Nanodomain organization of rhodopsin in native human and murine rod outer segment disc membranes

Abstract

Biological membranes display distinct domains that organize membrane proteins and signaling molecules to facilitate efficient and reliable signaling. The organization of rhodopsin, a G protein-coupled receptor, in native rod outer segment disc membranes was investigated by atomic force microscopy. Atomic force microscopy revealed that rhodopsin is arranged into domains of variable size, which we refer to herein as nanodomains, in native membranes. Quantitative analysis of 150 disc membranes revealed that the physical properties of nanodomains are conserved in humans and mice and that the properties of individual disc membranes can be variable. Examining the variable properties of disc membranes revealed some of the factors contributing to the size of rod outer segment discs and the formation of nanodomains in the membrane. The diameter of rod outer segment discs was dependent on the number of rhodopsin molecules incorporated into the membrane but independent of the spatial density of rhodopsin. The number of nanodomains present in a single disc was also dependent on the number of rhodopsin molecules incorporated into the membrane. The size of the nanodomains was largely independent of the number or spatial density of rhodopsin in the membrane.

Impact of Reduced Rhodopsin Expression on the Structure of Rod Outer Segment Disc Membranes

“…Atomic force microscopy (AFM) can provide much needed insights in these areas because it allows for the direct visualization of membrane proteins within the native environment of a lipid bilayer and physiological buffer conditions 12–14 . AFM has revealed that rhodopsin is organized into domains within ROS disc membranes that are composed of rows of dimeric receptor 15–17 . These domains are referred to as nanodomains since the dimensions are less than 100 nm 17 .…”

Nanodomains in Biomembranes with Recycling

“…Some works argue that even though a membrane is maintained out-ofequilibrium by active processes, equilibrium statistical mechanics is sufficient to account for the finite size of membrane nano-domains [21,18,22,19]. For example, the observation of nano-domains of rhodopsin (a GPCR) in cadavers' retina cells [8], very similar to those observed in fixed cells or membrane sheets, suggests that active processes are not essential. Alternatively, some other works propose that out-of-equilibrium mechanisms are necessarily at play, driven by active membrane recycling [25,29,14,30,31,26,32].…”

“…Since proteins represent about one half of the membrane mass, the protein surface fraction φ is on the same order of magnitude, even though certainly slightly smaller because many proteins protrude out of the membrane plane. Accepted values lie around φ = 0.2 [8,10]. We shall explore the range φ = 0.01 to 0.3 in this work and the SI.…”

“…E-cadherins constitute another example for which nano-patterning of the membrane has been shown to be involved in its biological function, clusters serving as adhesive foci in adherens junctions [14,15]. Grouping identical receptors in a same cluster can also optimize and make more reliable their response to external stimuli [6,8,16,17,19]. Clusters could also result in the inactivation or storage of proteins that would otherwise interact with their environment in a possibly unfavorable way.…”

“…The organization in nano-domains had been suspected for long, but a variety of super-resolution light microscopy techniques and atomic-force microscopy (AFM) have recently improved further our understanding of membrane supramolecular organization. They gave definitive evidence of the generic existence of nanometer sized functional domains on the cell membrane [2,4,5,6,7,8,9,10].…”