Derganc, J. | bioRxiv Science

bioRxiv2020

Exceptional stability of a perilipin on lipid droplets depends on its polar residues, suggesting multimeric assembly

Numerous proteins target lipid droplets (LDs) through amphipathic helices (AHs). It is generally assumed that AHs insert bulky hydrophobic residues in packing defects at the LD surface. However, this model does not explain the targeting of perilipins, the most abundant and specific amphipathic proteins of LDs. The gigantic Plin4 contains a highly repetitive AH that lacks bulky hydrophobic residues, and its LD targeting depends strongly on its length. We show that Plin4 forms a remarkably immobile protein layer at the surface of cellular or artificial LDs, making them stable over days. This Plin4 AH feature is not shared with the AHs of other perilipins, which display much faster dynamics on lipid surfaces. Plin4 AH stability on LDs is exquisitely sensitive to the nature and distribution of its polar residues. These results suggest that Plin4 forms stable arrangements of adjacent AHs via polar interactions.

cell biology↗

bioRxiv2020

Cortical stiffness of keratinocytes measured by lateral indentation with optical tweezers

Keratin intermediate filaments are the principal structural element of epithelial cells. Their importance in providing bulk cellular stiffness is well recognized, but their role in the mechanics of cell cortex is less understood. In this study, we therefore compared the cortical stiffness of three keratinocyte lines: primary wild type cells (NHEK2), immortalized wild type cells (NEB1) and immortalized mutant cells (KEB7). The cortical stiffness was measured by lateral indentation of cells with AOD-steered optical tweezers without employing any moving mechanical elements. The method was validated on fixed cells and Cytochalasin-D treated cells to ensure that the observed variations in stiffness within a single cell line were not a consequence of low measurement precision. The measurements of the cortical stiffness showed that primary wild type cells were significantly stiffer than immortalized wild type cells, which was also detected in previous studies of bulk elasticity. In addition, a small difference between the mutant and the wild type cells was detected, showing that mutation of keratin impacts also the cell cortex. Thus, our results indicate that the role of keratins in cortical stiffness is not negligible and call for further investigation of the mechanical interactions between keratins and elements of the cell cortex.

biophysics↗