Hierarchical mesoscale domain organization of the plasma membrane Akihiro Kusumi 1, 2 , Kenichi G.N. Suzuki 1, 3 , Rinshi S. Kasai 2 , Ken Ritchie 4 and Takahiro K. Fujiwara 1 1 Institute for Integrated Cell-Material Sciences (iCeMS), Kyoto University, Kyoto 606-8507, Japan 2 Institute for Frontier Medical Sciences, Kyoto University, Kyoto 606-8507, Japan 3 Precursory Research for Embryonic Science and Technology (PRESTO), Japan Science and Technology Agency (JST), iCeMS, Kyoto University, Kyoto 606-8507, Japan 4 Department of Physics, Purdue University, West Lafayette, IN 47907, USA Based on recent single-molecule imaging results in the living cell plasma membrane, we propose a hierarchical architecture of three-tiered mesoscale (2–300 nm) domains to represent the fundamental functional orga- nization of the plasma membrane: (i) membrane com- partments of 40–300 nm in diameter due to the partitioning of the entire plasma membrane by the ac- tin-based membrane skeleton ‘fence’ and transmem- brane protein ‘pickets’ anchored to the fence; (ii) raft domains (2–20 nm); and (iii) dimers/oligomers and greater complexes of membrane-associated proteins (3–10 nm). The basic molecular interactions required for the signal transduction function of the plasma mem- brane can be fundamentally understood and convenient- ly summarized as the cooperative actions of these mesoscale domains, where thermal fluctuations/move- ments of molecules and weak cooperativity play crucial roles. The important thing in science is not so much to obtain new facts as to discover new ways of thinking about them. William Lawrence Bragg Three-tiered architecture of the plasma membrane According to the fluid mosaic model proposed by Singer and Nicolson in 1972 [1], the cellular plasma membrane is a two-dimensional liquid, in which the lipid and protein molecules impregnated in the plasma membrane are mixed like a mosaic and undergo thermal diffusion. This model is still believed to represent the basic structure of the plasma membranes of all cells existing on earth. Such universality is comparable to that of the double helical structure of DNA. If the Singer–Nicolson model were en- tirely true, then all of the membrane molecules would be homogeneously distributed throughout the plasma mem- brane, undergoing simple Brownian diffusion. However, membrane researchers continue to uncover evidence show- ing that this is not the case [2]. In this review, we address the fundamental questions regarding the mechanisms, or general rules, for organizing membrane constituent molecules in plasma membranes. Specifically, we will discuss the three types of membrane inhomogeneities, or membrane domains, in the mesoscale in the plasma membrane, which form the basis for its hierarchical order. Here, we define the mesoscale as ap- proximately 2–300 nm (i.e. greater than a nanometer and smaller than a micron). One of the crucial reasons for evoking the mesoscale concept is that it is a scale in which both thermal fluctuation and weak cooperativity could play important roles. We try to avoid the nanoscale concept because it gives the impression of static structures and lack of cooperativity, owing to the small numbers of molecules involved. We hope to convey the idea of dynamic, restless and cooperative domains and molecular complexes, which continually form and disperse within the plasma mem- brane on various time scales. Furthermore, it would be misleading to refer to a domain with a 300-nm diameter as a nanodomain, because it is 300-fold larger than one nanometer. The most basic and largest domain in the hierarchy is the ‘membrane compartment’, which is created via the parti- tioning of the entire plasma membrane, owing to its inter- actions with the actin-based membrane skeleton (fence) and transmembrane (TM) proteins anchored to the membrane skeleton fence (pickets) (Figure 1a). The second in the hierarchy is the ‘raft domain’, created by various levels of molecular affinities and immiscibilities involving both lipids and proteins, as well as cooperative condensations within the plasma membrane, which can be modulated by stimu- lation-induced dimers and oligomers of raft-associable receptors (Figure 1b, [3,4]). The third, smallest domain includes the ‘dimers/oligomers and greater complexes of membrane-associated and integral membrane proteins’. We regard protein complexes as one of the mesodomains (protein complex domains, which are formed without any major involvement of lipids), rather than considering them as molecular complexes entirely different from membrane inhomogeneities and simply calling them protein com- plexes, for the sake of consistency in the definition of the mesoscale domains. The sizes of the third-tier protein com- plexes are largely in the mesoscale, ranging mostly between 3 and 10 nm (Figure 1c). We believe such a definition of mesodomains will clarify the understanding of the mecha- nisms which underlie plasma membrane function. Review Corresponding author: Kusumi, A. (akusumi@frontier.kyoto-u.ac.jp). 604 0968-0004/$ – see front matter ß 2011 Elsevier Ltd. All rights reserved. doi:10.1016/j.tibs.2011.08.001 Trends in Biochemical Sciences, November 2011, Vol. 36, No. 11