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Submembraneous Microtubule Cytoskeleton: Biochemical and Functional Interplay of TRP Channels with the Cytoskeleton Chandan Goswami and Tim Hucho

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Submembraneous Microtubule Cytoskeleton: Biochemical and Functional Interplay of TRP Channels with the Cytoskeleton Chandan Goswami and Tim Hucho MINIREVIEW Submembraneous microtubule cytoskeleton: biochemical and functional interplay of TRP channels with the cytoskeleton Chandan Goswami and Tim Hucho Department for Molecular Human Genetics, Max Planck Institute for Molecular Genetics, Berlin, Germany Keywords Much work has focused on the electrophysiological properties of transient actin; axonal guidence; cytoskeleton; growth receptor potential channels. Recently, a novel aspect of importance cone; myosin; pain; signalling complex; emerged: the interplay of transient receptor potential channels with the transient receptor potential channels; cytoskeleton. Recent data suggest a direct interaction and functional reper- tubulin; varicosity cussion for both binding partners. The bi-directionality of physical and Correspondence functional interaction renders therefore, the cytoskeleton a potent integra- C. Goswami, Department for Molecular tion point of complex biological signalling events, from both the cytoplasm Human Genetics, Max Planck Institute for and the extracellular space. In this minireview, we focus mostly on the Molecular Genetics, Ihnestrasse 73, 14195 interaction of the cytoskeleton with transient receptor potential vanilloid Berlin, Germany channels. Thereby, we point out the functional importance of cytoskeleton Fax: +49 30 8413 1383 components both as modulator and as modulated downstream effector. Tel: +49 30 8413 1243 E-mail: [email protected] The resulting implications for patho-biological situations are discussed. (Received 15 April 2008, revised 23 June 2008, accepted 30 July 2008) doi:10.1111/j.1742-4658.2008.06617.x The microtubule cytoskeleton plays a role in a variety with various membrane proteins, which often are part of cellular aspects such as division, morphology and of large protein complexes. The dynamic properties motility, as well as the transport of molecules and and the complexity of tubulin as an interacting protein organelles toward and from the cell membrane. in large complexes at the membrane just are beginning Although all these phenomena affect the plasma mem- to be unravelled. One apparent function is to serve as brane, however, most of the microtubule filaments do a scaffold protein and modulator of transmembrane not reach to the lipid membrane region, partially due signalling. to a thick hindering cortical actin network. However, recent studies indicate that a small number of dynamic Cytoskeletal components in signalling microtubules can extend rapidly to the cell membrane. complexes at membranes Although most contacts are established only tran- siently, there are membranous regions in which the The cytoplasmic domains of transient receptor pot- plus end of these pioneering microtubules is stabilized. ential (TRP) channels recruit large complexes of Stabilization appears to be mediated by the interaction proteins, lipids and small molecules. Depending on the Abbreviations FHIT, fragile histidine triad protein; MAP, microtubule-associated protein; RTX, resinferatoxin; TRP, transient receptor potential; TRPV, transient receptor potential vanilloid; TRPV1, transient receptor potential vanilloid subtype 1; TRPV1-Ct, C-terminal of transient receptor potential vanilloid subtype 1; TRPV1-Nt, N-terminal of transient receptor potential vanilloid subtype 1; TRPV2, transient receptor potential vanilloid subtype 2; TRPV4, transient receptor potential vanilloid subtype 4. 4684 FEBS Journal 275 (2008) 4684–4699 ª 2008 The Authors Journal compilation ª 2008 FEBS C. Goswami and T. Hucho TRP channels and cytoskeleton regulate each other preparation method, these structures have been domains like ankyrin repeats, Ca2+-sensing EF hands, referred to as ‘signalplex’, i.e. complexes involved in phosphorylation sites, calmodulin-binding sites and a signalling events [1], or ‘channelosome’, i.e. complexes so-called ‘TRP box’. Based on their sequence, the formed around functional ion channels [2], and ⁄ or as mammalian TRP family is differentiated into six ‘lipid raft complexes’, i.e. complexes localized to this subfamilies, namely TRP canonical (TRPC), TRP membranous subdomain [3]. Proteomic studies of vanilloid (TRPV), TRP melastatin (TRPM), TRP ‘signalplexes’ or ‘channelosomes’ purified from cell polycystin (TRPP), TRP mucolipin (TRPML) and lines, as well as from brain, give both direct and indi- TRP ankayrin (TRPA) ion channels [18]. All TRP rect evidence for the presence of the cytoskeleton as channels investigated to date are involved in the detec- well as ion channels. Scaffolding adaptors like inacti- tion and ⁄ or transduction of physical and chemical vation-no-afterpotential D [4–6], Na+ ⁄ H+ exchanger stimuli. regulatory factor [7] and ezrin ⁄ moesin ⁄ radixin-binding phosphoprotein 50 [8] are also found, some of which TRPV1 and the cytoskeleton interact directly with ion channels, e.g. TRP channels [9], but also contain binding motifs for cytoskeletal Physical interaction of TRPV1 with the proteins [8,10]. Accordingly, cytoskeletal proteins such cytoskeleton as spectrin, myosin, drebrin and neurabin, as well as tubulin and actin [1,2,6,11] are confirmed components TRPV1 is the founding member of the vanilloid sub- in signalplexes and channelosomes. family of TRP channels and detects several endo- Complementary proteomic studies of purified lipid genous agonists (e.g. N-arachidonoyl-dopamine) and rafts reveal the presence of several cytoskeletal proteins noxious exogenous stimuli, such as capsaicin (the main [12,13] such as a- and b-tubulin, tubulin-specific chaper- pungent ingredient of hot chilly) and high temperature one A (a folding protein involved in tubulin dimer (> 42 °C) [19,20]. TRPV1 is a nonselective cation assembly), KIF13 (a kinesin) [12], actin, nonconven- channel with high permeability for Ca2+. In recent tional myosin II and nonconventional myosin V [14]. years, TRPV1 has gained extensive attention for its Similarly, proteomics studies of the ‘membrane cyto- involvement in signalling events in the context of pain skeleton’, a submembranous fraction, which is attached and other pathophysiological conditions including to the cytoskeleton [15], and of cytoskeleton-associated cancer [21–27]. proteins in general [16], indicate the presence of lipid The interaction of TRPV1 with tubulin was first iden- raft membrane proteins as well as cytoskeletal proteins. tified through a proteomic analysis of endogenous inter- Together, these varying studies give strong evidence actors enriched from neuronal tissue [28]. The that cytoskeletal proteins are part of signalling com- interaction was then confirmed by biochemical plexes including transmembrane proteins and are approaches including co-immunoprecipitation, micro- involved in their organization at membrane. tubule co-sedimentation, pull-down and cross-linking experiments. In contrast to the tubulin cytoskeleton, the physical interaction of TRPV1 with actin or neurofila- Structural features of TRP channels ment cytoskeleton has not been observed to date [28,29]. The TRP family of ion channels is named after the The C-terminus of TRPV1 (TRPV1-Ct) is sufficient Drosophila melanogaster trp mutant, which is charac- for the interaction with tubulin while the N-terminus terized by a transient receptor potential in the photore- of TRPV1 (TRPV1-Nt) apparently does not interact ceptors in response to light [17]. In the meantime, [28]. Using deletion constructs and biotinylated pep- orthologues and paralogues of TRP channels have tides, the tubulin-binding region located within been described in organisms ranging from simple TRPV1-Ct was mapped to two short, highly basic eukaryotes to human. They share a high degree of regions (amino acids 710–730 and 770–797) [29]. If an homology in their amino acid sequence. TRP channels a-helical conformation is assumed, these two regions are formed by monomers with six transmembrane project all their basic amino acids to one side, thus regions that assemble into tetramers, which form the potentially enabling interactions with negatively functional cation-permeable pore. The most conserved charged residues (Fig. 1). Indeed, correspondingly, the region is the sixth transmembrane domain, which con- C-terminal over-hanging region of tubulin contains a stitutes most of the inner lining of the ion channel large number of negatively charged glutamate (E) resi- pore. The N- and C-termini of TRP channels are dues in a stretch characterized as unstructured region located in the cytoplasm and, depending on the respec- of the tubulin and referred as E-hook. These E-hooks tive TRP channel, consist of various functional are known to be essential for the interaction of tubulin FEBS Journal 275 (2008) 4684–4699 ª 2008 The Authors Journal compilation ª 2008 FEBS 4685 TRP channels and cytoskeleton regulate each other C. Goswami and T. Hucho A with various microtubule-associated proteins such as MAPs, Tau, as well as others. Indeed, binding of TRPV1-Ct with tubulin was abolished when the E-hooks containing over-hangs were removed by prote- B ase treatment [29]. The tubulin-binding region of TRPV1 apparently is under high evolutionary pressure as its sequence is highly conserved in all TRPV1 ortho- logues [29]. Also between homologues, the distribution of basic amino acids composing the tubulin-binding regions is conserved even though the overall amino acid conservation is rather limited. Based on these data an interaction of tubulin with TRPV2, TRPV3 and TRPV4 (Fig. 2) can also be predicted. These TRPV1 homo- logues have the highest conservation of basic charge distribution
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