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Regulation of Actin-Based Apical Structures on Epithelial Cells Thaher Pelaseyed1,* and Anthony Bretscher2

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Regulation of Actin-Based Apical Structures on Epithelial Cells Thaher Pelaseyed1,* and Anthony Bretscher2 © 2018. Published by The Company of Biologists Ltd | Journal of Cell Science (2018) 131, jcs221853. doi:10.1242/jcs.221853 REVIEW SUBJECT COLLECTION: CILIA AND FLAGELLA Regulation of actin-based apical structures on epithelial cells Thaher Pelaseyed1,* and Anthony Bretscher2 ABSTRACT related finger-like protrusions that are found on the apical membrane Cells of transporting epithelia are characterized by the presence of of hair cells. We highlight the contribution of bundled actin abundant F-actin-based microvilli on their apical surfaces. Likewise, filaments to apical organization, and the contribution of tip-linking auditory hair cells have highly reproducible rows of apical stereocilia to the structure and function of microvilli and stereocilia. (giant microvilli) that convert mechanical sound into an electrical Furthermore, we describe an emerging mechanism of how signal. Analysis of mutations in deaf patients has highlighted the bundled actin filaments are connected to the plasma membrane in critical components of tip links between stereocilia, and related a way that restricts their presence to the apical membrane. structures that contribute to the organization of microvilli on epithelial cells have been found. Ezrin/radixin/moesin (ERM) proteins, which Morphological features and arrangement of actin-based are activated by phosphorylation, provide a critical link between the surface structures plasma membrane and underlying actin cytoskeleton in surface Shaping the plasma membrane into a finger-like structure requires structures. Here, we outline recent insights into how microvilli and a core of bundled cytoskeletal filaments with lateral attachments stereocilia are built, and the roles of tip links. Furthermore, we to the plasma membrane. In the case of cilia, the filaments are highlight how ezrin is locally regulated by phosphorylation, and that microtubules, whereas filaments of actin support filopodia, this is necessary to maintain polarity. Localized phosphorylation is microvilli and stereocilia (DeRosier and Tilney, 2000; Lewis achieved through an intricate coincidence detection mechanism that and Bridgman, 1992). Here, we restrict our discussion to requires the membrane lipid phosphatidylinositol 4,5-bisphosphate microvilli and stereocilia, which are found on the apical domain of epithelial cells. [PI(4,5)P2] and the apically localized ezrin kinase, lymphocyte- oriented kinase (LOK, also known as STK10) or Ste20-like kinase The plasma membrane of microvilli is supported by a bundle of ∼ – (SLK). We also discuss how ezrin-binding scaffolding proteins 30 40 parallel, inherently polarized actin filaments with their regulate microvilli and how, despite these significant advances, it barbed ends at the microvillus tip (Mooseker and Tilney, 1975). remains to be discovered how the cell polarity program ultimately Early morphological studies showed that the F-actin core is bundled interfaces with these processes. and attached to the plasma membrane by lateral cross-bridges of ∼30 nm length, indicating that the actin filaments are both KEY WORDS: Actin, Cell polarity, Cytoskeleton, ERM proteins, crosslinked to one another and to the plasma membrane (Brunser Microvilli, Stereocilia and Luft, 1970; Matsudaira and Burgess, 1979; Mooseker and Tilney, 1975). On intestinal epithelia and the epithelium of the Introduction kidney proximal tubule, the microvilli are of uniform length and Cells throughout the vertebrate body are polarized. By regulating its generally hexagonally packed. Despite over 40 years of research, it internal organization and plasma membrane composition in a is still not clear what drives the assembly of actin filaments in polarized manner, the cell ensures organism viability and the microvilli, but much has been learned about their morphogenesis execution of central biological functions, such as cell division, by studying microvillar components. Today, we appreciate that proliferation, signaling and migration. One critical aspect of cell events such as cytoskeletal remodeling, membrane–cytoskeleton polarity is how cells build and restrict morphological features to one crosslinking and extracellular adhesion are parts of the cellular domain of a polarized cell. The epithelial cell serves as the program that shapes the apical brush border domain (Crawley et al., archetypal example of a polarized cell. Its apical membrane domain 2014b). We discuss some of these cellular programs in this Review. is decorated with finger-like protrusions called microvilli (Granger Stereocilia are found on the apical aspect of hair cells of the and Baker, 1950). Microvilli expand the interface between the cell vertebrate cochlea and vestibular systems. Early studies of bird and extracellular milieu, and accommodate membrane proteins that cochlea revealed that the length of hair cell stereocilia range from take part in signaling as well as ion and nutrient transport. By 1.5 to 5 µm, increasing in length from the proximal to distal end contrast, the basolateral membrane harbors a different composition (Tilney and Saunders, 1983). However, in contrast to what is seen in of membrane proteins and displays a membrane morphology that is birds, stereocilia on individual cells of the mammalian cochlea very distinct from the apical brush border membrane (Bryant and exhibit a staircase-like pattern with a predetermined graded increase Mostov, 2008). In this Review, we discuss recent insights into the in stereociliary length between rows (Fig. 1A, see Box 1). The regulation and assembly of the apical microvilli and stereocilia – mammalian cochlea contains two types of hair cells: a single row of inner hair cells with a linear array of stereocilia measuring ∼5µmin length, and three rows of outer hair cells with stereocilia arranged in 1Institute of Biomedicine, Department of Medical Biochemistry and Cell Biology, a V-shaped pattern and measuring 2–3 µm in length (Dallos, 1996; University of Gothenburg, SE-405 30 Gothenburg, Sweden. 2Weill Institute for Cell and Molecular Biology, Department of Molecular Biology and Genetics, Kaltenbach et al., 1994). Vestibular hair bundles in the utricle of Cornell University, Ithaca, NY 14853, USA. adult mouse range from ∼2 µm for the shortest stereocilia in the staircase to up to ∼15 µm (Li et al., 2008; Rzadzinska et al., 2004). *Author for correspondence ([email protected]) Each mature stereocilium contains 400–3000 actin filaments with T.P., 0000-0002-6434-3913; A.B., 0000-0002-1122-8970 their barbed ends at the tip and pointed ends associated with the Journal of Cell Science 1 REVIEW Journal of Cell Science (2018) 131, jcs221853. doi:10.1242/jcs.221853 A Fig. 1. Organization and regulation of stereocilia and microvilli. Stereocilia (A) Stereocilia of auditory hair cells are arranged in a staircase configuration. Tip link Stimulus Harmonin Actin bundles are assembled and stabilized through actin-binding proteins such as fimbrin, espin and fascin. SANS Stereocilia of each row are connected to stereocilia of adjacent rows through tip CDH23 links. Tip links require a ternary complex of Myo7a, USH1C (harmonin) and SANS K+ that binds to the cadherin CDH23. Ca2+ MET channel CDH23 forms a heteromeric interstereociliary link with protocadherin Myo7a PCDH15 PCDH15 on the adjacent stereocilium. The cytoplasmic region of PCDH15 interacts with the mechano-electrical transduction (MET) channels. Auditory stimulus causes deflection towards taller rows, which generates an influx of K+ and F-actin F-actin Ca2+ ions through the MET channels. (B) Key Microvilli covering the apical aspects of Fimbrin Espin Fascin intestinal epithelial cells require heterotypic extracellular linkages that are mediated by the protocadherin PCDH24 B and mucin-like protocadherin MLPCDH. By analogy with stereocilia, the Microvilli cytoplasmic tail of PCDH24 binds the ternary complex of USH1C (harmonin), Harmonin the SANS homolog ANKS4B and Myo7b. The actin cytoskeleton is ANKS4B assembled and stabilized by villin, espin PCDH24 and fimbrin. MLPCDH Myo7b F-actin F-actin Key Villin Espin Fimbrin base, where it tapers and inserts into the cuticular plate of the hair Apical membrane morphology: the cell polarity program and cell (Tilney et al., 1980, 1989; Zheng et al., 2000). In an extended membrane traffic series of studies, Tilney et al. have documented the assembly Microvilli and stereocilia are restricted to the apical domain of of stereocilia during development in ultrastructural detail their respective cells (Fig. 1A,B). This apical-basolateral polarity (Tilney et al., 1992). Stereocilia on mature hair cells are linked is directed by an epithelial polarity program that involves intrinsic by tip links that activate mechano-electrical transduction channels polarity proteins as well as signals from adjacent cells and the when the stereocilia are bent at their base in response to sound. extracellular matrix (Rodriguez-Boulan and Macara, 2014). This A prerequisite for proper mechano-electrical transduction is a program was elucidated mostly from studies in C. elegans and directional mechanosensitivity based on hair cells being arranged Drosophila (Campanale et al., 2017) and, in the fly, it consists of so that their axis of sensitivity is aligned with the direction of Bazooka, Partitioning defective-6 and Atypical protein kinase C stimulation (Shotwell et al., 1981). Recently, a series of studies has (Baz–Par-6–aPKC), which function antagonistically through revealed the molecular mechanisms that instruct the organization of aPKC
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