Dynamic mechanochemical feedback between curved membranes and BAR protein self-organization

In many physiological situations, BAR proteins reshape membranes with pre-existing curvature (templates), contributing to essential cellular processes. However, the mechanism and the biological implications of this reshaping process remain unclear. Here we show, both experimentally and through modelling, that BAR proteins reshape low curvature membrane templates through a mechanochemical phase transition. This phenomenon depends on initial template shape and involves the co-existence and progressive transition between distinct local states in terms of molecular organization (protein arrangement and density) and membrane shape (template size and spherical versus cylindrical curvature). Further, we demonstrate in cells that this phenomenon enables a mechanotransduction mode, in which cellular stretch leads to the mechanical formation of membrane templates, which are then reshaped into tubules by BAR proteins. Our results demonstrate the interplay between membrane mechanics and BAR protein molecular organization, integrating curvature sensing and generation in a comprehensive framework with implications for cell mechanical responses.

Membrane Constriction by ESCRT-III Proteins Induces Structural Lipid Bilayer Asymmetries

Cells utilize molecular machines to form and remodel their membrane-defined compartments' compositions, shapes, and connections. The regulated activity of these membrane remodeling machines drives processes like vesicular traffic and organelle homeostasis. Although molecular patterning within membranes is essential to cellular life, characterizing the composition and structure of realistic biological membranes on the molecular length scale remains a challenge, particularly during membrane shape transformations. Here, we employed an ESCRT-III protein coating model system to investigate how membrane-binding proteins bind to and alter the structural patterns within lipid bilayers. We observe leaflet-level and localized lipid structures within a constricted and thinned membrane nanotube. To map the fine structure of these membranes, we compared simulated bilayer nanotubes with experimental cryo-EM reconstructions of native membranes and membranes containing halogenated lipid analogs. Halogenated lipids scatter electrons more strongly, and analysis of their surplus scattering enabled us to estimate the concentrations of lipids within each leaflet and to estimate lipid shape and sorting changes induced by high curvature and lipid-protein interactions. Specifically, we found that cholesterol enriched within the inner leaflet due to its spontaneous curvature, while acidic lipids enriched in the outer leaflet due to electrostatic interactions with the protein coat. The docosahexaenoyl (DHA) polyunsaturated chain-containing lipid SDPC enriched strongly at membrane-protein contact sites. Simulations and imaging of brominated SDPC showed how a pair of phenylalanine residues opens a hydrophobic defect in the outer leaflet and how DHA tails stabilize the defect and "snorkel" up to the membrane surface to interact with these side chains. This highly curved nanotube differs markedly from protein-free, flat bilayers in leaflet thickness, lipid diffusion, and other structural asymmetries with implications for our understanding of membrane mechanics.

The identification of strain-stress curve for 5049 aluminum based on tube hydraulic bulging test

Tube hydraulic bulging tests with fixed-end conditions are carried out to explore tubular material characteristics for 5049 aluminium. Tube diameter at the center of specimen and pole thickness under different internal pressures are recorded during forming process. Based on experimental data, two types of theoretical models using membrane mechanics and total strain theory are applied to determine the flow stress curve of tubular specimens. A tension specimen is cut from the same tube along longitudinal direction and strain-stress curve is fitted by a universal tensile test. In order to test their accuracy, obtained material parameters from three methods are imported into a finite element model (FEM) and its predicted results are compared with bugle height measured from experiments. The comparison shows that the flow stress curve of 5049 aluminium tube can be identified by these three methods and simulated results from total strain model has a better agreement with experimental measures compared with the other two methods.

To form and function: on the role of basement membrane mechanics in tissue development, homeostasis and disease

The basement membrane (BM) is a special type of extracellular matrix that lines the basal side of epithelial and endothelial tissues. Functionally, the BM is important for providing physical and biochemical cues to the overlying cells, sculpting the tissue into its correct size and shape. In this review, we focus on recent studies that have unveiled the complex mechanical properties of the BM. We discuss how these properties can change during development, homeostasis and disease via different molecular mechanisms, and the subsequent impact on tissue form and function in a variety of organisms. We also explore how better characterization of BM mechanics can contribute to disease diagnosis and treatment, as well as development of better in silico and in vitro models that not only impact the fields of tissue engineering and regenerative medicine, but can also reduce the use of animals in research.

Abstract 13664: Membrane Stiffness as a Function of Plakophilin-2 Expression: Implications to Arrhythmogenic Right Ventricular Cardiomyopathy

Mutations in PKP2 , the gene coding for the desmosomal protein Plakophilin-2 (PKP2), can lead to an inheritable cardiac disease called Arrhythmogenic Right Ventricle Cardiomyopathy (ARVC). Various studies investigated the molecular and electrical properties of cardiomyocytes (CMs) with PKP2 deficiency. However, systematic studies on the mechanical properties of PKP2-deficient CMs, and their relation to the cardiomyopathic phenotype, are lacking. We studied the relation between PKP2 expression, membrane transverse Young’s modulus (tYM; a surrogate measure of membrane stiffness), and their correlation with tubulin expression and with substrate stiffness. Left and right ventricular (LV, RV) CMs were isolated from murine hearts control (ctrl), or with a cardiomyocyte-specific, tamoxifen-activated knockout of PKP2 (PKP2cKO). Three time points were studied: 14, 15 and 21 days post-tamoxifen injection (dpi). The phenotypes at these time points correspond to three disease stages: concealed, arrhythmogenic and cardiomyopathy of RV predominance, respectively. tYM was evaluated by mechanoSICM. Tubulin was visualized by confocal microscopy. tYM in LV myocytes from control hearts showed a tendency toward higher values when compared to RV myocytes (LV 2.9±0.2 kPa vs RV 2.6±0.2 kPa, n=53-59). At 14 dpi, LV and RV PKP2cKO CMs were softer than ctrl (LV 2.1±0.1 kPa, RV 2.1±0.1 kPa vs ctrl, n=30-59). The trend reversed a day later and by 21 dpi there was a clear increase in tYM, particularly in PKP2cKO RV myocytes (LV 4.5±0.3 kPa, RV 4.8±0.6 kPa vs ctrl, n=44-59). Z-groove index, a parameter of membrane organization, decreased in PKP2cKO CMs. Increased tYM at 21 dpi corresponded to upregulation of α-tubulin and particularly, acetylated tubulin, which was reduced at 14dpi but went up in 21dpi. We also observed an inverse correlation between membrane tYM, and substrate stiffness. We conclude that loss of PKP2 expression affects, in a disease stage-specific manner, CM mechanical properties and the MT network organization. Whether these effects are correlative or one is consequence of the other, remains to be determined. We speculate that changes in membrane mechanics at the single cell level are a component of mechanical dysfunction in hearts affected with ARVC.

A role for tectorial membrane mechanics in activating the cochlear amplifier

The mechanical and electrical responses of the mammalian cochlea to acoustic stimuli are nonlinear and highly tuned in frequency. This is due to the electromechanical properties of cochlear outer hair cells (OHCs). At each location along the cochlear spiral, the OHCs mediate an active process in which the sensory tissue motion is enhanced at frequencies close to the most sensitive frequency (called the characteristic frequency, CF). Previous experimental results showed an approximate 0.3 cycle phase shift in the OHC-generated extracellular voltage relative the basilar membrane displacement, which was initiated at a frequency approximately one-half octave lower than the CF. Findings in the present paper reinforce that result. This shift is significant because it brings the phase of the OHC-derived electromotile force near to that of the basilar membrane velocity at frequencies above the shift, thereby enabling the transfer of electrical to mechanical power at the basilar membrane. In order to seek a candidate physical mechanism for this phenomenon, we used a comprehensive electromechanical mathematical model of the cochlear response to sound. The model predicts the phase shift in the extracellular voltage referenced to the basilar membrane at a frequency approximately one-half octave below CF, in accordance with the experimental data. In the model, this feature arises from a minimum in the radial impedance of the tectorial membrane and its limbal attachment. These experimental and theoretical results are consistent with the hypothesis that a tectorial membrane resonance introduces the correct phasing between mechanical and electrical responses for power generation, effectively turning on the cochlear amplifier.