TRENDS in Genetics Vol.18 No.12 December 2002

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636 Review

Hildegard Tekotte

Ilan Davis*

Wellcome Trust Centre forCell Biology, ICMB, King’sBuildings, The Universityof Edinburgh, MayfieldRoad, Edinburgh, UK EH9 3JR.*e-mail:ilan.davis@ed.ac.uk

In general, mRNA localization provides a moreefficient mechanism of concentrating proteins at theirsite of function in the cell rather than transportingmany individual protein molecules after theirsynthesis. With few exceptions [1,2], mislocalizedmRNAs lead to incorrectly localized protein.Moreover, in many cases mRNA localization workshand in hand with the regulation of translation, sothat the translation of transport intermediates isrepressed until they reach their correct destination. A detailed description of translational regulation isbeyond the scope of this article and has been coveredextensively in several excellent recent reviews [3,4].There are many variations on this functional themethat have been comprehensively reviewed elsewhere[1,5] (also described in Fig. 1 and Boxes 1 and 2).mRNA localization is known to have important roles in cell polarity, segregation of cytoplasmicdeterminants during asymmetric cell divisions,targeting signals to particular groups of cells,specifying germ cells, initiating or restrictingdiffusion of nuclear transcription factors and in thespecification of embryonic axes. Although the field ofmRNA localization initially focused on functionalstudies, more recently it has made considerableprogress in determining how mRNA becomesdistributed asymmetrically in the cell.

The mechanism of mRNA localization

Several possible mechanistic models have beenproposed for how mRNA becomes localized. However,these have remained largely untested until recently(reviewed in [4]). The models include directionalexport of mRNA from the nucleus [6], generaldegradation and selective stabilization at the site oflocalization [7], diffusion and specific retention at thesite of localization [8] or active transport along thecytoskeleton using molecular motors [9]. With theexception of directional nuclear export of mRNA [10],the other mechanisms are all thought to have a role in

some specific cases. For example, hsp83 mRNA isselectively localized to the posterior of the Drosophilablastoderm embryo by degradation in all other partsof the embryo and selective stabilization at theposterior [7], and osk mRNA localization can involveat least some diffusion and anchoring [8]. In manycases, mRNA localization depends on an intactcytoskeleton [1], but it is not always known whetherthe cytoskeleton is required for active transport,anchoring or both. For example, Vg1 mRNAlocalization depends on both actin and microtubules(MTs) [9], and bicoid (bcd) mRNA localization in theDrosophila oocyte is MT dependent [11]. Although therole of the cytoskeleton in anchoring remains poorlycharacterized, in the past two years several clearcases of transport of mRNA along the cytoskeletonhave emerged (Fig. 2). It seems likely that activetransport along the cytoskeleton will be the mostcommon mechanism of mRNA localization.

Yeast ASH1 mRNA localization requires myosin V class motorsThe best understood example of an mRNA that issegregated during asymmetric cell division is ASH1 mRNA in budding yeast. Ash1p [12–14] is atranscription factor that suppresses expression of HO,an endonuclease that promotes mating-type switching.Localization of the ASH1 mRNA to the bud, the futuredaughter cell [15,16], restricts Ash1p expression to thedaughter cell and allows mating-type switching only inthe mother cell (Fig. 1). More recently, localization ofanother yeast transcript, IST2, has been shown tofunction in the targeting of a membrane protein to thebud [17], and other examples are likely to follow. Themovement of particles containing ASH1 mRNA can bevisualized in living yeast cells by tagging the mRNAwith a sequence to which a green-fluorescent protein(GFP) fusion binds specifically [18]. These studies and others have highlighted the actin-dependentmovement of the particles from the mother cell to thebud [18,19]. Screens for mutations that disrupt ASH1mRNA localization have uncovered five loci named theShe genes, which are all essential for the process.She1/myo4, an unconventional (non-muscle) class Vmyosin motor [12,13] (Fig. 3) is required for thetransport of ASH1 mRNA along actin into the bud (see the end of the article for a description of some of the other SHE mutants).

In contrast to yeast, myosin has not yet beenshown to be required directly for mRNA localization

Intracellular mRNA localization is a common mechanism of post-transcriptional

regulation of gene expression. In a wide range of organisms, mRNA localization

coupled with translational regulation target the proteins to their site of function.

Here, we describe recent exciting evidence that some mRNAs are transported as

particles along the cytoskeleton by the molecular motors dynein, kinesin or

myosin. We discuss the key questions of how localized mRNAs might be linked

to motors and what determines their cytoplasmic destinations.

Published online: 01 November 2002

Intracellular mRNA localization:

motors move messages

Hildegard Tekotte and Ilan Davis

in Drosophila. However, the myosin-related protein,Tropomyosin II (TmII) is required for osk mRNAlocalization [20], which is both actin and MTsdependent [8]. In skeletal muscle, TmII regulatesmuscle contraction by affecting the interactionbetween myosin and actin, but its role in othercellular contexts is unclear. In tmII mutants, oskmRNA is unlocalized, but this defect is not due to adisruption of MT polarity. Although the precise role

of TmII protein in osk mRNA localization is unclear, it could be required for actin-and myosin-dependentanchoring at the posterior of the oocyte.

Drosophila osk mRNA localization requires the Kinesin Imotor for transport to the plus ends of MTsFor some time it has been known that mRNA encodingmyelin basic protein (BMP) is localized inoligodendrocytes by an MT-dependent mechanism thatrequires kinesin, a plus-end-directed MT-associatedmotor [21]. More recently, tau mRNA particles havebeen found to localize to axons and growth cones in akinesin-dependent manner [22]. It seems highly likelythat many more examples of kinesin-based motility ofmRNA will be discovered in nerve cells and other celltypes. Two years ago it also became clear that kinesin I(Fig. 3) is required for osk mRNA localization at theposterior of the Drosophila oocyte [23], because oskmRNA is unlocalized in kinesin heavy chain (khc) [24]null mutant egg chambers. This disruption is not dueto a change in MT polarity and it seems most likely thatkinesin is involved in the transport of osk mRNA to theposterior where the plus ends of microtubules arelocated. However, in the absence of direct visualizationof osk mRNA motility to the posterior, there are otherpossible interpretations. For example, a recent studysuggests that kinesin I also acts to exclude osk mRNAfrom the oocyte cortex where some MT minus ends arelocated [25]. Targeting Osk function to the posteriorinvolves additional mechanisms acting together withosk mRNA localization, probably to ensure fidelity ofprotein targeting. First, diffusion and posterioranchoring might also have a role in addition to activetransport [8]. Second, osk mRNA is only enriched at the posterior [26] and the unlocalized mRNA istranslationally repressed. Third, localized Osk proteinis specifically stabilized through phosphorylation byPar-1 [27], a kinase with a role in axis specification thatis conserved between worms and flies [28].

Pair-rule, wingless and probably bcd mRNA localizationrequire cytoplasmic dyneinIn addition to localized maternal transcripts, such asosk, a large number of zygotically transcribed mRNAsalso show asymmetric distributions in the embryo(Box 1, Fig. 1). Recently, the localization of wingless (wg)mRNA and the pair-rule mRNAs fushi tarazu (ftz) andrunt (run) was shown to be dependent on cytoplasmicdynein (Figs 2,3). First, using modified assays thatbuilt on previous studies of the fate of injected mRNA [8,29], it was discovered that wg, ftz and runRNAs labeled in vitro with fluorescent UTP, move asparticles in direct paths to the apical cytoplasm wheninjected into the basal cytoplasm or yolk [10]. Therefore,all the factors required for their localization can berecruited in the cytoplasm. This process requires MTsbut not actin [10], in agreement with previousobservations [29,30]. Second, it was also shown thatapical localization of blastoderm mRNA requirescytoplasmic dynein and dynactin [10], using inhibitory

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TRENDS in Genetics

ASH1 mRNA in budding yeast

Ant

Ant

Post

Post

PostAnt

Dorsal

Ventral

(b)(a)

(c)

(d)

(e) (f)

gurken (grk), bicoid (bcd), oskar (osk) and nanos(nos) mRNA in a fly oocyte

Apically localizedfushi tarazu (ftz) mRNAin a fly syncytialblastoderm embryo

Unlocalized Krüppel (Kr)anterior bcd andposterior osk and nosmRNA in a fly syncytialblastoderm embryo

Vg1 and An mRNA in a frog oocyte actin mRNA in a chicken fibroblast cell

Fig. 1. Key examples of the function of mRNA localization. (a) ASH1 mRNA (red) localized to the budtip in a budding yeast cell, thus targeting the Ash1 protein only to the bud nucleus. Ash1p suppressesmating-type switching in the bud, so that it only occurs in the mother cell. (b) A Drosophila eggchamber showing the pattern of localization of four key mRNAs in the oocyte. bicoid (bcd) mRNA(green) is localized in an anterior ring, gurken (grk) mRNA (red) is localized in the dorso-anteriorcorner near the oocyte nucleus and oskar (osk) and nanos (nos) mRNA (blue) are localized at theposterior pole (see Box 1 for further details). (c) A Drosophila syncytial blastoderm embryo showingthe pattern of expression of the pair-rule transcript fushi tarazu (ftz) in seven expression stripes, orrings, around the periphery of the embryo. ftz mRNA, like the segment polarity transcript, winglessand all other pair-rule mRNAs is apically localized ‘above’ each nucleus in the expression stripe (see Box1 for further details). (d) A Drosophila syncytial blastoderm embryo showing the pattern ofexpression of the gap transcript Krüppel (Kr, red) and posteriorly localized osk and nos mRNA (blue)and anteriorly localized bcd mRNA (green). Kr is expressed in a central domain and unlocalized withrespect to the peripheral nuclei, being found ‘above’ (apically) and ‘below’ (basally) the nuclei. (e) A Xenopus oocyte showing vegetal localization of some mRNAs, such as Vg1 (green) and animallocalization of other mRNAs, such as An1 (red) (see Box 2 for further details) (f) Actin mRNA (red)localization to the leading edge of chicken fibroblast cells enriches actin protein at its site ofpolymerization and is important for cell motility (see Box 2 for further details).

antibodies, mutants that reduce the speed of motilityand an excess of dynamitin, which acts as a dominantinhibitor of dynactin. Two transacting factors that arerequired for oocyte specification and implicated in thefunction of cytoplasmic dynein, Bicaudal-D (BicD) andEgalitarian (Egl), are required for pair-rule mRNAlocalization in the embryo and are recruited by theinjected pair-rule RNA [31] (see below).

However, the potential role that cytoplasmic dyneinplays in mRNA localization in the oocyte is less clear.The Swallow (Swa) protein which is required for bcd

mRNA localization (Fig. 2) binds both RNA and thedynein light chain (Fig. 3), as shown by yeast interactionassays [32]. Swa might therefore be the protein thatlinks bcd RNA to dynein motors. An alternativeinterpretation is that Swa could be transported to theanterior independently of bcd RNA where it functions toanchor the bcd mRNA once it becomes localized by othermeans [5]. Other circumstantial evidence for motilityof bcd mRNA in the oocyte comes from studies ofExuperantia (Exu), a protein required for bcd mRNAlocalization. Exu–GFPfusions form particles that move

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In Drosophila, mRNA localization is very common. More than 20localized maternal mRNAs have been studied, many in great detail [a],and perhaps as many as 10% of all transcripts present in the oocyte arelocalized [b]. The best studied examples are the maternal transcriptsgurken (grk), oskar (osk), bicoid (bcd) and nanos (nos) and eachrepresents a variation on the functional theme of targeting proteins totheir site of function (Fig. 1).

grk encodes a TGF-α homologue, which plays a central role inestablishment of the antero-posterior and dorso-ventral axes of theoocyte and embryo [c,d]. Grk signaling is required at two stages indevelopment, when it instructs the overlying monolayer of columnarepithelium (follicle cells) to adopt particular identities. The localization of grk mRNA targets the signal so that it is only sent to restricted groupsof follicle cells. The first Grk signal instructs posterior follicle cells thatthen signal back to the oocyte and repolarize their MTs and establish the antero-posterior axis. The oocyte nucleus then migrates to thedorso-anterior corner of the oocyte, where the second Grk signalinstructs the overlying follicle cells to become dorsal, thus setting up thedorso-ventral axis. There is now some evidence that kinesin and dyneinare both required for grk mRNA localization and dorso-ventral axisspecification [e,f]

osk mRNA is localized at the posterior of the oocyte and embryo,where it is the key determinant of posterior structures and germ cellformation. Mislocalizing osk mRNA at the anterior is sufficient to inducegerm cell formation at the anterior [g] and Osk protein recruits theproducts of many other posterior group genes [h]. osk mRNAlocalization is not complete, so the translation of unlocalized osk mRNAmust be repressed [i].

nanos mRNA is localized to the posterior of the oocyte, where it isrecruited by Osk protein [h]. The majority of nos mRNA remainsunlocalized in the embryo and must be translationally repressed [i,j].Nos protein is required to suppress the maternally encoded Hunchbacktranscription factor at the posterior. Nos also has an independentfunction in later development in germ cell migration and the differentfunctions of Nos map to distinct domains of the protein [k].

bcd mRNA localization to the anterior of the oocyte is essential for setting up the antero-posterior axis when it is translated in theembryo. It encodes a transcription factor that regulates the expression of a hierarchy of segmentation genes that define the details ofantero-posterior patterning. Bcd protein diffuses away from its anteriorsource, thus giving rise to a gradient of nuclear protein with a highconcentration near the anterior and low concentration at the posterior.Proteins required for bcd mRNA localization include: Staufen (Stau),Exuperantia (Exu) and Swallow (Swa) [l] (see text for further details).

Zygotic transcripts are specifically localized in Drosophilablastoderm embryos, for a variety of functions. Gap transcripts arelocalized to sub-regions of the antero-posterior axis that are specifiedby the Bcd protein gradient. Like bcd mRNA, gap transcripts provide asource of translated protein that form local morphogenetic gradientsof nuclear transcription factors in particular regions of the syncytialblastoderm. In essence, the gap proteins function by providingdifferent combinations of protein gradients at different positions alongthe antero-posterior axis, thus activating specific downstream pair-rulegenes, such as fushi tarazu (ftz), hairy (h) and runt (run). Each of thepair-rule proteins is expressed in seven narrow segmental stripes, thus specifying fine positional information and segmental patterns in the embryo. Segment polarity genes, such as wingless (wg),

which encodes a Wnt-like signal, refine the positional information even further.

In contrast to gap proteins, pair-rule proteins do not diffuse in thesyncytial blastoderm embryo. Instead they are expressed in sevennarrow stripes that accurately mirror the 7 stripes of mRNA from whichthey are translated (Fig. 1). Pair-rule and wingless (wg) transcripts areapically localized in caps above each syncytial nucleus within anexpression stripe, whereas gap transcripts are unlocalized, beingpresent above and below each nucleus in their expression domains [m].It is thought that apical localization of pair-rule transcripts restrictsdiffusion of the protein before import into the blastoderm nuclei, thusmaintaining a tight correspondence between protein and RNA stripepattern at a single cell level [m]. By contrast, apical localization of wgmRNA is required to target the secretion of the Wg signal to the apicalsides of cells in the gastrulating embryo [n]. crumbs mRNA is alsoapically localized and encodes a membrane protein required forepithelial polarity [o] in the embryo.

References

a Palacios, I.M. and Johnston, D.S. (2001) Getting the message across: theintracellular localization of mRNAs in higher eukaryotes. Annu. Rev. CellDev. Biol. 17, 569–614

b Dubowy, J. and Macdonald, P.M. (1998) Localization of mRNAs to the oocyteis common in Drosophila ovaries. Mech. Dev. 70, 193–195

c Gonzalez-Reyes, A. et al. (1995) Polarization of both major body axes inDrosophila by Gurken-Torpedo signaling. Nature 375, 654–658

d Neuman-Silberberg, F.S. and Schüpbach, T. (1993) The Drosophiladorsoventral patterning gene gurken produces a dorsally localized RNA andencodes a TGF α-like protein. Cell 75, 165–174

e Januschke, J. et al. Polar transport in the Drosophila oocyte requires Dyneinand Kinesin I cooperation. Curr. Biol. (in press)

f Brendza, R. et al. (2002) Dorsal–ventral axis determination in Drosophilarequires kinesin I, a plus-end-directed microtubule motor. Curr. Biol.121, 1541–1545

g Ephrussi, A. and Lehmann, R. (1992) Induction of germ cell formation byoskar. Nature 358, 387–392

h Ephrussi, A. et al. (1991) Oskar organizes the germ plasm and directslocalization of the posterior determinant nanos. Cell 66, 37–50

i Bergsten, S.E. and Gavis, E.R. (1999) Role for mRNA localization intranslational activation but not spatial restriction of nanos RNA.Development 126, 659–669

j Gavis, E.R. and Lehmann, R. (1994) Translational regulation of nanos byRNA localization. Nature 369, 315–318

k Arrizabalaga, G. and Lehmann, R. (1999) A selective screen reveals discretefunctional domains in Drosophila nanos. Genetics 153, 1825–1838

l St Johnston, D. et al. (1989) Multiple steps in the localization of bicoid RNAto the anterior pole of the Drosophila oocyte. Development 107, 13–19

m Davis, I. and Ish-Horowicz, D. (1991) Apical localization of pair-ruletranscripts requires 3′ sequences and limits protein diffusion in theDrosophila blastoderm embryo. Cell 67, 927–940

n Simmonds, A.J. et al. (2001) Apical localization of wingless transcripts isrequired for wingless signaling. Cell 105, 197–207

o Tepass, U. et al. (1990) Crumbs encodes an EGF-like protein expressed onapical membranes of Drosophila epithelial-cells and required fororganization of epithelia. Cell 61, 787–799

Box 1. The function of mRNA localization in Drosophila

from the nurse cells to the oocyte and within theoocyte [33]. Furthermore, fluorescently labeled bcdmRNA is able to move from the nurse cells into theoocyte and then localize within the oocyte [34]. In theabsence of a direct visualization of bcd mRNA particlemovement and a definitive genetic or other test ofwhether this movement is dynein dependent, themechanism of localization of bcd mRNA remains to befully determined.

Transacting factors required for mRNA localization

In most cases, the destination of mRNAs is determinedby specific sequences in their 3′ untranslated regions(UTRs), often referred to as ‘zip codes’. In the case ofVg1 and actin mRNAs, the zip codes are small primaryRNA sequences [35], but in most cases they are difficultto define and are probably recognized at the level ofRNA structure. The zip codes are recognized bytransacting proteins that determine their correctcytoplasmic destinations. However, identifying the key

transacting factors that provide specificity has notbeen easy because, during its transcription, mRNAbecomes coated with many proteins required for adiverse range of functions. These include cap binding,

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One of the most compelling demonstrations of the functional importance of mRNAlocalization has come from the study of actin mRNA localization in chicken fibroblastcells (Fig. 1). Actin mRNA localization is important for the polarization of fibroblastcells, by targeting the assembly of actin fibres to the leading edge of the cell. Here,actin-rich outgrowths provide the mechanism and specify the direction of cell motility.When actin mRNA localization is disrupted, using antisense oligonucleotides againstthe cis-acting zip code (the RNA signal required for localization) [a], cell polarity andmotility are disrupted and the cells become sick and disorientated [b].

mRNA localization is also important in other vertebrate somatic cells such asoligodendrocytes [c]. In particular, different mRNAs are sorted to distinct regions of dendrites and axons [d]. When coupled with translational regulation, mRNAlocalization provides a rapid means of regulating nerve cell function independentlyof transcription in the nucleus. Such control is particularly important in the nervoussystems of large animals, where axons can be many metres long and even fasttransport takes many hours or days.

mRNA localization is important for muscle cell specification in primitive ascidianchordate eggs [e] and also has an important role in at least some vertebrate oocytesand embryos. In Xenopus oocytes a large number of transcripts are specificallylocalized to either the animal or vegetal hemisphere. The first to be identified wereAn1, An2, and An3 mRNAs in the animal hemisphere, and Vg1 mRNA in a vegetalcrescent (Fig. 1) [f]. More recently, many more localized transcripts have beendiscovered [g]. Although the functional significance of many of these is still beinginvestigated, in the case of VegT, mRNA localization in the vegetal hemisphere isthought to have a crucial role in targeting Wnt and TGF-βsignals during mesoderminduction, the process by which vegetal tissues signal to a region of animal cellsinstructing them to become mesoderm rather than ectoderm (reviewed in [h]). It is stillunresolved whether mammalian follicles (oocytes) have any pre-localized mRNAsthat function in embryonic axis specification similar to those in flies and frogs.

References

a Kislauskis, E.H. et al. (1994) Sequences responsible for intracellular-localization ofβ-actin messenger RNA also affect cell phenotype. J. Cell Biol. 127, 441–451

b Kislauskis, E.H. et al. (1997) Beta-Actin messenger-RNA localization and proteinsynthesis augment cell motility. J. Cell. Biol. 136, 1263–1270

c Ainger, K. et al. (1993) Transport and localization of exogenous myelin basic proteinmRNA microinjected into oligodendrocytes. J. Cell Biol. 123, 431–441

d Job, C. and Eberwine, J. (2001) Localization and translation of mRNA in dendrites andaxons. Nat. Rev. Neurosci. 2, 889–898

e Nishida, H. and Sawada, K. (2001) macho-1 encodes a localized mRNA in ascidian eggsthat specifies muscle fate during embryogenesis. Nature 409, 724–729

f Rebagliati, M.R. et al. (1985) Identification and cloning of localized maternal RNAs fromXenopus eggs. Cell 42, 769–777

g Mowry, K.L. and Cote, C.A. (1999) RNA sorting in Xenopus oocytes and embryos.FASEB J. 13, 435–445

h Kloc, M. et al. (2002) Mechanisms of subcellular mRNA localization. Cell 108, 533–544

Box 2. The function of mRNA localization in vertebrate cells

TRENDS in Genetics

Minusends

+

Mic

rotu

bule

s

kinesinIcytoplasmicdynein

Swallow?

myosinV

She2She3

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Actin

oskarmRNA

ASH1mRNA

fushi tarazumRNA

bicoidmRNA

wingless

mRNA

?

EgalitarianBicaudal-D?

Motors

Linkers

Fig. 2. Recently discovered examples of molecular-motor-mediatedtransport of mRNA along MTs and actin, and the putative linkers thatcould provide specificity. Different coloured ants represent differentkinds of motors and the putative linkers between RNA and motors areshown in the key. MTs are the predominant cytoskeletal element in theinterior of cytoplasm and are polymerized from their minus ends, whichare often colocalized in interphase (shown as a circle). They provide theprinciple ‘tracks’ on which long distance transport by the molecularmotors dynein and kinesin occurs. Dynein transports to the minusends, and kinesin to the plus ends of MTs. Actin is predominantly at thecell cortex and is thought to be required for short-range transport.There is a large family of so called unconventional myosins, withdifferent family members being thought to be required for transport ofdistinct cargo. MyosinV is required for Ash1 mRNA localization.

splicing, polyadenylation, stability, RNA surveillance(monitoring translatability by nonsense-mediateddecay) and maintaining RNA structure andtranslatability in the cytoplasm. Furthermore, thecomposition of RNA particles changes during mRNAprocessing, its export from the nucleus [36] and atdifferent stages of localization in the cytoplasm. Forexample, Squid, the Drosophilahomologue of hnRNPA1,which is required for general mRNA export from thenucleus, is also required for at least one step of grkmRNA localization. There are three isoforms of Squid,one isoform is probably bound to grk mRNA before andanother after its export from the nucleus. It is thoughtthat the cytoplasmic isoform of Squid is requiredspecifically for grk localization [37]. Biochemicalapproaches have been partly successful in identifyingnew factors in Drosophila [38] and Xenopus [39] thatbind to mRNA and are required for mRNA localization.

In the future it is likely that such approaches will becombined with mass spectroscopy to identify all thefactors present in particles containing localized mRNA.However, a determination of the function of individualprotein components in RNA particles will also requiregenetic and cell biological methods.

In the case of pair-rule and wg transcripts, all thefactors required for apical localization are recruited inthe cytoplasm and are maternally supplied [40].However, osk and probably other transcripts mustrecruit factors within the nucleus that function inmRNA localization in the cytoplasm [41,42]. Work inhuman cells shows that in all intron-containingtranscripts, the site at which an intron is removed ismarked by proteins that form an exon–exon junctioncomplex (EJC). In Drosophila, this complex includesY14, Mago nashi and probably recruits Barentsz in the cytoplasm. Each of these factors is required for osk mRNA localization to the posterior of the oocyte [43–45]. The EJC also includes spliceosomecomponents [41,46], as well as factors required torecruit NXF1 (yeast Mex67 and human TAP). NXF1 isnot part of the EJC and functions as a receptor forexport of all mRNAs from the nucleus [47]. It probablydoes not have a specific role in cytoplasmic localizationof mRNA [10]. A big challenge for the future is toexplain how a complex such as the EJC, which binds toall spliced mRNAs and interacts with general nuclearexport machinery, can perform specific roles in thelocalization of only a specific subset of mRNAs.

Another transacting factor with multiple functionsis Staufen (Stau), probably the best understoodRNA-binding protein required for mRNA localizationin Drosophila. Stau is a double-stranded RNA(dsRNA)-binding protein that is required forlocalization, translation derepression, and anchoringof several different mRNAs in several different celltypes. It colocalizes with osk mRNA at the posterior of the oocyte and is required for its localization,anchoring and translation. Stau is also required forprospero (pros) mRNA and protein localization inembryonic neuroblasts, where pros functions inassignment of cell identity during asymmetric celldivisions. In addition, Stau is involved in anchoringbcd mRNA at the anterior (reviewed in [48]). Stauprotein consists of five dsRNA-binding domains anddistinct domains mediate the different functions ofthe protein [49]. Domain 2 is required for theMT-dependent localization of osk mRNA, and domain 5is required for the translational derepression oflocalized osk mRNA and actin-dependent localizationof pros mRNA. However, it is still not known whetherStau provides the link between osk mRNA and theKinesin I motor that transports it to the posterior.

Linkers between motors and RNA cargo

Linkers between RNA and cytoskeletal motors arelikely to be key transacting determinants of thedestination of mRNA transport. It is probable thatsome linkers are shared among different classes of

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(a) (b)

(c)

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Light chainHeavy chain

Motor domain

Globular tail

Receptor

Heavy chainmotor domain

Intermediateand light chains

Dynein

Dynactin

Kinesin

Myosin

p150/gluedp24p50/dynamitinARP1cappingp25, p27

Actin filament

Microtubule

Cytoplasmic dynein

Fig. 3. A basic overview of the molecular motors kinesin, dynein and myosin. In general, all themotors are large multiprotein complexes with a dimer structure. They each consist of a globular headdomain, which contacts the cytoskeleton and provides the force for motility through hydrolysis of ATP,and tail domains, which are more divergent and likely to be involved in cargo recognition. (a) Kinesinis in general a plus-end-directed MT motor. However, there are many kinds of kinesins, includingminus-end-directed motors. (b) Cytoplasmic dynein, the major MT minus-end-directed motor in thecell, is responsible for the transport of many cargoes. It is the largest of the motors and has anaccessory multiprotein complex, dynactin, which is required for dynein processivity to MTs, but notfor force generation. Dynactin consists of many proteins including p150/glued and p50/dynamitin, as well as actin-related proteins and could be involved in cargo recognition. (c) Non-muscle myosins(so called unconventional myosins) are a large and diverse family of actin-based motors.

cargo including mRNAs. Comprehensive lists ofmotor-cargo interactions have been published veryrecently in two excellent reviews [50,51], but a fewparticularly relevant examples will be described here.

Myosin linkersASH1 is the best characterized myosin-dependentlocalized mRNA. SHE genes were identified in ascreen for factors essential for the transport of ASH1mRNA to the bud. She1/Myo4 is a type V Myosin, She2is a new type of dsRNA-binding protein that binds tothe localization signals within ASH1 mRNA and She3links She2 to myosin heavy chain [19]. It seems likelythat myosin-based mRNA transport systems could bewidespread in many organisms and equivalent linkersto She2 and She3 could exist for other classes ofunconventional myosins and other classes of mRNAs.However, such linkers remain to be discovered.

Dynein linkersThe 3′ UTR of parathyroid hormone mRNA bindsdirectly to dynein light chain [52], raising the intriguingpossibility that some mRNAs are transported oranchored by direct binding to dynein or dynactin.However, as with myosin, it is most likely that theinteractions between dynein and its RNA cargoes aremediated by specific protein linkers. Several differentcomponents of the dynein and dynactin complexeshave been implicated in cargo binding; for example,dynactin [50,51], the dynein accessory complex thatinteracts with MTs and increases the processivity of the motor [53] without providing the power formovement. The p150/glued subunit of dynactininteracts with the dynein intermediate chain, whichexists in several isoforms [54]. This interaction isimportant for the function of the motor [54], raisingthe interesting possibility that different subtypes ofdyneins transport different kinds of cargo.

BicD [55] and Egl, factors required duringoogenesis for specification of the oocyte, are goodcandidates for linkers between dynein and pair-ruletranscripts. Both factors are present in the samecomplex [56] and are recruited by injected pair-ruleRNA in the blastoderm embryo [31]. Both genes are required for apical localization of endogenouspair-rule transcripts [31]. BicD does not bind RNA,but interacts with dynein in mouse and human cells,suggesting that it acts as an indirect linker betweenpair-rule transcripts and dynein [31]. Lissencephaly-1(Lis-1) has been implicated in several dynein- andBicD-dependent processes, such as nuclear migrationand position in the oocyte and other tissues [57,58].However, it remains unclear whether dynein,BicD and Lis-1 are also involved in anchoring.

Kinesin linkersosk mRNA localization to the posterior pole requireskinesin I, but the identity of the linker or linkersbetween the RNA and motor are unknown, despite the growing list of factors required for its localization

(see above). The C-terminal domain of kinesin I containssix imperfect 34-amino acid tetratricopeptide repeats(TPR). The function of this domain is not clear, but it isprobably involved in protein–protein interactions andpossibly in cargo binding [50,51]. Candidates forproteins that bind to the TPR and could act as motorreceptors include kinectin, but this molecule is notpresent in axons and it has no homologues in flies andworms. Perhaps a better candidate for a linker isamyloid precursor protein (APP), which is known toform a complex with the kinesin light chain TPRdomain. However, kinesin light chain is not requiredfor kinesin heavy chain-dependent cytoplasmicstreaming and posterior localization in Drosophila [60].Another good candidate for a kinesin linker is Sundaydriver (Syd), which is thought to bind the TPR domainof the kinesin light chain [59]. Future work willestablish which of these candidates or indeed otherproteins are the linkers for osk mRNA and any othermRNAs which will undoubtedly be discovered torequire kinesin I or other kinesins for their localization.

It has recently been shown that Rab11, a memberof the largest family of monomeric ras-relatedGTPases, is required for osk mRNA localization [61].Rab11 is thought to be required for the organization of MT plus ends and could define the identity ofposterior membranes required to anchor factorsinvolved in osk mRNA localization and translation.However, Rabs also play a key role in determining thespecificity of membrane transport and fusion ofvesicles, and they have been proposed to be candidatelinkers between membranes and their specificmotors, although additional adapters could existbetween Rabs and the motors [62]. Theseobservations lead to an interesting speculation thatRab11 is a linker between osk mRNA and kinesin I.

Concluding remarks

The study of the mechanism of mRNA localization isnow at an exciting intersection with the field ofcytoskeletal motors and the increasing numbers ofdifferent factors that decorate mRNA during itsbiogenesis in the nucleus and movement in thecytoplasm. New assays in living cells are leading to therapid identification of cis-acting mRNA localizationsignals (zip codes) and the identification of thetransacting factors that provide the specificity neededfor localization. It is likely that many more exampleswill be discovered of mRNAs that are transported bykinesin, other plus end motors, myosin familymembers or different isoforms of cytoplasmic dynein.We also anticipate the wider application ofbiochemical methods for identifying the factorsrequired for localization and the visualization of themovement of endogenous transcripts together withthe factors required for their localization. The next fewyears should lead to a much better understanding ofthe factors that influence which motors mRNAs attachto and how the distinct cytoplasmic destinations ofdifferent mRNAs are determined.

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Acknowledgements

We thank David Tollervey,Andreas Merdes,Nina MacDougall,Alejandra Clark,David Ish-Horowicz and Véronique Van de Borfor their very usefulcomments on themanuscripts, and Adrian Oprins for helpwith the illustrations.

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