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FEBS Letters 581 (2007) 2194–2201

Minireview Tunneling nanotubes: A new route for the exchange of components between animal cells

Hans-Hermann Gerdes*, Nickolay V. Bukoreshtliev, Joa˜o F.V. Barroso Department of Biomedicine, University of Bergen, Jonas Lies vei 91, Bergen 5009, Norway

Received 16 January 2007; revised 27 March 2007; accepted 28 March 2007

Available online 4 April 2007

Edited by Thomas So¨llner

Very recently, a new principle of -to-cell communication Abstract Recently, highly sensitive nanotubular structures mediating membrane continuity between mammalian cells have based on the formation of thin membrane channels between been discovered. With respect to their peculiar architecture, mammalian cells was discovered [4]. These tubes, referred to these membrane channels were termed tunneling nanotubes as tunneling nanotubes (TNTs), mediate membrane continuity (TNTs). TNTs could form de novo between animal cells leading between connected cells and permit the direct intercellular to the generation of complex cellular networks. They have been transfer of diverse components [5]. In contrast to gap junc- shown to facilitate the intercellular transfer of as well tions, TNTs were also found to accomplish long-range, direc- as, on a limited scale, of membrane components and cytoplasmic ted communication between dislodged cells. In the following, molecules. It has been proposed that TNTs represent a novel and we summarize the current knowledge of TNTs by focusing general biological principle of cell-to-cell communication and it on their structure and formation, compare them to plasmodes- becomes increasingly apparent that they fulfill important func- mata of plants, and speculate on the emerging implications of tions in the physiological processes of multicellular organisms. Ó 2007 Published by Elsevier B.V. on behalf of the Federation of this new route of intercellular exchange of information. European Biochemical Societies.

Keywords: ; Plasmodesmata; Membrane 2. Architecture and formation continuity; Cell-to-cell communication; Intercellular transfer; F- TNTs were first characterized between cultured rat pheo- chromocytoma PC12 cells (Fig. 1A) [4]. Here they comprise thin membranous channels mediating continuity between the plasma membranes of connected cells (Fig. 1B) [4]. The diame- 1. Introduction ter of these channels ranges between 50 and 200 nm and their length reaches up to several cell diameters [4]. Similar structures Cell-to-cell communication is a fundamental process for the have been described for other permanent cell lines as well as for development and maintenance of multicellular organisms. Di- primary cell cultures (Table 1). For human -derived verse mechanisms for the exchange of molecular information macrophages, murine macrophage J774 cells, Epstein Barr between cells have been documented. The most common Virus (EBV) transformed human line 721.221 or periph- mechanism depends on the secretion of molecules and their eral blood natural killer (NK) cells, an average length of 30 lm subsequent binding to specific receptors of target cells and is and in some cases above 140 lm has been reported [6]. In the involved in synaptic [1] as well as in para- and endocrine sig- case of macrophages, two distinct classes of nanotubes, i.e. thin naling. A more direct mechanism relies on the transfer of small (diameter < 0.7 lm) and thick (diameter P 0.7 lm) were ob- cytoplasmic components, e.g. calcium, between adjacent cells served [7]. Between DU 145 human prostate cancer cells also through proteinaceous channels, known as gap junctions [2]. thinner (diameter of 100–200 nm) and wider (diame- In addition, immune cells were shown to secrete membrane ter P 1 lm) intercellular bridges were seen ranging from a vesicles referred to as exosomes. These vesicles are contained few microns to at least 100 lm in length [8]. The diameter of within multivesicular bodies, which, upon fusion with the plas- TNTs detected between human endothelial progenitor cells ma membrane, are released into the extracellular space [3]. (EPCs) and neonatal rat cardiac myocytes ranged from 50 to 800 nm and their length from 5 to 120 lm [9]. Finally, TNTs be- tween THP-1 measured up to 100 lm in length [10]. *Corresponding author. Fax: +47 55586360. For PC12 cells, TNTs are stretched between interconnected E-mail address: [email protected] (H.-H. Gerdes). cells attached at their nearest distance [4,11]. Thus, in contrast Abbreviations: TNT, tunneling nanotube; PDs, plasmodesmata; EPCs, to other cellular protrusions, TNTs are not associated with the endothelial progenitor cells; NK, natural killer; ER, endoplasmic substratum, but hover freely in the cell culture medium reticulum; SEL, size exclusion limit; EBV, Epstein Barr Virus; EGFP, (Fig. 2A, bottom). This peculiar morphology of TNTs is less enhanced green fluorescent protein; f-EGFP, EGFP fused to the evident the flatter and the more irregular-shaped the connected 0 0 farnesylation signal of c-Ha-Ras; DiI, 1,1 -dioctadecyl-3,3,3 , cells are, as is the case for e.g. normal rat kidney (NRK) cells 30-tetramethylindocarbocyanine perchlorate; DiO, 3,30-dioctadecylox- acarbocyanine perchlorate; DiD, 1,10-dioctadecyl-3,3,30,30-tetramethy- (S. Gurke, A. Rustom, H.H. Gerdes, unpublished data) or l-indodicarbocyanine perchlorate macrophages [7]. In retrospect, we believe that the apparent

0014-5793/$32.00 Ó 2007 Published by Elsevier B.V. on behalf of the Federation of European Biochemical Societies. doi:10.1016/j.febslet.2007.03.071 H.-H. Gerdes et al. / FEBS Letters 581 (2007) 2194–2201 2195

the target cell [4]. This may point to a chemo-tactical guidance of the protrusions or the involvement of other signals. In sup- port of this view, a directed outgrowth of long and thin cellular protrusions referred to as towards a basic fibroblast growth factor (bFGF) gradient has been observed in in vitro cultures of Drosophila cells [14]. The velocity of TNT elonga- tion between dislodging immune cells was determined to be approximately 0.2 lm per second [6]. This speed may also be valid for the de novo formation of TNTs from outgrowing fil- opodia, being consistent with the velocity range of processes driven by actin-polymerization [15], which is the most likely mechanism underlying TNT directed outgrowth. Interestingly, H2O2 was recently found to promote both actin polymeriza- tion and the formation of TNTs in primary cultures of rat [13]. Furthermore, the parallel observation that the phosphorylation of the p38 mitogen-activated protein ki- nase (MAPK) was also triggered by H2O2, may suggest an involvement of this pathway in the regulation of TNT forma- tion [13]. Finally, the importance of F-actin for TNT forma- tion and stabilization is further corroborated by their absence in PC12 cell cultures upon treatment with the actin- depolymerising drug latrunculin B [4]. After a filopodia-like structure encounters the target cell, both membranes fuse to form a stretched membrane connec- tion (Fig. 2A, bottom). In the case of PC12 cells, this process was accompanied by the degeneration of the remaining protru- sions, possibly reflecting the existence of a feedback mechanism [4]. The complete process of TNT formation between PC12 cells is accomplished on a time-scale of several minutes [4]. Interestingly, TNT formation is not only a single event re- stricted to pairs of cells, but frequently leads to the formation of local networks between groups of cells [4,10]. Between Fig. 1. Ultrastructure of tunneling nanotubes (TNTs). (A) Scanning PC12 cells, network formation starts directly after plating of electron microscopic (SEM) or (B) transmission electron microscopic individual cells and culminates after 2 h [4]. However, this time (TEM) image of a TNT connecting two cultured PC12 cells. The boxed areas in (A) and (B) are shown as higher magnification images (insets schedule presumably differs with respect to the growth condi- A1–A3, B1, B2). (B1) and (B2) represent consecutive 80 nm sections, tions and/or cell lines which are under investigation. This is which demonstrate membrane continuity between the connected cells. exemplified for cardiac myocytes and EPCs, where TNT struc- (C) Bars: 10 lm (A); 2 lm (B); 200 nm (insets A1–A3, B1, B2). tures first became visible after a few hours, and whose forma- Modified from Rustom et al., Science 303, 1007. tion culminated after 24 h and declined after 48 h [9]. Because TNTs were frequently found between diverging cells, it is pos- spheroidal shape of PC12 cells significantly contributed to the sible that they also exist between associated cells. This may indi- discovery of TNTs by emphasizing their unique characteristics cate that a preexisting membrane channel between cells which among the great variety of different cellular protrusions. It is of are in close contact is elongated as they start to diverge from note that these characteristics share striking similarities with one another (Fig. 2B). Such a force-driven process may resem- artificial nanotubes generated between giant uni- ble the generation of long membrane tethers in vitro [16–18]. lamellar liposomes mediating also membrane continuity [12]. TNTs between PC12 cells show a pronounced sensitivity to Whereas in most cases TNTs appeared as straight lines, the mechanical stress, chemical fixation and prolonged light expo- occurrence of branched connections could only be observed sure during widefield microscopy [4]. The latter leads to visible in rare cases [4,7,9]. vibrations and subsequent rupture of the thin membrane tubes Another structural property of TNTs is the prominent F-ac- [4]. Likewise, TNTs between other cell types, e.g. cardiac myo- tin bundle spanning the entire length of the nanotube cytes, EPCs and THP-1 monocytes, have been shown to display [4,7,10,13] (Fig. 2A, bottom). have not been a similar sensitivity [9,10]. This pronounced sensitivity may be found in TNTs connecting PC12 cells [4]. In the case of human one reason why theses structures were not recognized earlier monocyte-derived macrophages, thin TNTs contain only F-ac- and could hamper further structural and functional analyses. tin while the thick ones have both F-actin and microtubules On the other hand, this characteristic may provide a tool to [7]. The presence of microtubules was also observed in mem- interfere with the TNT-dependent cell communication route. brane channels between cultured prostate cancer cells [8]. TNTs between PC12 cells were seen to form de novo be- 3. Generic cargo tween dislodged cells [4]. Their de novo formation appears to be initiated by the outgrowth of filopodia-like cell protrusions First clues towards the function of TNTs between connected (Fig. 2A, top). Interestingly, in the case of PC12 cells, the cells were provided by videomicroscopic studies of neuroendo- growing filopodia-like structures seem to be directed towards crine PC12 cells. Differential interference contrast (DIC) 2196

Table 1 Cell cultures shown to form TNTs and their respective transferred cargo Cells shown to form TNTs Transferred cellular components/signalsa Cell-organelles Plasma membrane components Cytoplasmic components Permanent cell-lines Rat pheochromocytoma Endosome-related Lipid anchored proteins Cytoskeletal proteins (EGFP-actin); PC12 cells [4] (DiO, DiI, DiD, LysoTracker, (f-EGFP) motor proteinsb (myosin Va) synaptophysin-EGFP) Human embryonic kidney Endosome-related –– (HEK)-293 cells [4,59] (DiO, DiI, DiD) Normal rat kidney (NRK) cells [4]c Endosome-related – Cytoskeletal componentsb (F-actin) (DiO, DiI, DiD) Between PC12 and HEK-293 cellsc Endosome-related –– (DiO, DiI) Between HEK-293 and NRK cellsc Endosome-related –– (DiO, DiI) Between PC12 and NRK cellsc Endosome-related –– (DiO, DiI) EBV-transformed human – Glycosylphosphatidylinositol – B cell line (721.221) [6] (GPI)-GFP Murine macrophage J774 cells [6] –– – DU 145 human prostate Endosome-related –– cancer cells [8] (DiO, DiI) THP-1 monocytes [10] – Surface receptorsb (HLA-A,B,C Calcium fluxes; dyes (Lucifer yellow) class I MHC) Hepatic HepG2 cells [60] –– – TRVb-1 cells [60] –– – Bovine mammary gland epithelial –– – (BMGE) cells [60] Primary cultures Between 721.221 cells and human – Surface receptorsb –

peripheral blood NK cells [6] (HLA-Cw6-GFP) 2194–2201 (2007) 581 Letters FEBS / al. et Gerdes H.-H. Human monocyte-derived Endosome-relatedb (DiO) – Cytoskeletal components (microtubules)b macrophages [6,7] Between macrophages and NK cells [6] –– – Between neonatal rat Mitochondria (MitoTracker) – Soluble proteins (GFP) cardiomyocytes and adult human EPCs [9] Primary cultures of rat astrocytes [13] – – Cytoskeletal proteinsb (F-actin); motor proteinsb (myosin Va) Myeloid-lineage dendritic cells Surface receptorsb – Calcium fluxes prepared from peripheral (HLA-A,B,C class I MHC) blood monocytes [10] Between dendritic cells and THP-1 – – Calcium fluxes monocytes [10] Hematopoietic stem and progenitor –– – cells [58] Primary cultures of mouse medullac –– – aMarkers which were tested experimentally appear in brackets. bThese markers were shown inside TNTs, but their intercellular transfer was not proven. cS. Gurke, A. Rustom, H.-H. Gerdes, unpublished data. H.-H. Gerdes et al. / FEBS Letters 581 (2007) 2194–2201 2197

Fig. 2. Model of TNT formation. (A) One cell forms an actin-driven protrusion directed towards the target cell (top). Fusion of the cell protrusion with the membrane of the target cell results in TNT formation (bottom). (B) TNTs may form between adjacent cells, which subsequently diverge. Red line, F-actin; arrows, direction of filopodium (A) and cell (B) movement, respectively. microscopy showed bulges moving uni-directionally along membrane tube, is also the donor cell for the delivery of organ- TNTs at a speed of approximately 25 nm/s [4]. This process elles, although the opposite scenario cannot be ruled out. In showed striking similarities to the demonstrated transfer of li- this respect, transfer along membrane nanotubes pid containers between liposomes connected via phospolipid could also be an explanation for the thus far unexplained bilayer nanotubes [12,19]. The assumption that the moving transfer of melanosomes from melanocytes to keratinocytes bulges of PC12 cells indeed represent membrane containers [20,21]. As a variation of this theme mitochondria and endo- was subsequently verified by transfer experiments employing some-related vesicles were both seen to move bidirectionally fluorescent markers for different cellular organelles. These within thick nanotubes connecting macrophages [7]. Notably, experiments, performed with two differentially labeled PC12 these nanotubes contained both F-actin and microtubules cell populations, showed that the moving containers represent and the organelle movement was shown to depend on the latter small organelles belonging to the endosomal/lysosomal system cytoskeletal structure [7]. The occurrence of an intercellular [4] (Table 1). This, together with the finding that F-actin is transfer of organelles raises the question as to what kind of present inside TNTs, suggests that the movement of organelles information is transferred and how this information is inte- along TNTs is facilitated by an acto-myosin-dependent mech- grated in the target cells. In the case of endosome-related anism (Fig. 3). Such a view is further corroborated by the pres- organelles, it is conceivable that any molecules packaged into ence of the actin-specific motor protein myosin Va in TNTs or associated with these structures can be transferred over long [4,13] (Table 1), partly colocalizing with the respective organ- distances between cells. Thus, it is notable that the early endo- elles [4]. Alternatively, the transfer of organelles could be dri- somal system in particular is one of the major reloading points ven by the polymerization of actin itself, which could be for a variety of signaling complexes functioning at the cell sur- regarded as a process of intercellular ‘‘treadmilling’’. Accord- face. Furthermore, transfer studies using PC12 cells revealed ing to this model, organelles are transferred by their associa- that after being transferred along TNTs, endosome-related tion with the constantly sliding F-actin rod. Irrespective of structures are able to fuse with their counterparts in the respec- the existent mechanism, the uni-directional transport observed tive target cells [4]. These facts provoke the intriguing model implies that the individual actin fibers inside the nanotubes that the transfer of organelles and their subsequent fusion with have the same polarity. membranes of the target cell conveys information between Apart from the transfer of small organelles, subsequent stud- connected cells. ies showed that TNTs also facilitate the intercellular transfer Initial studies of PC12 cells showed that in addition to of larger organelles such as mitochondria from cardiac myo- organelles, plasma membrane components such as lipids or li- cytes to EPCs [9]. Like the uni-directional transport of endo- pid-anchored proteins are also transported along TNTs to the some-related structures between PC12 cells, this transport connected target cells [4] (Fig. 3). This lateral transfer in the has been reported to occur in one direction only [9]. It is tempt- plane of the cell surface is consistent with membrane continu- ing to speculate that the cell, which initiates the formation of a ity between the connected cells. In the case of PC12 cells, mem- brane transfer is quite slow as has been shown by the intercellular transport of EGFP fused to the farnesylation sig- nal of c-Ha-Ras (f-EGFP) (Table 1), a lipid-anchored mem- brane marker [4]. The slow transfer of membrane components from thin tubular projections to spherical cell sur- faces has also been predicted by mathematical models [22]. Nevertheless, in the case of cell signaling molecules, the lateral transfer of a few molecules can fulfill important roles in inter- cellular communication. The existence of membrane continuity mediated by mem- brane channels should result in the occurrence of a cytoplasmic Fig. 3. Schematic representation of a TNT indicating selective transfer bridge that, in principle, facilitates the transfer of cytoplasmic of different cellular components as indicated. material between interconnected cells (Fig. 3). For this reason, 2198 H.-H. Gerdes et al. / FEBS Letters 581 (2007) 2194–2201 it was surprising to find that TNTs between PC12 cells seem to thickness of the two connected cells. This is contrasted by impede the intercellular flow of even small molecules such as TNTs, which show a significant variation in length and con- the 400 Da dye molecule calcein [4]. This suggests that the pas- nect even distant cells through the extracellular space. Despite sive diffusion of small cytoplasmic molecules through TNTs is their difference in length, the diameter of PDs is 50 nm significantly slowed. However, the transfer of actin as the [26,27,29–31] and thus is comparable to that of TNTs. structural backbone of TNTs could be detected [4], a finding PDs form by two distinct mechanisms, either as a conse- which is consistent with the proposed intercellular ‘‘treadmil- quence of incomplete cell plate/wall formation during cytoki- ling’’ process. It is possible that the F-actin rod tightly nesis, resulting in primary PDs, or de novo across the cell wrapped by the surrounding membrane functions as a plug wall, resulting in secondary PDs [32]. By the latter mechanism for passive diffusion. However, Lucifer yellow was transferred cells are able to increase their intercellular traffic and connect TNT-dependent between THP-1 cells [10] and GFP seems to with others not cytokinetically related to them. To date, TNTs be transferred between TNT-connected cardiac myocytes and have only been found to form de novo between mammalian EPCs [9] (Table 1). Furthermore, calcium fluxes through rela- cells [5]. However, this does not exclude that during certain tively large diameter TNTs were observed between dendritic developmental stages they may also form during cytokinesis cells and THP-1 monocytes [10] (Table 1). This may indicate as has been documented for the formation of ring canals that TNTs possess different permeabilities for soluble mole- [33,34] and smaller intercellular bridges connecting Drosophila cules, which in turn may reflect the existence of a gating mech- germ line cells [35] as well as broad intercellular bridges be- anism or structural differences between the respective tween spermatids of mammals [36]. intercellular structures of different cell types. In agreement Although both PDs and TNTs facilitate membrane continu- with this view, two different morphological classes of TNTs ity between connected cells, thus suggesting a strong functional have been described [7,8]. Further support for this concept is relationship, the ultrastructure of PDs from higher plants ap- also provided by the observed differences in permeability for pears to be distinct from that of mammalian TNTs [4,37].A structurally related membrane nanotubes in plants know as characteristic feature of PDs from higher plants is the presence plasmodesmata, e.g. during different stages of plant develop- of an appressed endoplasmic reticulum (ER) concentric to the ment [23,24] (see below). limiting plasma membrane sleeve that runs the length of the PD and links the ER of the two connected cells. This structure is also known as the desmotubule [4,37] (Fig. 4). Electron 4. Relation between TNTs and plasmodesmata microscopic studies clearly show that TNTs lack such a struc- ture [4,37]. Interestingly, PDs of the characean algae Chara Eduard Tangl first described in 1879 channels between plant corallina also lack an appressed ER and may represent a prim- cells providing ‘‘offene Kommunikationen’’. These channels itive precursor of higher plant PDs [37]. The major transport were later (1901) named ‘‘Plasmodesmen’’ by Eduard Stras- route of molecules through higher plant PDs occurs within burger [25]. In 1968 Roberts finally described plasmodesmata the region between the desmotubule and the plasma membrane (PDs; singular ) at the electron microscopic level known as the cytoplasmic sleeve [32] (Fig. 4). This space is as a structure with membraneous outlines [26,27]. These small filled with proteinaceous material suggested to form a helical membrane conduits have been shown to bridge neighbouring channel, which determines the size exclusion limit (SEL) of cells [28] and thus their length is determined by the cell wall the molecules free to flow between connected cells (Fig. 4).

Fig. 4. Schematic comparison of TNTs and PDs. Structural components (top table) and generic transferred cargo (bottom table) are compared between the two membranous channels. For both tables the following symbols indicate: (+) present, () absent, (?) unknown. H.-H. Gerdes et al. / FEBS Letters 581 (2007) 2194–2201 2199

The SEL of PDs appears to be tissue- and possibly cell-type and soluble molecules between dislodged cells (Fig. 3) indicates specific, and is developmentally regulated [32,37]. unique physiological functions fulfilled by these nanotube con- PDs allow the transfer of nutrients and signals necessary for nections. In this respect, two recent studies present evidence plant growth and development [32]. The transport of such mol- for a TNT-dependent communication between various cells ecules via PDs is thought to occur by two independent mech- of the immune system, which suggests a crucial role in immune anisms known as non-targeted and targeted movement [37].In defence [6,10]. In one case, it was shown that dendritic cells are the first case small tracer molecules smaller than the SEL of the able to propagate calcium fluxes through TNTs upon stimula- channel move by passive diffusion from cell to cell. This im- tion, leading to the activation of a large group of intercon- plies that hormones, minerals, amino acids and sugars may nected immune cells [10]. This TNT-mediated transfer of move freely between cells. In the second case, molecules are calcium ions led to the flattening and membrane extension of transported by a selective and regulated mechanism, if they the receiving cells as a physiological response in preparation possess an intrinsic trafficking signal(s). Such signal(s) leads for phagocytosis [10]. Furthermore, in the second study, class to a dilation of the plasmodesmal pore allowing movements I major histocompatibility complex (MHC) was detected in of macromolecules larger than its basal SEL [32]. Such mole- TNTs between EBV transformed human B cell line 721.221 cules include viral proteins, viral genomes, RNA or ribonu- transfected with HLA-Cw6-GFP and human peripheral blood cleoparticles (RNPs) and transcription factors including NK cells [6]. This may point to a key role of TNTs in a faster homeodomain proteins [37–39] (Fig. 4). and more efficient presentation of antigens, in particular at the Over the last century several constituents of PDs have been immunological synapse [44]. identified. These include the conserved calcium-sequestering If TNTs indeed play key roles in cell-to-cell signaling, it can protein calrecticulin [40], the contractile protein centrin [41], be expected that alterations in their structure and function may the unconventional myosin VIII [42] and F-actin [43]. Microtu- lead to severe pathological defects in the respective organisms. bules, however, appear to be absent in PDs [28] as well as in This may be relevant for, e.g., the development and progres- TNTs characterized from PC12 cells [4]. In conjunction with sion of cancer as one of the most severe disorders of modern ultrastructural studies, showing constrictions at the neck re- times. For the past decade evidence has accumulated that can- gions of PD channels, centrin is thought to be involved in cer derives from an arrest of stem cell maturation rather than the regulation of transport of molecules through the cytoplas- from the dedifferentiation of somatic cells [45]. How this arrest mic sleeve [41]. It has been hypothesized that such constriction is evoked is still a matter of debate, but several intriguing sug- can result from the contraction of centrin nanofilaments, gestions have been proposed. One idea is that normal stem which are located at the neck region of PDs (Fig. 4), regulated cells fuse with somatic cells leading to genetic reprogramming presumably by calcium in concert with calreticulin [32,37].A and the generation of cancer stem cells [46]. The generation of second mechanism for the regulation of pore closure may be such a hybrid cell may involve a transient fusion event between provided by callose deposition at the extracellular space facing the respective cell types mediated by TNTs, enough to allow the neck regions of PDs, leading to the formation of a contrac- the transfer of components necessary to induce cancer stem cell tile sphincter [32] (Fig. 4). Conversely, degradation of callose formation. Furthermore, several lines of evidence suggest that would lead to dilation of the plasmodesmal pore to promote inappropriate cell-cell fusion between tumor and somatic cells macromolecular traffic [37]. Furthermore, regulation of plas- may contribute to cancer progression by generating hybrids modesmal transport may also occur along their entire length with more aggressive properties including the ability to metas- by an acto-myosin-dependent mechanism of contraction/relax- tasize and resistance to chemotherapy [46]. Again, TNTs may ation between the desmotubule and the plasma membrane [32]. play a key role in this process. A higher resistance to cytostatic Accordingly, F-actin spirals around the desmotubule and con- drugs could reflect an increased level of ABC transporters nects it to the plasma membrane via myosin VIII (Fig. 4). (drug-efflux pumps) in the tumor cells [45]. In this respect, it Again, the contraction/relaxation activity of this complex is has been reported that the P-glycoprotein, an ABC transporter thought to be regulated by calcium and calreticulin [32].In that mediates multidrug resistance, is transferred between dif- agreement with the observation that both TNTs and PDs rep- ferent tumor cells in vitro and in vivo [47]. Although the under- resent membrane conduits between animal and plant cells, lying mechanism for this intercellular transfer is still unknown respectively, it is conceivable that they share many functions [47], the data presented is consistent with a TNT-dependent and properties. In this respect, it would not be surprising, if fu- transfer mechanism of plasma membrane components [48] ture research shows that, e.g. the acto-myosin complex in (Fig. 3). TNTs together with so far unidentified contractile elements The striking similarities between TNTs and PDs (Fig. 4) are involved in a gating mechanism as in PDs or, conversely, strongly suggest that both may play similar roles in the phys- F-actin in conjunction with myosin facilitates the movement iology of the respective organisms. Thus, in light of the finding of cargo along PDs as proposed for TNTs (Fig. 4). In conclu- that PDs play fundamental roles in pattern-formation during sion, the knowledge of the functions of PDs may be instructive plant development, e.g. by RNA transfer [24,39,49], it is tempt- for understanding the role of TNTs and vice versa. ing to speculate that TNTs also orchestrate essential parts of cell differentiation in animal organisms, as has recently been implied for the differentiation of EPCs [9,50] and developmen- 5. Emerging physiological implications tal processes in general [51]. With respect to the latter, TNTs may open an alternative route for, e.g., the selective distribu- The discovery of TNTs has revealed a novel type of intercel- tion of morphogens during embryogenesis [52]. Intercellular lular communication between animal cells. In particular, the transport of morphogens through tubular connections via ability to establish selective long-range transfer of various cel- membrane containers, in addition to passive diffusion through lular components such as organelles, membrane constitutents the extracellular space, seems plausible because morphogens 2200 H.-H. Gerdes et al. / FEBS Letters 581 (2007) 2194–2201 such as Sonic hedgehog are often closely membrane-associated nanotubes between human macrophages support long-distance [53–55]. Similarly, also RNA, which was shown to exert exten- vesicular traffic or surfing of bacteria. J. Immunol. 177, sive cell–cell movement via PDs [56], may distribute through 8476–8483. [8] Vidulescu, C., Clejan, S. and O’Connor, K.C. (2004) Vesicle TNTs between animal cells resulting in regulation of gene traffic through intercellular bridges in DU 145 human prostate expression by means of post-transcriptional silencing (Fig. 4). cancer cells. J. Cell. Mol. Med. 8, 388–396. 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