Classical morphological descriptions of tissue structures do not consider how shape determines function. Nonetheless, the inti‑mate interrelation between these features is often evident at the level of individual cells. For example, the slime mould Dictyostelium discoideum becomes asymmetrically polar‑ized in response to the chemoattractant cyclic AMP (cAMP) and forms two discrete poles: the leading edge (which is enriched in β‑actin) and the uropod at the trailing edge1. The uropod can be defined as a plasma membrane protrusion in which specific organelles, cytoskeletal proteins and adhesion and signalling receptors are con‑centrated (FIG. 1; TABLE 1). As D. discoideum cells migrate along a chemotactic gradient of cAMP, they align themselves, forming streams of cells that migrate together in a head‑to‑tail manner to form multicellular aggregates in a process called streaming. This process requires the polarization of adenylate cyclase at the uropod, generating a trail of cAMP for other cells to follow2.

Uropods are not exclusive to D. discoi-deum. Other cells that display prominent uropods during migration along chemotac‑tic gradients include Entamoeba histolytica, zebrafish immunocytes3 and vertebrate leukocytes, including T and B lymphocytes, natural killer (NK) cells, monocytes, granulo‑cytes and dendritic cells4,5. Irrespective of the cell type, uropods protrude at the rear of motile cells, lifting away from the

substratum, they contain a microtubular meshwork and accumulate N‑linked glyco‑sylated receptors with high affinity for the lectins concanavalin A and phytohaemag‑glutinin. The leukocyte uropod was first identified during studies of the interactions between T lymphoblasts and macrophages by McFarland and colleagues6, who coined the term uropod to describe a “cell append‑age which was both a tail and a connecting stalk” and noted that uropod formation is the most prominent morphological feature of motile leukocytes.

Leukocytes must migrate along chemo‑tactic gradients to exit the bloodstream and reach target tissues. Migration requires a polarized morphology to enable leukocytes to convert mechanical force into net forward locomotion. Leukocyte polarization is gener‑ated by a coordinately regulated bi directional actin cycle. Actin polymerization at the leading edge pushes the cell front forward, whereas filamentous (F)‑actin near the plasma membrane moves rearwards. At the front of the cell, a dynamic process of membrane recycling maintains the effective concentrations of chemotactic receptors, ena‑bling directed cell migration. Motile leuko‑cytes reorientate the microtubule‑organizing centre (MTOC), the Golgi apparatus and cisternae of the endoplasmic reticulum (ER) towards the uropod. By contrast, fibroblast‑like motile cells generally do not form uro‑pods, as their tails neither extend from the

substratum nor accumulate the characteristic glycoproteins that are observed in amoeboid motile cells. Fibroblasts localize the MTOC between the nucleus and the leading edge, in part as a consequence of their strong spreading, which is a prerequisite for their locomotion7.

Although the signalling and structural requirements of uropod formation are being characterized in molecular detail, a clear understanding of uropod function is lacking. Here, we offer a perspective on the functional aspects of uropod biology.

The uropod as a structural unitAs mentioned above, the uropod contains cytoskeletal components, organelles and adhesion receptors (FIG. 1; TABLE 1).

Cytoskeletal components. The uropod con‑tains the MTOC, the nucleation point from which the microtubule array radiates4,8. Interestingly, microtubules that are closer to the MTOC are stabilized by acetylation9, which suggests that this subset of micro‑tubules has roles in dynein‑mediated vesicu‑lar transport or as providers of mechanical support for the uropod. However, contrary to its effect on the polarization of adherent cells, microtubule disruption does not impair uropod formation in amoeboid cells but rather promotes it10,11. This is probably due to the activation of myosin light chain kinase (MLCK; also known as MYLK)12, which can promote myosin assembly and actomyosin ring contraction at the uropod neck, collaps‑ing the network of the microtubule‑associated intermediate filament vimentin towards the uropod. This might eventually result in the protrusion of the plasma membrane. Although vimentin‑deficient cells still form uropods11, it is unknown whether uropods can form in the absence of both microtubules and intermediate filaments.

Cell polarization and uropod formation are also regulated by polymerized actin net‑works. Asymmetric distribution of F‑actin in round‑shaped leukocytes precedes the development of chemoattractant‑induced cell polarity13. At the leading edge, actin adopts a lamelli podial shape, which involves a dendritic network and active polymeriza‑tion and depolymerization7. At the uropod,

o p i n i o n

Bringing up the rear: defining the roles of the uropodFrancisco Sánchez-Madrid and Juan M. Serrador

Abstract | Renewed interest in cell shape has been prompted by a recent flood of evidence that indicates that cell polarity is essential for the biology of motile cells. The uropod, a protrusion at the rear of amoeboid motile cells such as leukocytes, exemplifies the importance of morphology in cell motility. Remodelling of cell shape by uropod-interfering agents disturbs cell migration. But even though the mechanisms by which uropods regulate cell migration are beginning to emerge, their functional significance remains enigmatic.

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Nature Reviews | Molecular Cell Biology

Myosin IIChemoattractantreceptor

Active integrin Inactive integrin

ICAMs Mucins CD44 MTOC MicrotubulesF-actin

Inactive ERMsActive ERMs

Uropod

Leadingedge

Chemoattractant

PtdIns(4,5)P2

P

αβγ

ROCK

ROCK

ROCK ROCK

PKC?

MLCK

ATP ATP ATPADP+Pi

P P P P PDBL

RhoAP PP P P PP P

P PP P

P PP P

P PP P

P PP P

P PP P

P PP P

ADP+Pi ADP+Pi

WASP Arp2/3?

mDIA?

a b c

d e f

Substratum

Cell front Cell rear

Plasmamembrane

Plasmamembrane

P P P P P P

↑ [Ca2+] RhoA PP2APP1 PIPKI

F‑actin orientates parallel to the long axis of migration, which is consistent with the spe‑cific localization of the actin‑binding pro‑teins spectrin14, the ezrin–radixin–moesin (ERM) proteins15 and the motor protein myosin16. These proteins connect actin to the plasma membrane cell cortex.

Uropods are also enriched in micro‑spikes and microvilli17,18, which are actin‑based plasma membrane projections that are specialized for tactile exploration

and attachment to other cell structures. Microvilli are regulated by phosphorylation‑mediated activation of ERM proteins, which bridge actin filaments to adhesion recep‑tors19,20. Interestingly, chemoattractants induce microvilli disassembly throughout the unpolarized cell immediately before lymphocyte polarization. This is mediated by transient dephosphorylation of ERMs and activation of the GTPase RAC1 (REF. 21), suggesting that subsequent cycles of ERM

phosphorylation and dephosphorylation might induce rearward redistribution of microvilli to the uropod or simply reassemble them within it.

Organelles. Active organelles contained in uropods include the ER, the Golgi apparatus and mitochondria17. Although the main part of the cytosol accumulates passively at the rear of motile cells, Golgi and ER cisternae translocate to the uropod in association with

Figure 1 | Mechanisms of uropod formation. a | The distribution of the adhesion receptors cD44, intercellular adhesion molecules (icAMs; icAM1–3) and mucins (cD43 and P-selectin glycoprotein ligand 1 (PsGL1)); microtubules; chemoattractant receptors; ezrin–radixin–moesin (eRM) pro-teins; and integrins in spherical non-polarized non-motile leukocytes. b | chemoattractants polarize leukocytes, inducing both the binding of adhesion receptors to activated eRM proteins and the myosin ii-mediated rearward redistribution of adhesion receptors to form clusters at the cell pole, opposite the zone of chemoattractant sensing. c | Actin polymeriza-tion and actomyosin contraction provide the force that protrudes uropods in leukocytes. d–f | Putative signalling events that involve cytoskeletal elements and adhesion receptors at each step of uropod formation. d | chemoattractant activation of surface receptors at the cell front initiates divergent signalling pathways (red) that are transmitted to the rear. The schematic depicts the possible roles of RhoA–ROcK (Rho kinase) and ca2+–MLcK (myosin light chain kinase) signalling in myosin ii assembly, of protein phosphatases PP2A and PP1 in the transient inactivation of eRM proteins, and of PiPKi (type i phosphatidylinositol-4-phosphate 5-kinase; also known as PiP5K1) activation to produce phosphatidylinositol-4,5-bisphosphate

(Ptdins(4,5)P2), which converts eRM proteins to an active form at the inner

surface of the plasma membrane.The chemoattractant receptor-coupled α, β and γ subunits of heterotrimeric GTP-binding proteins are also depicted. e | Roles of eRM proteins and myosin ii in RhoA–ROcK activation and receptor clustering. Ptdins(4,5)P

2-activated eRM proteins bind to adhesion

receptors at the plasma membrane. ROcK or other kinases (for example, protein kinase c (PKc)) phosphorylate eRM proteins at their carboxyl termini and activate the guanine nucleotide-exchange factor DBL by recruiting it to the plasma membrane, providing a positive-feedback path-way for RhoA–ROcK activation that promotes actin binding and polymeri-zation (to form filamentous (F)-actin). complexes of eRM proteins and adhesion receptors are redistributed to form clusters by an actomyosin-based mechanism. f | The coordinated actions of ROcK-mediated myosin assembly, actomyosin contraction near the plasma membrane and actin polymerization — through an unknown mechanism that might involve the formin mammalian diaphanous (mDiA1; also known as DiAPH1) or the actin-nucleating complex WAsP–ARP2/3 (Wiskott–Aldrich syndrome protein–actin-related protein 2/3) — result in the protrusion of the uropod. MTOc, microtubule-organizing centre.

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the MTOC during chemoattractant‑induced polarization22. Mitochondria also redistri‑bute towards the uropod by a microtubule‑mediated mechanism that requires fission of mitochondria by DRP1 (REF. 23).

Adhesion and antigen receptors. A plethora of adhesion molecules relocate to the uropod, some in microvilli, endowing this compart‑ment with an adhesive function. These molecules include immunoglobulin (Ig) superfamily members, such as inter cellular adhesion molecules (ICAMs; ICAM1, ICAM2 and ICAM3) and CD2, mucins (CD43 and P‑selectin glycoprotein ligand 1 (PsGL1; also known as sELPLG)), L‑selectin, the hyaluronic acid receptor CD44, the heparan sulphate proteoglycan syndecan 1, the tetraspanins CD9 and CD81 and associ‑ated proteins of the EwI family (EwI2 (also known as IGsF8) and EwIF), integrins, adhesion‑related proteases (ADAM17 and cathepsin X) and the antigen receptors of B and T cells7,24–30. some of these receptors are actively targeted to the uropod by the interaction of their juxtamembrane regions with ERM proteins, which requires clusters of positively charged amino acids and crucial ser residues31–33

Signalling at the uropodMigrating cells that have uropods coordinate their activities by segregating different sig‑nalling pathways to specific compartments. Net advance is achieved by protrusion and attachment to the substratum at the front of the cell, and retraction and detachment at the rear. These opposing actions are carried out through the segregation of lipid domains and targeted polarization and activation of signalling components (FIG. 1; TABLE 1).

Lipid rafts. Chemoattractant‑induced cell polarization causes an asymmetric redistribu‑tion of cholesterol‑ and glycosphingo lipid‑rich lipid rafts towards the leading edge and the uropod. Lipid rafts in the uropod are enriched in cholesterol and the ganglioside GM1, whereas GM3 is concentrated at the leading edge34,35. Moreover, raft formation has been linked to actomyosin activity36, suggesting that the selective segregation of raft domains at these opposite compartments supports the signalling pathways that ultimately control actomyosin‑dependent cell polarization and uropod function.

Cyclic GMP and AMP. Chemoattractant‑induced production of intracellular cyclic GMP (cGMP), the product of guanylate cyclase, regulates leukocyte chemotaxis

through the activity of uropod‑located G kinase. G kinase binds to and transiently phosphorylates vimentin, causing cell polarization and the co‑redistribution of both proteins from the cytosol to the nascent uropod37. Another cyclic nucleotide, cyclic AMP (cAMP), which doubles as an extra‑cellular chemoattractant and an intracellular second messenger in D. discoideum, is essen‑tial for polarization and chemotaxis1,2. In leukocytes, most cAMP actions are mediated through the activation of cAMP‑dependent protein kinase A (PKA), which localizes to the Golgi apparatus in the uropod38,39. cAMP agonists induce uropod formation40, and activated PKA phosphorylates the GTPase RhoA on ser188, regulating cell polarization and migration39.

Rho GTPases. The Rho GTPases RhoA, Rac proteins and CDC42 are key players in the organization of leukocyte polarity5,41. These are GTP‑ and GDP‑binding proteins of the Ras superfamily, the activities of which are finely regulated by guanine nucleotide‑exchange factors (GEFs) and GTPase‑activating proteins (GAPs).

Activation of members of the Rho family seems to be restricted to either the uropod or the leading edge, and, during cell migra‑tion, Rho GTPases generate divergent signals that originate from compartments at opposite cell poles. Myosin II‑dependent retraction at the trailing edge is controlled by RhoA activation at the uropod, whereas actin polymer ization and protrusion at the leading edge are stimulated by Rac activ‑ation7,42. Negative‑feedback mechanisms allow crosstalk of this compartmentalized activation with specific Rho GTPase signal‑ling at the front and rear poles of the cell, thereby maintaining leukocyte polarization. Although crosstalk between Rac proteins and RhoA seems to negatively regulate myosin II‑dependent contraction at the rear, recent evidence indicates that RAC1, but not RAC2, is also required for Rho‑dependent actomyosin contraction at the trailing edge43. Although the exact mechanism by which RAC1 regulates the activation of Rho at the uropod is currently unknown, chemo‑attractants can induce Rac protein activation at both the leading and the trailing edges, which is compatible with regional activation

Table 1 | organelles, proteins and small signalling molecules in the uropod

uropod features components refs

Organelles Golgi apparatus, endoplasmic reticulum, MTOc, mitochondria and secretion vesicles

4,6,8,23

Plasma membrane Microvilli and microspikes 17,18

Adhesion receptors icAMs (icAM1–3), cD2, mucins (cD43 and PsGL1 (also known as seLPLG)), cD44, syndecan 1, tetraspanins (cD9 and cD81), eWi family proteins (eWi2 (also known as iGsF8) and eWiF) and integrins (LFA1, MAc1 and β1)

7,24–26,30

Proteases ADAM17 and cathepsin X 27,28

signalling receptors BcR and TcR 29,30

cytoskeletal elements Microtubules and HDAc6 8,9

Microfilaments, spectrin and eRM proteins 14,15,53

intermediate filaments (for example, vimentin) and associated plectin

11

Motor proteins (for example, myosin ii) 15,16,55

septins 81

Membrane constituents GM1, cholesterol and phosphatidylserine 34,35,76

signalling molecules G kinase, PKA, PKc, RhoA, PiPKiβ and PiPKiγ, Ptdins(4,5)P

2 and Ptdins(3,4,5)P

3

5,37–39, 50,51

vesicle trafficking machinery clathrin, AP2, dynamin 2 and PsTPiP1 52,53

Apoptosis elements cD95 and PLscR1 75,76

PDZ-containing platforms scribble and DLG 67

AP2, adaptor protein 2; BcR, B cell receptor; DLG, Discs large; eRM, ezrin–radixin–moesin; HDAc6, histone deacetylase 6; icAM, intercellular adhesion molecule; LFA1, leukocyte function-associated molecule 1; MAc1, macrophage 1 antigen; MTOc, microtubule-organizing centre; PiPKi, type i phosphati-dylinositol-4-phosphate 5-kinase; PKA, protein kinase A; PKc, protein kinase c; PLscR1, phospholipid scramblase 1; PsGL1, P-selectin glycoprotein ligand 1; PsTPiP1, Pro-ser-Thr phosphatase-interacting protein 1; Ptdins(4,5)P

2, phosphatidylinositol-4,5-bisphosphate; Ptdins(3,4,5)P

3, phosphatidylinositol-3,4,5-

trisphosphate; TcR, T cell receptor.

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of RAC1 at the uropod. By contrast, activa‑tion of RAC2 is confined to the front of the cell, where, in coordination with CDC42, RAC2 might regulate the polymerization of actin. CDC42 seems to help to maintain the uropod by inhibiting the formation of additional protrusions at the cell rear, as overexpression of a dominant‑negative CDC42 mutant impairs chemotaxis induced by the chemoattractant stromal cell‑derived factor (sDF1; also known as CXCL12) and promotes aberrant formation of lamellipodia at the uropod44.

Phosphoinositides. Phospholipid metabolic activity is in part concentrated in uropods. The phospholipid phosphatidylinositol‑4,5‑bisphosphate (PtdIns(4,5)P2) is a key second messenger in the integration of front‑to‑rear signalling in both D. discoideum and leuko‑cytes. PtdIns(4,5)P2 phosphorylation by phosphoinositide 3‑kinase (PI3K) results in the formation of phosphatidylinositol‑3,4,5‑trisphosphate (PtdIns(3,4,5)P3), a process that is counteracted by the phosphatase PTEN. In D. discoideum and leukocytes, PtdIns(3,4,5)P3 is produced mainly at the leading edge, where it regulates actin poly merization and directional migration45. PTEN is restricted to the uropod in D. discoideum but is evenly distributed throughout the cell body of polarized leukocytes. Although earlier reports suggest that neither PtdIns(3,4,5)P3 nor PTEN are absolutely essential for amoeboid‑based chemotaxis46–48, more recent findings show that, in response to the chemoattractant formyl‑Met‑Leu‑Phe (fMLP), PTEN is transiently localized at the uropod of neutrophils. Moreover, chemo‑kines that interfere with fMLP‑mediated chemotaxis disperse PTEN throughout the plasma membrane, suggesting that PTEN might regulate directional leukocyte motil‑ity, establishing hierarchical responses to chemoattractants49.

PtdIns(4,5)P2 is synthesized by type I phosphatidylinositol‑4‑phosphate 5‑kinase (PIPKI; also known as PIP5K1), and the PIPKI isoforms β and γ localize to the uropod50,51. Local production of PtdIns(4,5)P2 is associated with many functions, includ‑ing endocytosis, and PIPKIγ colocalizes at the uropod with several components of the endocytic complex that regulate membrane trafficking during cell migration. These include the coat protein clathrin, the adap‑tor protein AP2, the GTPase dynamin 2 and the PCH (pombe Cdc15 homology) family member PsTPIP1 (Pro‑ser‑Thr phosphatase‑interacting protein 1), which is involved in the regulation of actin‑based cellular

functions, including chemotaxis52,53. Recent studies suggest that calcium fluxes and pro‑tein kinase C (PKC) activities might also be localized to the uropod and are associated with cytoskeletal rearrangements during cell motility54.

Myosin ii function in the uropodThe formation of uropods in motile cells is promoted by chemoattractants, the crosslink‑ing of certain adhesion receptors and activating stimuli, such as concanavalin A, phytohaemagglutinin, CD3 antibodies and certain cytokines7,40 (FIG. 1). This shape change is associated with the rearwards redistribu‑tion of adhesion receptors and myosin II activity at the rear40,55. Both uropod forma‑tion and the concomitant co‑redistribution of adhesion receptors and ERM proteins are abolished by the treatment of cells with specific myosin ATPase inhibitors15,55. The mechanistic link between the activities of ERMs and myosin II is provided by RhoA. ERM proteins activate RhoA through a positive‑feedback mechanism (FIG. 1).

First, RhoA, through its effector Rho kinase (ROCK), can induce the phosphoryla‑tion of ERM proteins in vitro and in vivo56. second, phosphorylated ERMs strongly activate the cycling of RhoA binding to GTP and GDP through the binding and mem‑brane recruitment of the GEF DBL57. RhoA‑dependent activation of ROCK induces the phosphorylation of myosin light chain at the rear of the cell and the sub sequent assembly and activation of myosin II16,42. supporting this model, small inter fering RNA (siRNA)‑mediated downregulation of myosin heavy chain IIA inhibits uropod formation in T lymphoblasts58, and inhibition of ROCK prevents the detachment of the trailing edge in cells that are migrating on ICAM1 (REF. 59). A role in chemoattractant‑induced uropod formation has also been suggested for the small GTPase RAP1 (also known as TERF2IP)60. However, many unresolved questions remain, including whether RAP1 activates cell polarization through a Rho‑dependent mechanism, whether PKC phos‑phorylates ERM proteins in response to chemoattractants, and how myosin II regulates ERM‑mediated redistribution of adhesion receptors in motile cells.

old and new functions for uropodsMany functions of leukocytes depend on their ability to redistribute cellular components towards polarized structures. The uropod has been increasingly implicated in processes that are distinct from its initially identified functions in adhesion and motility (FIG. 2).

Intercellular adhesion and communication. several adhesion receptors and tetraspanins are concentrated at the uropod, raising the possibility that tetraspanin‑enriched micro‑domains might organize adhesion receptor nanoclusters in this structure, as described recently for endothelial cells61. This could provide a general mechanism by which leuko cytes connect with other cells to estab‑lish cell–cell interactions. In fact, a large body of morphological evidence suggests roles for uropods in intercellular processes (FIG. 2a).

First, interactions between ICAM1‑enriched uropods on macrophages mediate interferon‑γ‑induced formation of multi‑nucleated giant cells62. second, antigen‑specific motile B cells with uropods form contacts with particulate antigen‑bearing macrophages on the subcapsular area of lymph nodes, and transport antigens on their uropods towards follicular dendritic cells29. Third, the allogenic reaction of leuko‑cytes is characterized by interconnections between the lymphocyte uropods and other leukocytes6. Fourth, ICAM2‑enriched ezrin‑mediated uropods on target cells facilitate their interaction and destruction by NK cells63. Fifth, thrombin‑activated platelets bind to the uropods of fMLP‑stimulated neutrophils64. Last, during T cell and NK cell chemotaxis and trans migration, uropods establish contacts with other cells to amplify the recruitment of bystander leukocytes to inflammatory foci65,66. These findings suggest that uropod‑mediated interactions with other cells would be facilitated by the topological organization of adhesion receptors and counter‑receptors in uropods and target cells.

Additionally, the uropod can be consid‑ered to be a structure in which the cellular activation machinery (organelles, adaptors and signalling and cytoskeletal‑associated molecules) is pre‑assembled to facilitate the transmission of information between cells. A network of PDZ‑containing adaptor pro‑teins, most notably scribble and Discs large (DLG), is concentrated in the uropod30,67. Proteins of the PDZ family are scaffolds that interact with C‑terminal conserved domains of kinases and adaptor proteins to assemble signalling complexes. These signalling com‑plexes, through their binding to cytoskeletal proteins at specific cellular compartments, are responsible for asymmetrical cell shape and polarity and regulate the activity of other determinants of cell polarization. The organ‑ization of scribble and DLG complexes in the uropod holds together many of the trans‑membrane proteins and signalling complexes of the immune synapse, mirroring their distribution in cognate cell–cell interactions.

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b Cell migration

a Intercellular adhesion

d Apoptosis

c Vesicle trafficking

Passage throughconstricted spaces

Retr

actio

n

De-adhesion

Caspase activation

Raft-associatedexocytosis

Clathrin-mediatedendocytosis

Myosin IIChemoattractantreceptor

Active integrin Inactive integrin

ICAMs

Mucins CD44 FAS FASL

MTOCMicrotubules F-actin

Active ERMs Ezrin

Leadingedge

Substratum

Phosphate group

Cell motility and migration. The precise role of the uropod in regulating cell motil‑ity remains unknown. RhoA‑mediated cell retraction is an important step not only for net advance but also for the passage of cells across endothelial monolayers68 (FIG. 2b). Leukocyte migration is promoted by the activity of cathepsin X, which localizes to the uropod and modulates the binding of the β2 integrin leukocyte function‑associated molecule 1 (LFA1) to ICAM1 at the rear of the cell28,69. Here, contraction forces not only promote the detachment of the uropod from the substratum but also generate a rearward wave of contractility that produces mechanical traction from the flow of intracellular material towards the leading edge70.

A recent study, however, postulates that integrin‑mediated adhesion and uropod contraction might be dispensable for leuko‑cytes that migrate in three‑dimensional matrices. The authors propose that the main force that drives migration in three dimensions is actin‑mediated protrusion at the cell front, and that uropod‑mediated retraction is not coordinated with protru‑sion of the leading edge but is instead acti‑vated sporadically to enable the nucleus to pass through constricted spaces71. This is consistent with the proposal that the fully packed rigid microtubule network in the uropod facilitates leukocyte deformation and migration8. Thus, the uropod seems to control de‑adhesion and retraction at the cell rear during leukocyte transmigration

across endothelial monolayers, and might also enable cells to navigate through the constricted spaces of interstitial tissues.

Vesicle recycling. Bidirectional transport of vesicles is a characteristic feature of uropods (FIG. 2c). Clathrin‑mediated endocytosis of transferrin–transferrin receptor complexes takes place preferentially at the uropod plasma membrane, mainly as large clathrin structures that form complexes with the adaptor protein AP2 (REF. 52). In addition, the biosynthetic transport of proteins towards the distal region of the uropod occurs through their incorporation into lipid rafts34. The localization of clathrin and lipid rafts to the uropod depends on RhoA–ROCK activation and myosin II contractility36,52.

Figure 2 | Biological functions of uropods. a | Uropods might function as connecting stalks that facilitate the interaction of a cell with others in pro-cesses such as antigen transport, cytotoxicity, extravasation or in the foci of cell proliferation. b | A dual role for uropods in cell migration has been pro-posed. First, myosin ii-dependent contraction at the rear enables de-adhesion from the substratum and cell body retraction, thereby facilitating net cell advance. second, the package of microtubules, intermediate filaments and actomyosin-dependent forces in the uropod reduces rigidity and facilitates

the generation of forces that are required for cell deformability, facilitating cell passage through constricted spaces during leukocyte transmigration. c | The uropod is a site of active bidirectional traffic, in which clathrin- mediated endocytosis and lipid raft-associated exocytosis to the distal pole are coordinately regulated. d | Under some circumstances, FAs-mediated apoptosis is initiated in the uropod of polarized cells by a mechanism that involves ezrin. eRM, ezrin–radixin–moesin; F-actin, filamentous actin; FAsL, FAs ligand; MTOc, microtubule-organizing centre.

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The highly polarized distribution of the Golgi apparatus, clathrin and lipid rafts at the trailing edge, and the abilities of these struc‑tures to mediate endocytosis and exo cytosis through distinct routes, support the role of the uropod as a specialized platform for vesicle trafficking. The fact that interfering with vesicle trafficking at the uropod disrupts cell polarization and migration indicates that these processes are interlinked. Hence, the uropod might function as an endocytic sink for plasma membrane recycling and afflux towards the leading edge.

Viral infection and apoptosis. several studies provide evidence that uropods are important for viral spreading. HIv‑1 can initiate infec‑tion as free virions, but viral spreading might be enhanced by contact through the uropods of infected and target cells72. Lipid rafts have important roles in the preferential budding of HIv‑1 from uropods and in the lateral assem‑blies that are required for HIv‑1 infection73. Moreover, the fusion of plasma membranes to form syncytia, which are required for the pro‑apoptotic effects of HIv‑1, occurs through the establishment of contacts between the uropods of infected monocytes74.

Other studies indicate a role for the uropod as a multimolecular death sig‑nalling complex (FIG. 2d). For example, ezrin‑mediated polarization of FAs to the leukocyte uropod is associated with increased susceptibility to FAs‑mediated apoptosis75. supporting this view, movement of phosphatidylserine to the uropod plasma membrane outer leaflet is a marker of apop‑tosis in fMLP‑stimulated neutrophils, prob‑ably as a consequence of the localized activity of phospholipid scramblase 1 (PLsCR1)76. However, given that ezrin is a transmitter of cytoskeletal‑mediated anti‑apoptotic and survival signals77, the precise mechanism by which the binding of FAs to ezrin at the uropod might interfere with surviving signals remains uncertain.

Conclusions and perspectivesThere is now a whole body of evidence that indicates that uropods are essential for important cellular functions. In leukocytes, uropods contribute to a range of immune functions, from facilitating cell motility and chemotaxis towards inflamed tissues to establishing intercellular adhesion and cell–cell communication and sustaining vesicular trafficking. Although rapid progress is being made towards defining the mechanisms of uropod formation and the functions that they have, the physiological significance of uropods remains poorly understood.

Uropod‑deficient neutrophils from Rac1–/– mice are poorly recruited to the lung in a model of fMLP‑induced inflammation78, and leukocytes from patients with wiskott–Aldrich syndrome display reduced cell polarization and migration in response to chemoattractants, indicating that wiskott–Aldrich syndrome protein (wAsP) partici‑pates in the cytoskeletal rearrangement that leads to uropod formation79. Furthermore, elevated levels of collapsin response media‑tor protein 2 (CRMP2), a semaphorin signal‑transducing protein that localizes to the uropod, are associated with increased transmigration of T cells in patients with neuroinflammatory diseases80.

Other questions need to be addressed to obtain a complete understanding of the role of the uropod in leukocyte migration. For example, it is unclear whether mDIA1 (also known as DIAPH1), the mammalian homologue of Drosophila melanogaster Diaphanous, or the actin‑nucleating com‑plex between wAsP and the actin‑related protein (Arp)2/3 complex link upstream Rho GTPase signals to actin polymerization and branching during uropod formation (FIG. 1). It is also possible that the leukocyte uropod works as a chemoattractant pump, as described for D. discoideum during streaming. The polarized secretion of chemo‑attractants at the uropod of leukocytes would provide insights into why vesicle trafficking in uropods regulates cell migration.

Other questions include how the uro‑pod controls rigidity, deformability and cell lengthening during leukocyte movement. In this regard, it is unknown whether corti‑cal tension at the plasma membrane differs between the rear and the front of the cell, and nanoscale analysis with advanced microscopy techniques, such as atomic force microscopy and optical magnetic tweezers, will help to address this issue. A role in controlling rigidity and deform‑ability is supported by the presence of a cell‑ular ‘corset’ of septins at the midbody that controls uropod extension in leukocytes81. In neurons, septins are localized at the neck of dendritic spines, and function as a diffu‑sion barrier that controls receptor trafficking and spine compartmentalization82. whether septin rings in leukocytes also act as cellular barriers that create specialized plasma mem‑brane microdomains at the rear of the cell deserves to be investigated. Elucidation of the mechanisms by which multiple extracellular inputs are integrated into the uropod should provide a more fine‑tuned hypothesis that will help to reveal the role that this enigmatic cell compartment has in leukocyte biology.

Francisco Sánchez-Madrid and Juan M. Serrador are at the Departamento de Biología Vascular e

Inflamación, Centro Nacional de Investigaciones Cardiovasculares (CNIC), Madrid, Spain.

Francisco Sánchez-Madrid is also at Servicio de Inmunología, Hospital Universitario de la Princesa, Universidad Autónoma de Madrid, Madrid, Spain.

Correspondence to F.S.-M. e-mail: fsanchez.hlpr@salud.madrid.org

doi:10.1038/nrm2680 Published online 17 April 2009

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AcknowledgementsF.S.-M. is supported by grants SAF2008-02635, INSINET-0159/2006 from Comunidad de Madrid, RD06/0014-0030 from Red Temática de Investigación Cooperativa en Enfermedades Cardiovasculares and FIPSE (Fundación para la Investigación y Prevención del SIDA en España) 36289/02. J.M.S is supported by grants PI070356, SAF2008-01339-E and Contrato-Investigador FIS (Ministerio de Ciencia e Innovación, Spain). Editorial support was provided by S. Bartlett. The authors thank A. Shaw, R. Gonzalez Amaro and M. Vicente Manzanares for critical reading of the manu-script. The Centro Nacional de Investigaciones Cardiovasculares (CNIC) is supported by the Spanish Ministry of Science and Innovation and the Pro-CNIC Foundation.

DATABASESOMiM: http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=OMiMWiskott–Aldrich syndromeUniProtKB: http://www.uniprot.orgcD2 | cD9 | cD43 | cD44 | cD81 | cDc42 | eWi2 | icAM1 | icAM2 | icAM3 | L-selectin | MLcK | PsGL1 | PTeN | RAc1 | RAc2 | RAP1 | RhoA | syndecan 1 | vimentin

FURTHER inFoRMATionFrancisco sánchez-Madrid’s homepage: http://www.cnic.es/index1.php?inc=2&secc=investigacion&idioma=uk

all links are active in the online pdf

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