Small heat shock proteins (sHSPs) have been demonstrated to interact with lipids and modulate the physical state of membranes across species. Through these interactions, sHSPs contribute to the maintenance of membrane integrity. HSPB1 is a major sHSP in mammals, but its lipid interaction profile has so far been unexplored. In this study, we characterized the interaction between HSPB1 and phospholipids. HSPB1 not only associated with membranes via membrane-forming lipids, but also showed a strong affinity towards highly fluid membranes. It participated in the modulation of the physical properties of the interacting membranes by altering rotational and lateral lipid mobility. In addition, the in vivo expression of HSPB1 greatly affected the phase behavior of the plasma membrane under membrane fluidizing stress conditions. In light of our current findings, we propose a new function for HSPB1 as a membrane chaperone.

Lipid metabolic disturbances are associated with several diseases, such as type 2 diabetes or malignancy. In the last two decades, high-performance mass spectrometry-based lipidomics has emerged as a valuable tool in various fields of biology. However, the evaluation of macroscopic tissue homogenates leaves often undiscovered the differences arising from micron-scale heterogeneity. Therefore, in this work, we developed a novel laser microdissection-coupled shotgun lipidomic platform, which combines quantitative and broad-range lipidome analysis with reasonable spatial resolution. The multistep approach involves the preparation of successive cryosections from tissue samples, cross-referencing of native and stained images, laser microdissection of regions of interest, in situ lipid extraction, and quantitative shotgun lipidomics. We used mouse liver and kidney as well as a 2D cell culture model to validate the novel workflow in terms of extraction efficiency, reproducibility, and linearity of quantification. We established that the limit of dissectible sample area corresponds to about ten cells while maintaining good lipidome coverage. We demonstrate the performance of the method in recognizing tissue heterogeneity on the example of a mouse hippocampus. By providing topological mapping of lipid metabolism, the novel platform might help to uncover region-specific lipidomic alterations in complex samples, including tumors.