Junichi Ikenouchi

For epithelia to function as barriers, the intercellular space must be sealed. Sealing two adjacent cells at bicellular tight junctions (bTJs) is well described with the discovery of the claudins. Yet, there are still barrier weak points at tricellular contacts, where three cells join together. In this study, we identify tricellulin, the first integral membrane protein that is concentrated at the vertically oriented TJ strands of tricellular contacts. When tricellulin expression was suppressed w

Snail is a transcription repressor that plays a central role in the epithelium-mesenchyme transition (EMT), by which epithelial cells lose their polarity. Claudins and occludin are integral membrane proteins localized at tight junctions, which are responsible for establishing and maintaining epithelial cell polarity. We examined the relationship between Snail and the promoter activity of claudins and occludin. When Snail was overexpressed in cultured mouse epithelial cells, EMT was induced with

The tricellular tight junction (tTJ) forms at the convergence of bicellular tight junctions (bTJs) where three epithelial cells meet in polarized epithelia, and it is required for the maintenance of the transepithelial barrier. Tricellulin is a four transmembrane domain protein recently identified as the first marker of tTJ, but little is known about how tricellulin is localized at tTJs. As for the molecular mechanism of association of tricellulin with tight junctions (TJs), we found that tricel

The molecular mechanisms of how primordial adherens junctions (AJs) evolve into spatially separated belt-like AJs and tight junctions (TJs) during epithelial polarization are not well understood. Previously, we reported the establishment of ZO-1/ZO-2-deficient cultured epithelial cells (1[ko]/2[kd] cells), which lacked TJs completely. In the present study, we found that the formation of belt-like AJs was significantly delayed in 1(ko)/2(kd) cells during epithelial polarization. The activation of

Tight junctions (TJs) are essential cell adhesion structures that act as a barrier to separate the internal milieu from the external environment in multicellular organisms. Although their major constituents have been identified, it is unknown how the formation of TJs is regulated. TJ formation depends on the preceding formation of adherens junctions (AJs) in epithelial cells; however, the underlying mechanism remains to be elucidated. In this study, loss of AJs in α-catenin-knockout (KO) EpH4 ep

The Par-3/Par-6/aPKC/Cdc42 complex regulates the conversion of primordial adherens junctions (AJs) into belt-like AJs and the formation of linear actin cables during epithelial polarization. However, the mechanisms by which this complex functions are not well elucidated. In the present study, we found that activation of Arf6 is spatiotemporally regulated as a downstream signaling pathway of the Par protein complex. When primordial AJs are formed, Par-3 recruits a scaffolding protein, termed the

Body and organ surfaces in multicellular organisms are covered with a sheet of epithelial cells. The tight junction (TJ) is an adhesive structure that seals the gap between epithelial cells and functions as a selective barrier to prevent the entry of antigens and pathogenic microbes from the extracellular environment. Several transmembrane proteins that constitute the TJ (claudin, occludin, tricellulin and angulin) have been identified. As over-expression of these proteins does not enlarge TJs o

The actin cytoskeleton usually lies beneath the plasma membrane. When the membrane-associated actin cytoskeleton is transiently disrupted or the intracellular pressure is increased, the plasma membrane detaches from the cortex and protrudes. Such protruded membrane regions are called blebs. However, the molecular mechanisms underlying membrane blebbing are poorly understood. This study revealed that epidermal growth factor receptor kinase substrate 8 (Eps8) and ezrin are important regulators of

Adherens junctions (AJs) control epithelial cell behavior, such as collective movement and morphological changes, during development and in disease. However, the molecular mechanism of AJ remodeling remains incompletely understood. Here, we report that the conformational activation of α-catenin is the key event in the dynamic regulation of AJ remodeling. α-catenin activates RhoA to increase actomyosin contractility at cell-cell junctions. This leads to the stabilization of activated α-catenin, i

Cellular architectures require regulated mechanisms to correctly localize the appropriate plasma membrane lipids and proteins. Microvilli are dynamic filamentous-actin-based protrusions of the plasma membrane that are found in the apical membrane of epithelial cells. However, it remains poorly understood how their formation is regulated. In the present study, we found that sphingomyelin clustering underlies the formation of microvilli. Clustering of sphingomyelin is required for the co-clusterin

The role of tight junctions (TJs) in the establishment and maintenance of lipid polarity in epithelial cells has long been a subject of controversy. We have addressed this issue using lysenin, a toxin derived from earthworms, and an influenza virus labeled with a fluorescent lipid, octadecylrhodamine B (R18). When epithelial cells are stained with lysenin, lysenin selectively binds to their apical membranes. Using an artificial liposome, we demonstrated that lysenin recognizes the membrane domai

High-resolution spatio-temporal monitoring of the cell membrane lipid order provides visual insights into the complex and sophisticated systems that control cellular physiological functions. Solvatochromic fluorescent probes are highly promising noninvasive visualization tools for identifying the ordering of the microenvironment of plasma membrane microdomains. However, conventional probes, although capable of structural analysis, lack the necessary long-term photostability required for live ima

Epithelial cells have characteristic membrane domains. Identification of membrane proteins playing an important role in these membrane domains has progressed and numerous studies have been performed on the functional analysis of these membrane proteins. On the other hand, the precise roles of membrane lipids in the organization of these membrane domains are largely unknown. Historically, the concept of lipid raft arose from the analysis of lipid composition of the apical membrane, and it can be

The plasma membrane is generally associated with underling actin cytoskeleton. When the plasma membrane detaches from actin filaments, it is expanded by the intracellular pressure and the spherical membrane protrusion which lacks underlying actin cortex, termed bleb, is formed. Bleb is widely used for migration across species; however, the molecular mechanism underlying membrane blebbing remains largely unknown. Our recent study revealed that 2 small GTPases, Rnd3 and RhoA, are important regulat

When the plasma membrane (PM) detaches from the underlying actin cortex, the PM expands according to intracellular pressure and a spherical membrane protrusion called a bleb is formed. This bleb retracts when the actin cortex is reassembled underneath the PM. Whereas this phenomenon seems simple at first glance, there are many interesting, unresolved cell biological questions in each process. For example, what is the membrane source to enlarge the surface area of the PM during rapid bleb expansi