we discuss various experimental methods in lipid-chromatin research, including biophysical, structural, and cell biology approaches, pointing out their strengths and weaknesses. We take the view that nuclear lipids have a far more widespread impact on chromatin than is currently acknowledged. This gap in comprehension is mostly due to existing experimental challenges in the study of lipid-chromatin biology. Several new, interdisciplinary approaches are discussed that could aid in elucidating the roles of nuclear lipids in chromatin regulation and gene expression.

Keywords: biophysical methods; cell biology; chromatin regulation; histone lipidation; nuclear lipids.

FIGURE 1

Characterization of lipid‐chromatin interactions. (a) Sphingolipids binding to histone proteins modulate histone modifications. (b) Cholesterol integration to chromatin structures aids in chromatin compaction and gene expression. (c) Phosphatidylinositol interaction with chromatin remodeling complexes regulates chromatin accessibility. (d) Methodologies of lipid‐chromatin interaction characterization including: MST, ITC, NMR, x‐ray crystallization, MS spectrometry, Raman microscopy, and cell biology approaches. For details, see text.

FIGURE 2

Theoretical, integrated pipelines for investigating nuclear protein‐lipid interactions. (a) Sample preparation: Cells are cultured in 96‐well plates, followed by fixation, permeabilization, and incubation with specific antibodies and dyes to label lipids and chromatin, or fluorescent liposomal nanovesicles are used to target lipids. (b) Screening: Fluorescent images are captured across different channels using automated microscopes, or fluorescent liposomal nanovesicles are incubated with a proteome microarray containing purified nuclear proteins. (c) Data analysis: Overlapping fluorescence signals are analyzed to detect potential chromatin‐lipid associations, and microarrays are examined for positive interactions. (d) Identification of candidates: Algorithms are used to identify candidates based on fluorescence overlap, and the data are integrated with microarray results. (e) Kinetics and structural determination: Candidate proteins are subjected to MST to determine binding kinetics (Kd) and analyzed by Cryo‐EM for detailed structural characterization. (f) Chromatin‐anchoring factors and lipid membranes: Lipid membranes are employed to investigate chromatin‐anchoring factors interacting with lipids. (g) Single‐cell omics: Direct lipid manipulation, such as single‐cell injections followed by single‐cell omics can provide further insights into the functional roles of lipids and identified interaction candidates.