c ell p olarity in y east

P1: FKZ/FDB P2: FDS/FGO QC: FDS/anil T1: FDX

September 13, 1999 10:34 Annual Reviews AR092-12

?Annu. Rev. Cell Dev. Biol. 1999. 15:365–91

Copyright c© 1999 by Annual Reviews. All rights reserved

CELL POLARITY IN YEAST

John ChantDepartment of Molecular and Cellular Biology, Harvard University, 7 Divinity Avenue,Cambridge, Massachusetts 02138; e-mail: [email protected].

Key Words actin, microtubules, cytoskeleton, GTPases, cell biology

■ Abstract Subcellular asymmetry, cell polarity, is fundamental to the diversespecialized functions of eukaryotic cells. In yeast, cell polarization is essential todivision and mating. As a result, this highly accessible experimental system servesas a paradigm for deciphering the molecular mechanisms underlying the generationof polarity. Beyond yeast, cell polarity is essential to the partitioning of cell fate inembryonic development, the generation of axons and their guidance during neuronal de-velopment, and the intimate communication between lymphocytes within the immunesystem. The polarization of yeast cells shares many features with that of these morecomplex examples, including regulation by both intrinsic and extrinsic cues, conservedregulatory molecules such as Cdc42 GTPase, and asymmetry of the cytoskeleton as itscenterpiece. This review summarizes the molecular pathways governing the generationof cell polarity in yeast.

CONTENTS

Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 366A Description of the Polarized Yeast Cell. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 366Cell Polarity in Yeast Controls Cell Fate Asymmetrically. . . . . . . . . . . . . . . . . . . 367Two Steps in Polarizing the Cell. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 368

Choosing a Direction for Polarization: Orienting Axes forBudding and Mating. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 368Budding in Cell-Type-Specific Patterns. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 368Mating Toward a Partner. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 375

Building an Axis in the Chosen Direction: Polarity EstablishmentMachinery. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 377Cdc42 GTPase. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 377Other Rho GTPases. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 380Other Factors. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 381

Cytoskeleton and Secretory Machinery. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 382Actin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 382Microtubules . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 383Secretion. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 383

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Further Directions. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 384Closing Remarks. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 385

INTRODUCTION

A Description of the Polarized Yeast Cell

The yeast life cycle has three major phases: cell division by budding, mating be-tween haploid cells, and sporulation of diploids. During budding and mating, cellsare highly polarized. Sporulating cells are not polarized, at least by conventionaldefinitions. There are three cell types in yeast: haploida andα cells and diploida/α cells. Haploid cells can divide vegetatively until they encounter a cell of theopposite mating type, in which case they mate to form ana/α diploid. a/α cellscan also divide vegetatively by budding. Upon starvation conditions they undergomeiosis and sporulation, thus reestablishing the haploid phase.

Figure 1 illustrates the polarized organizations of budding and mating cells.During both processes the overall cellular organization is similar, although bud-ding cells have a neck constriction between mother and bud. The polarized actincytoskeleton consists of two prominent structures: patches and cables. Actin di-rects secretion to the bud or mating projection thereby causing this region to growselectively. Accordingly, the exocytic machinery is highly polarized (Finger &Novick 1998). Actin also contributes to the efficient partitioning of certain or-ganelles, such as mitochondrion, to the daughter (Simon et al 1995, Hermann &Shaw 1998). The relative contributions of actin patches or cables to polarizedgrowth remain unresolved. Patches, thought to be analogous to cortical actin ofhigher cells, cluster around regions of cell surface growth and may serve as dockingsites for vesicles or as endocytosis sites for retrieval of membrane. Remarkably,these actin patches appear to move within the plane of the membrane at rapid rates

Figure 1 The polarized organization of yeast cells while budding and mating.

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?YEAST CELL POLARITY 367

(Doyle & Botstein 1996, Waddle et al 1996). Movement is likely powered bymyosin molecules associated with the patches, rather than by actin treadmilling(Belmont & Drubin 1998). Cables, emanating from the bud site or mating projec-tion tip, probably serve as tracks for myosin-directed movement of vesicles andorganelles to the growth site. In recent years, more attention has focused on thestudy of the actin patches; however, a recent paper highlights the importance ofactin cables (Pruyne et al 1998).

Astral microtubules move the nucleus and orient the spindle in relation to thecellular axis of polarization. A separate set of microtubules forms the spindle.The distinction between spindle and astral microtubules is particularly clear inyeast because the nuclear membrane never breaks down during mitosis. Duringbudding, astral microtubules direct movement of the nucleus to the mother-budneck and then orient the spindle parallel to the mother-bud axis (Carminati &Stearns 1997, Shaw et al 1997). During mating, astral microtubules move thenucleus to the tip of the mating projection in preparation for nuclear fusion, whichfollows cellular fusion (Read et al 1992, Kurihara et al 1994). Astral microtubulesexhibit dynamic instability and appear to be oriented by stabilization at sites onthe cortex (Carminati & Stearns 1997, Shaw et al 1997). Astral microtubules playno role in the targeting of secretion.

In terms of other cytoskeletal elements, yeast cells have no intermediate fila-ments, but they do possess septin filaments, which encircle the mother-bud neckfor the entire cell cycle and serve as a scaffold for cytokinetic activities (Longtineet al 1996, Chant 1996, Frazier et al 1998). Although septins have long beenknown in yeast, recent work has shown that they are found in all eukaryotic cells,where they mediate cytokinesis with possibly broader roles (Kinoshita et al 1997).

Cell Polarity in Yeast Controls Cell Fate Asymmetrically

During development, polarization is a mechanism by which cell fate is segre-gated through the partitioning of RNA or protein determinants. Remarkably, inwild-type haploid yeast, the mother and daughter of each division have distinctcell fates. (Daughter refers to the cell that just formed by budding). If a straincarries a functionalHO gene, the mother cell switches mating type following celldivision, while the daughter cell maintains its identity (Strathern & Herskowitz1979). For many years it was known that the mother cell can switch matingtype because it transcribesHO, a gene encoding an endonuclease that intitiatesswitching, whereas the daughter cell cannot switch becauseHO is silent (Nasmyth1983). The mechanism by which this asymmetry inHO expression is producedremained unsolved for over a decade. We now know that the mRNA encoding atranscriptional repressor (Ash1) of theHO gene is partitioned to the daughter bymyosin-mediated movement of Ash1 ribonucleoprotein particles along polarizedactin cables (Takizawa et al 1997, Long et al 1997, Bertrand et al 1998). Additionalreading concerning this exciting finding can be found in several reviews (Chang& Drubin 1996, Amon 1998).

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?368 CHANT

Two Steps in Polarizing the Cell

The formation of an axis of polarization in yeast occurs in two steps, which unfoldin temporal sequence and can be distinguished genetically.

Step 1: Choosing a direction for polarization. The cell integrates relevant in-formation to choose a direction for polarization. Budding cells use intrinsic spatiallandmarks from previous cell divisions to produce precise patterns of polarizationfrom which bud formation and cell division follow. Mating cells polarize in thedirection of their mating partner as defined by gradients of secreted peptide matingpheromones.

Step 2: Building an axis. The cell builds an axis in the chosen direction asreflected by asymmetry of the cytoskeleton, new cell wall growth, and the positionof secretion and membrane insertion. This step is controlled by Cdc42 and otherRho-type GTPases.

CHOOSING A DIRECTION FOR POLARIZATION:Orienting Axes for Budding and Mating

Budding in Cell-Type-Specific Patterns

Yeast cells divide in two precise spatial patterns (Figure 2a; Freifelder 1960,Hicks et al 1977, Chant & Pringle 1995). Haploida andα cells divide in theaxial pattern in which the mother and daughter cell are constrained to form theirbuds immediately adjacent to the previous site of cell separation. Diploid cellsdivide in a bipolar pattern in which mother and daughter are constrained to bud atthe poles of their ellipsoidal shapes, overlayed with biases toward using a cell’sbirth or distal end for budding events. Figure 2a illustrates these patterns as onewould observe by watching individual cells growing on a solid substrate (upper)or by staining of bud scars, permanent remnants marking each division site on thesurface of mother cells (lower). This second method allows detailed assessmentof a cell’s budding pattern over many generations.

Three classes of genes are involved in producing these patterns. The first classis specifically required for the axial pattern (AXL1, BUD10/AXL2, BUD3, BUD4;Chant & Herskowitz 1991, Fujita et al 1994, Roemer et al 1996, Halme et al 1996).The products of these genes are involved in marking the mother-bud neck duringone cell cycle as a site for budding in the next cell cycle. Additionally,AXL1 isthe immediate target of cell-type control. The second class is specifically requiredfor the bipolar pattern (BUD8, BUD9, and others; Zahner et al, 1996). It is likelythat Bud8 and Bud9 proteins are involved in marking the poles of diploid cellsas sites for budding. The third class is required for both patterns (BUD1/RSR1,BUD2, BUD5; Bender & Pringle 1989, Chant & Herskowitz 1991, Chant et al1991). These gene products likely read the axial or bipolar marks provided by thefirst two classes of gene products and convey this information to the machineryinvolved in building the axis.

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Figure 2 The orientation of axes of polarization during different phases of the yeastlife cycle. (A) Patterns of budding and associated axes of polarization (arrows withincells). Upper. The axial and bipolar patterns of cell division as observed by growingcells on a solid surface.Lower. The patterns of scars on the yeast cell surface resultingfrom the two modes of budding. On each cell a single birth scar marks the pole atwhich the cell was attached to its mother. Bud scars, which are smaller in diameter andmore distinct, mark each site of cell division. Scars can be visualized by staining witha dye (Calcofluor) or by scanning electron microscopy. The patterns of bud scars, inrelation to each other and to the birth scar, precisely document a cell’s budding patternhistory. In the axial pattern, scars form a continuous chain. In the bipolar pattern, scarscluster around the poles. (B) Polarization toward a mating partner. Haploid cells ofopposite mating type will polarize toward each other, elongate, and fuse.

The Mechanism of Choosing Bud Sites in the Axial PatternInitial geneticanalysis ofBUD3, BUD4, AXL1, andBUD10/AXL2showed that these genes werespecifically required for the axial pattern of cell division (Chant & Herskowitz1991, Fujita et al 1994, Roemer et al 1996, Halme et al 1996).AXL1 plays anadditional role in mating pheromone processing, but this role appears entirelyindependent and unrelated to axial budding (Adames et al 1995). Bud3, Bud4, andBud10/Axl2 proteins localize in the mother-bud neck (Figure 3; Chant et al 1995,

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?370 CHANT

Figure 3 The mechanism of axial budding. The spatial and temporal dynamics ofproteins involved in marking the mother-bud neck region for axial budding. Severalproteins, presumably associated in a complex, assemble in the mother-bud neck toform the spatial landmark for the next round of axial budding. See text for details.

Sanders & Herskowitz 1996, Roemer et al 1996, Halme et al 1996). Immediatelyprior to cytokinesis, each of these proteins can be seen as a double ring encirclingthe mother-bud neck (Figure 3, Panel 4). At cytokinesis, this double ring splitsinto two single rings (Figure 3, Panel 4–5). One single ring persists on eachprogeny cell until approximately the time at which a new axis of polarizationforms in axial orientations (Figure 3, Panel 5). Axl1 localization has not beenreported. The genetics and localization data have led to the view that these proteins,marking the mother-bud neck, act as a spatial memory of this position from onecell cycle to the next—the inherited landmark for axial budding (Chant et al 1995,Sanders et al 1996, Roemer et al 1996, Halme et al 1996). The behavior of thisstructure is consistent with the physiological properties of axial budding: Analysisof consecutive budding events has shown that the new bud site forms adjacent tothe last division site with no influence from prior division sites, i.e. the landmarkis shortlived as are the inherited single rings of Bud3, Bud4, and Bud10 (Figure 3,Panels 5–6; Chant & Pringle 1995).

An interesting aspect of the proposed axial budding mechanism is that it iscyclical, with no beginning or end. The axial landmark at the site of cell separation

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directs the next site for bud emergence and mother-bud-neck formation (Figure 3,panels 5, 6). A new landmark forms in the mother-bud neck. Thus progeny cellsinherit a landmark of their previous division site, and this structure directly deter-mines where the next landmark will form.

These results have raised a number of interesting questions. How do theseproteins localize to the neck region? What is the mechanism by which theseproteins signal to direct the next axis of polarization? What regulates usage of thislandmark differentially between haploid and diploid cells?

Localization of Bud3, Bud4, and Bud10Bud3 and Bud4 form double rings en-circling the neck coincident with the initiation of mitosis (Figure 3). Localizationof Bud3 and Bud4 is dependent upon the septin rings, a cytokinetic scaffold thatencircles the mother-bud neck for the entire cell cycle (Chant et al 1995, Sanders& Herskowitz 1996).BUD3andBUD4mRNAs exhibit sharp peaks of abundancein the cell cycle corresponding to the appearance of localized protein products(Sanders & Herskowitz 1996; M Mischke & J Chant unpublished data). It is suffi-cient to postulate that when the mRNAs are translated the soluble products diffuseand bind to the septins already located at the neck. Direct binding to the septinshas not been demonstrated, although it seems likely.

In contrast, Bud10 arrives at the bud site early in the cell cycle and remains inthe mother-bud neck for the remainder of the cell cycle (Figure 3; Roemer et al1996, Halme et al 1996). Bud10, a single-pass transmembrane protein, is deliveredto the surface via the secretory pathway. How Bud10 localizes tightly to certaindomains of the cell surface is not yet understood. Perhaps, Bud10 is delivered tothe cell surface via secretory vesicles specifically at the time of bud emergenceand then remains immobile. Alternatively, Bud10 could be delivered to varioussites on the cell membrane and then laterally diffuse to regions where it is retainedthrough interaction with some other factor.

The Mechanism of Bud3, Bud4, and Bud10 ActionThe mechanism by whichthese proteins communicate with downstream machinery to polarize cellular com-ponents remains unknown. Bud3 and Bud4 are large proteins with uninformativesequences (Chant et al 1995, Sanders & Herskowitz 1996). Bud10 is a single-pass transmembrane protein with a 500–amino acid extracellular domain and a300–amino acid intracellular domain, but Bud10 exhibits no further similaritiesto other proteins in the database (Roemer et al 1996, Halme et al 1996). It istempting to speculate that Bud10 functions in a manner analogous to noncatalyticreceptors, such as T cell receptors or integrins for which tight clustering appears tobe important for sending a signal. To extend this analogy, perhaps the extracellulardomain of Bud10, which is highly glycosylated, serves to anchor the protein inthe cell wall with Bud3 and Bud4 serving to tightly cluster Bud10 to generate apotent signal (Roemer et al 1996, Halme et al 1996). It remains a possibility thata specific extracellular ligand for Bud10 exists, although there is no evidence tosupport this.

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?372 CHANT

How do these proteins send a signal? The most favored view is that the axialspatial cue triggers the local activation of Bud1 GTPase, which in turn activatesCdc42 GTPase (Figure 4a; as described in considerable detail below). Perhaps, theBud10 cytoplasmic tail serves to localize one or more Bud1 regulators—Bud5 GEF(guanine nucleotide exchange factor) (Chant et al 1991) or Bud2 GAP (GTPase-activating protein) (Park et al 1993). Bud1 GTPase, which is closely related toRas (rather than Rho or some other GTPase subfamily), appears to be uniformlylocalized to the plasma membrane with little significant soluble pool (Michelitch& Chant 1996, Park et al 1997). Targeting of Bud1 regulators by Bud10 orassociated proteins would allow localized Bud1 activation by GTP binding orGTP/GDP cycling. Activated Bud1 could then communicate further downsteamvia Cdc24 and Cdc42.

The Mechanism of Choosing Buds in the Bipolar PatternDiploid yeast cellsuse spatial cues for producing the bipolar pattern that are entirely distinct fromthose used in the axial pattern. Physiological experiments have shown that thebipolar landmarks exist at the poles of diploid cells and that they are persistent fornumerous cell cycles (Chant & Pringle 1995).

To date, two genes,BUD8andBUD9, have been reported that have highly spe-cific effects on the bipolar pattern (Zahner et al 1996). Mutations in these genesaffect the bipolar pattern in distinct and interesting ways.bud8mutants are unableto bud at the distal pole (the pole opposite the birth end) and instead produce aunipolar pattern with all buds forming at the proximal pole (the birth pole). Theresulting pattern is distinct from the axial pattern in that bud sites do not occur ina sequential chain as viewed by bud scar staining; rather, they occur as a clusterin the vicinity of the pole in no particular order. The complementary phenotypeis produced bybud9mutants: They are unable to bud at the proximal pole andbud only at the distal pole. Of importance is the fact thatbud8 bud9null mutantsproduce a random pattern in diploids, with the axial pattern of haploids remainingunaffected (JR Pringle, personal communication). Genetically, the properties ofBUD8 andBUD9 are exactly what one would predict for the bipolar landmarks.The subcellular localizations of these proteins have not yet been reported.

The sequences of Bud8 and Bud9 predict related transmembrane proteins with450–500 amino acid extracellular domains, membrane spanning domains, shortcytoplasmic loops, second membrane spanning domains, and short extracellular

−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−→Figure 4 The molecular machinery guiding axes of polarization during budding andmating. (A) The pathway governing axial and bipolar polarization in haploids ordiploids undergoing cell division by budding. (B) The pathway governing chemotropicpolarization during mating. In both panels solid arrows indicate physical interactionbetween proteins or conformational transitions of proteins. Dashed arrows indicatelikely points of regulation where no physical interaction has been demonstrated.

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?374 CHANT

domains (Figure 4a) (JR Pringle, personal communication). The cytoplasmicloops of the two proteins are related in sequence, suggesting that they dock somecommon factor(s), perhaps Bud5 or Bud2, as speculated for the axial landmarkproteins above.

It should be noted that mutations in a large number of genes will shift the bipolarpattern to a random pattern without affecting the axial pattern. These genes includeBni1, Spa2, Rvs161, Rvs167, Sac6, Vrp1(verprolin),Act1(actin-specific alleles),Aip3/Bud6, Pea2, Sec3, Sec4, Sec9, andPho85(Snyder 1989, Sivadon et al 1995,Valtz & Herskowitz 1996, Zahner et al 1996, Finger & Novick 1997, Vaduva et al1997, Yang et al 1997, Tennyson et al 1998). It seems unlikely that all these geneproducts have a direct mechanistic role in bipolar budding. Most, if not all, ofthese genes affect the actin cytoskeleton or secretory pathways. For reasons thatare not clear, it appears that slight perturbations in these pathways abolish bipolarbudding in a seemingly nonspecific manner.

Regulation by Cell Type That haploid and diploid cells polarize and bud in twodifferent spatial patterns provides a paradigm of morphogenetic differentiation.Budding patterns are regulated by cell type rather than ploidy (Hicks et al 1977,Chant & Pringle 1995). Cell type is controlled by transcriptional regulators pro-duced by the mating type loci (MATa andMATα). a/α (MATa/MATα) cells aredifferent froma(MATa) orα (MATα) cells because they produce the transcriptionalcorepressora1-α2 (thea1 subunit encoded byMATa; theα2 subunit encoded byMATα) (Herskowitz et al 1992). For a number of years, it was speculated that thecell-type control in budding pattern was produced by the transcriptional repressionof some gene critical to axial budding bya1-α2 in diploid cells. According to thishypothesis, bipolar budding would occur by default.

Fujita and colleages (1994) identified this gene asAXL1. AXL1is expressedin haploids but not in diploids, where it is repressed bya1-α2. Loss ofAXL1 inhaploids results in the bipolar pattern, while forced expression ofAXL1in diploidsleads to the axial pattern. ThusAXL1 is the single gene differentially regulatedbetween haploids and diploids that is responsible for differences in budding pattern.All other genes specifically required for the axial pattern (BUD3, BUD4, BUD10)are expressed in diploids, and the protein products are present and concentratedin the neck (Chant et al 1995, Roemer et al 1996, Halme et al 1996, Sanders &Herskowitz 1996). Apparently, Axl1 is responsible for this landmark being activespecifically in the haploid cell types.

How Axl1 acts remains a mystery. There are two simple schemes by whichAxl1 may act (Figure 5). The most simple is that Axl1 promotes recognition anduse of the axial landmark in haploid cells (Figure 5a). For instance, Axl1 mightcouple Bud10 to the regulators of Bud1 GTPase (Bud2, Bud5). A second schemeis that an inhibitor of the axial budding landmark exists that prevents its use indiploid cells (Figure 5b). Accordingly, Axl1 would block the inhibitor’s action inhaploid cell types (Fujita et al 1994). Some support is provided for the second

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Figure 5 Cell-type control of budding patterns. (A) A scheme for cell-type controlwhere Axl1 acts positively on axial landmark proteins. (B) An alternate scheme whereAxl1 acts indirectly by blocking an inhibitor of the axial landmarks.

scheme by the existence of mutations,rax1andbud7, reported to have propertiessimilar to those predicted to affect the gene encoding the inhibitor (Fujita et al1994, Zahner et al 1996).

Mating Toward a Partner

Haploid yeast cells are able to redirect their polarization axes in order to mate with apartner (Figure 1b, Figure 2b). Polarization toward a mating partner is chemotropicin that cells of one mating type respond to peptide mating pheromone secreted bythe opposite mating type. Cells can be induced to polarize and grow toward an

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artificially produced gradient of pheromone, demonstrating that pheromone is asufficient chemotropic signal (Segall 1993). Mating pheromones (a andα factor)are sensed by seven transmembrane receptors on each cell type:a cells expressα-factor receptor;α cells expressa-factor receptor. As in other eukaryotic cells,these seven transmembrane receptors are coupled to a heterotrimeric G proteinthat orchestrates cellular responses (Sprague & Thorner 1992, Herskowitz 1995).

Mutations of two types affect polarization during mating. Some mutationsalter the ability of cells to orient their axes chemotropically, i.e. cells carryingsuch mutations form mating projections at wild-type efficiencies but in randomdirections with respect to the pheromone gradient. Mutations of this sort includecertain special alleles ofSTE4(encoding Gβ), FAR1, andCDC24, which encodesthe exchange factor for Cdc42 GTPase (Valtz et al 1995, Nern & Arkowitz 1998,Butty et al 1998). The second class of mutations prevent polarization entirely.Mutations in the genes originally required for budding polarization (i.e.CDC24,CDC42, and associated factors), as well as mutations in several other genes (SPA2,BNI1, PEA2), prevent assembly of an axis (Gehrung & Snyder 1990; Chenevertet al 1992, 1994; Valtz & Herskowitz 1996; Evangelista et al 1997). Genesrequired for orienting axes of budding polarity (BUD andAXLgenes) are largely,if not completely, dispensible for polarization during mating.

Background on Mating Signal Transduction Chemotropic polarization is oneof several well-characterized responses produced by haploid cells in response topheromone. Other responses include transcription, cell cycle arrest, and aggluti-nation, all of which can be elicited for the most part with synthetic pheromone.Much is known about the signal transduction pathways governing these responses(Sprague & Thorner 1992, Herskowitz 1995). Binding of pheromone to the ap-propriate receptor promotes exchange of GTP for GDP on the Gα subunit, leadingto dissociation of the Gβγ heterodimer (Figure 4b). In yeast, Gα has no knownfunction beyond sequestering theβγ subunit. Freeβγ at the plasma membranerecruits and activates Ste20, a PAK kinase, and Ste5, a MAP kinase cascade scaf-fold protein (Whiteway et al 1995, Inouye et al 1997, Pryciak & Huntress 1998,Leeuw et al 1998). These two effectors activate a MAP kinase cascade composedof Ste11 MAP kinase kinase kinase, Ste7 MAP kinase kinase, and Fus3 MAPkinase (Herskowitz 1995). Fus3 phosphorylates Ste12, a transcription factor thatactivates pheromone-inducible genes, and Far1, which binds and inhibits the G1cyclin-dependent kinase complex, to effect cell cycle arrest (Peter & Herskowitz1994). For many years it had been suspected, and now it has been shown, thatmost of this heavily studied signal transduction pathway has no role in orientingaxes of polarization during mating. Remarkably, however, Far1 is a bifunctionalprotein with a second independent role in mating polarization, as described below(Valtz et al 1995, Butty et al 1998).

The Pathway for Orienting Toward a Partner The problem of deciphering thelinks between exogenous pheromones and ultimately cytoskeletal polarization in

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yeast is highly analogous to the problem of chemotaxis in higher eukaryotic sys-tems. Recently, major progress has resulted in our molecular understanding ofmating chemotropism. Far1 protein appears to be the physical link between sig-nal transduction and the Cdc42 module (Figure 4b). A very recent report fromPeter and coworkers (Butty et al 1998) shows that Far1 protein binds directly tothe Gβ protein. Versions of Far1 protein that are unable to interact with Gβ aredefective in orienting mating projections. Likewise, mutant versions of Gβ unableto interact with Far1 result in misoriented mating projections (Nern & Arkowitz1998, Butty et al 1998). Completing the linkage between Gβ and Cdc42, theauthors demonstrate that Far1 binds Cdc24, the Cdc42 GEF (Butty et al 1998;Figure 4b).

The two functions of Far1 protein, cell cycle arrest and mating polarization,are entirely separable (Valtz et al 1995, Butty et al 1998). Cell cycle arrest islikely mediated by a nuclear pool of Far1 protein, whereas mating polarity ismediated by Far1 at the cell cortex (Butty et al 1998). In cells growing vegetatively,prior to pheromone exposure, Far1 is largely, if not exclusively, found in thenucleus. Treatment with pheromone results in relocation of some Far1 protein tothe cytoplasm.

Far1 has no homologs to potentially guide chemotaxis in higher cells. Bindingof Far1 to Gβ occurs through a RING finger domain similar to a domain of Ste5,another target of Gβ (Butty et al 1998). Interestingly, the domain of Cdc24 thatbinds Far1 is conserved in other exchange factors for Rho type GTPases, notablyDbl, an exchange factor for Cdc42 in mammalian cells.

BUILDING AN AXIS IN THE CHOSEN DIRECTION:Polarity Establishment Machinery

Once the cell has integrated spatial cues from budding landmarks or matingpheromone, this information is fed to the polarity establishment machinery, whichis responsible for polarization of the cytoskeleton and cellular components alongthe chosen axis. Prominent among the polarity establishment machinery are Rho-type GTPases, suggesting that cell polarization mechanisms are highly conservedamong eukaryotic cells.

Cdc42 GTPase

Conserved at both the sequence and functional level in all eukaryotes, Cdc42appears to be the central factor in polarizing the cell. Through the exchange factorCdc24, Cdc42 is the convergence point for polarization machinery during buddingand mating (Figure 4).cdc24andcdc42mutants were found as mutants unableto form an axis of polarization during budding (Sloat & Pringle 1978, Adamset al 1990). In contrast to wild-type cells, which target secretion and new cellsurface growth to the bud site or mating projection, these mutants are defective

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in growth targeting (Field & Schekman 1980). As a result, they grow uniformlylarge and round rather than asymmetrically to form a bud or projection (Sloat &Pringle 1978, Adams et al 1990). The localization of Cdc42 cemented its place asa central regulator of polarization. During G1, Cdc42 localizes as a tight corticalpatch at the future bud site or at the future position of the mating projection (Zimanet al 1993). Many proteins localize to the developing growth site, but, in contrastto most secretory machinery and actin-associated proteins, Cdc42 localizes inan actin-independent manner (Ayscough et al 1997). Taken together, these datasuggest that the position of Cdc42 on the cortex defines the axis of cytoskeletalpolarization. These findings have led to two important questions. How does Cdc42become localized? How does Cdc42 exert its effects on the cytoskeleton?

How does Cdc42 Become Localized?The answer to this question is incomplete.The current view is that prior to polarity establishment, Cdc42 is GDP-bound andinactive. Antecedent to bud or mating projection formation, Cdc42 becomes locallyconverted to its GTP-bound form at one position on the cell membrane to definethe point of polarization. Conversion of Cdc42 to its GTP-bound conformationwould be promoted by localized Cdc24 activity in response to Bud1 GTPase duringbudding or in response to Far1 during mating (Figure 4) (Zheng et al 1995, Park et al1997, Butty et al 1998). In concordance with such a scheme, it has been found thatCdc24 is localized to the bud site and to the mating site (Nern & Arkowitz 1999).Ultimately, localized Cdc42 activity would serve as a docking site or activationsite for its effectors.

It is possible that the GTPase activating proteins for Cdc42 are spatially regu-lated. Localized GAP activities might serve to dock Cdc42 and promote localizedrelease of Cdc42 effectors, or GAP activities might be inhibited at the bud or mat-ing site allowing build up of the GTP-bound form of Cdc42. Two Cdc42 GAPs areknown: Rga1/Dbm1 and Bem3, with possibly a third, Rga2 (Stevenson et al 1995,Chen et al 1996, Zheng et al 1994).rga1 mutants are morphologically aberrantin a number of ways, but unlikecdc42nulls, rga1 nulls are viable (Stevensonet al 1995). bem3single mutants exhibit no obvious phenotype, and nothing ispublished concerningRGA2(Zheng et al 1994). Schemes can also be imaginedin which both types of regulators are controlled spatially to produce positionalprecision in Cdc42 activity.

How Cdc42 is brought to the membrane is not understood. It is likely thatCdc42-GDP is soluble, with GTP-binding conferring affinity for the membrane.In other systems, it has been suggested that Rho class GTPases are sequestered inthe cytoplasm by guanine nucleotide dissociation inhibitor (GDI) proteins. Search-ing the yeast genome predicts only one Rho-type GDI protein (Rdi1). Despite thefact that this GDI interacts biochemically with both Cdc42 and Rho1, disruptionof the gene encoding this GDI (RDI1) produces no apparent phenoytpe (Masudaet al 1994, Koch et al 1997). In the GDI-deficient cells, Cdc42 can still be detectedin cytoplasmic fractions. Thus how Cdc42 remains soluble in the absence of Rdi1and the function of Rdi1 remain uncertain.

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How does Cdc42 Control the Cytoskeleton?Elucidation of the pathways con-trolled by Cdc42 to regulate cytoskeletal polarization is a major unsolved problemof cell biology in both yeast and higher cells. Four effectors of Cdc42 are known todate (Ste20, Cla4, Gic1, and Gic2), with Bni1 representing a possible fifth effectoraccording to one report (Cvrckova et al 1995, Brown et al 1997, Chen et al 1997,Evangelista et al 1997). However, evidence is perhaps more strongly in favor ofthe view that Bni1 is an effector of Rho1 (Kohno et al 1996), therefore it is treatedas such below.

Ste20 and Cla4 are members of the PAK kinase family, with Cdc42/Rac inter-active binding (CRIB) domains through which they bind to Cdc42. Currently, theextent of their contributions to cell polarization and cytoskeletal rearrangementsis unresolved and controversial. Genetic studies suggest that these related kinaseshave some overlapping as well as distinct roles (Cvrckova et al 1995). For example,Ste20 is involved in mating signal transduction, whereas Cla4 is not. Conversely,cla4 mutants have a defect in cell shape and cytokinesis, whereasste20mutantshave no detectable vegetative defect. However,ste20 cla4mutants are inviable;they arrest as unbudded cells with actin appearing to be polarized (Cvrckova et al1995). Based on these data, one can conclude that these PAK kinases are notrequired for Cdc42 to polarize the actin cytoskeleton. This interpretation concurswith experiments in mammalian cells where Cdc42 molecules unable to bind PAKkinases are fully able to induce cytoskeletal rearrangements when injected intofibroblasts (Lamarche et al 1996, Joneson et al 1996). However, conflicting opin-ions can be found in the literature (Sells et al 1997, Eby et al 1998). Even if PAKkinases do not affect actin polarization, they likely have a morphogenetic role indirecting septin assembly or secretion to the bud site.

Gic1 and Gic2 are Cdc42 effectors that contribute to cytoskeletal polarization(Brown et al 1997, Chen et al 1997). These two proteins were identified by scan-ning the entire yeast genome for anonymous open reading frames with the CRIBmotif (Burbelo et al 1995). Remarkably, the presence of this 8-amino acid coremotif was an accurate predictor of proteins that bind Cdc42 specifically, and Gicfunction is dependent upon an intact CRIB motif (Brown et al 1997, Chen et al1997). Double mutants have defects in actin polarization that resemble those ofcdc42mutants but are considerably less severe (Brown et al 1997, Chen et al1997). The mildness of thegic1gic2phenotype, combined with the lack of identi-fiable homologs in other eukaryotes, suggests that other important Cdc42 effectorsremain to be discovered.

When comparing yeast to other eukaryotic systems, it is interesting that someproteins that are direct effectors of Cdc42 in mammalian cells have counter-parts in yeast, which apparently do not bind Cdc42. In mammalian cells, WASP(Wiskott-Aldrich Syndrome Protein) and its isoforms are effectors of Cdc42,which contribute to actin rearrangements (Kirchhausen & Rosen 1996, Mikiet al 1998). In yeast, the WASP-related protein Las17/Bee1 has a role modu-lating actin dynamics, but it has little or no affinity for Cdc42 (Li 1997, Karpovaet al 1998). Perhaps the systems have diverged such that the exact linkages of

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protein networks have changed but the basic functions of proteins have remainedconstant.

Similarly, mammalian IQGAP proteins bind Cdc42 avidly and almost certainlyare Cdc42 effectors, but one is hard pressed or unable to demonstrate interactionbetween yeast Cdc42 and the IQGAP-related protein Iqg1/Cyk1 (Epp & Chant1997, Lippincott & Li 1998, Osman & Cerione 1998). Iqg1 appears to be involvedin cytokinesis rather than the establishment of cell polarity. Iqg1 directs actin ringformation and other cytokinetic activities to the mother-bud neck toward the endof the cell cycle. In higher cells, the roles of IQGAP proteins remain unknown.Perhaps they also play a role in cytokinesis, and their intimate association withCdc42 reflects a role for Cdc42 in cytokinesis (Drechsel et al 1997), a role possiblynot preserved in yeast.

Other Rho GTPases

Four other Rho class GTPases in yeast have been studied with a fifth protein of thisclass predicted in the database. Genetic data place the four studied Rho proteinsinto two pairs related by function: Rho1 and Rho2, Rho3 and Rho4.

Rho1 and Rho2 Rho1 is thought to maintain polarization of the actin cytoskele-ton and cell wall biosynthetic activities during bud growth, with Rho2 playing asimilar but minor role. Rho1 mutants arrest growth as small-budded cells and lyse(Yamochi et al 1994). Rho1 protein is localized to the bud site and then to thetip of the growing bud (Yamochi et al 1994). The pathway immediately upstreamof Rho1 is well understood. Rho1 GEFs, Rom1 and Rom2, are localized to thebud site where presumably they recruit Rho1 from a soluble pool (Ozaki et al1996, Manning et al 1997). Rom2 activity is dependent upon the lipid kinase Tor2(Schmidt et al 1997). The activity of Rom2 requires its PH domain, suggesting theattractive hypothesis that Tor2 lipid kinase produces a phospholipid that activatesRom2, thereby activating Rho1. (Rom1 is likely regulated in a similar fashionby analogy to Rom2.) How Tor2 lipid kinase is activated remains unknown, butperhaps Cdc42 directly or indirectly plays a role. Notably, many GEF proteinshave PH domains immediately adjacent to their GEF catalytic domain, suggestingthat regulation by phosphoinositides is widespread.

Rho1 has three well-defined effectors, glucan synthase, protein kinase C (Pkc1),and the formin Bni1 (Nonaka et al 1995, Qatoda et al 1996, Drgnova et al 1996,Kohno et al 1996, Kamada et al 1996). Direct linkage between Rho1 and the cellwall biosynthetic activity, glucan synthase, makes perfect sense both geneticallyand physiologically: Lysis ofrho1 mutants is readily explained by a deficiencyin glucan synthase activity, and the polarization of cell wall biosynthetic activitiesis exactly what one would predict, given that yeast cells grow in such a highlyasymmetric fashion (Qatoda et al 1996, Drgnova et al 1996).

How Rho1 affects the actin cytoskeleton is less clear. Two Rho1 effectors,Bni1 and Pkc1, have been speculated to control actin. The role of Bni1 as an actin

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regulator was intitially buttressed by its interactions with several actin binding pro-teins, profilin, tropomyosins, and Bud6/Aip3 (Kohno et al 1996, Evangelista et al1997). As provocative as such interactions may be, a recent report argues stronglythat Pkc1 mediates the effect of Rho1 upon the actin cytoskeleton (Helliwell et al1998). Pkc1 activates a MAP kinase pathway made up of Bck1 (MAP kinasekinase kinase), Mkk1,2 (MAP kinase kinases), and Mpk1/Slt2 (MAP kinase).Whether PKC controls actin via this pathway or an independent pathway remainsunknown.

Rho3 and Rho4 Rho3 and Rho4 appear to be involved in the establishment ormaintenance of polarity and overlap in function (Matsui & Toh-e 1993, Imai et al1996). However, these proteins are less well studied than Cdc42 and Rho1. Takaiand coworkers have made a strong case for Bnr1, the second yeast formin, as aRho3 effector (Bnr1: Bni1 related; Imamura et al 1997). The formin family isdefined by two domains, the formin homology 1 domain (a proline-rich domain)and the formin homology 2 domain. In other systems such asDrosophilaandfission yeast, these proteins are involved in cytokinesis. Even though the twoformins appear to act as effectors for different GTPases, genetic analyses suggestthat they overlap in function (Imamura et al 1997). Singlebni1 or bnr1 mutantsare phenotypically mild; however, double mutants have a defect in cell shape withassociated disorganization in the actin cytoskeleton. Interactions of Rho3 with aclass V myosin (Myo2) and an exocyst component (Exo80) have recently beenreported (Robinson et al 1999). While the role of these interactions in vivo remainto be fully explored, such a tangible connection linking Rho type GTPases to thecytoskeleton and secretory machinery could prove a major breakthrough.

Other Factors

Several additional factors have effects on polarity establishment, but their functionsare not easily placed in the pathways defined above.

Bem1, a protein with two SH3 domains, binds to Cdc24, Cdc42, Bud1, andFar1 (Chenevert et al 1992, Zheng et al 1995, Park et al 1997, Butty et al 1998).bem1mutants are mildly deficient in polarity establishment during budding, witha more severe phenotype at elevated temperature, andbem1mutants are unableto form mating projections (Chenevert et al 1992). Two ligands of the Bem1SH3 domains are known: the related proteins Boi1 and Boi2 (Bender et al 1996,Matsui et al 1996).boi1 boi2double mutants have moderate polarization defects.Given the multiple interactions of Bem1 with other polarity establishment proteinsand the effect ofbem1mutations on polarization, it is speculated that the Bem1protein acts as a scaffold for all of this machinery to facilitate the efficiency of theprocess.

BEM4was identified in screens surroundingRHO1andCDC42. bem4mutantshave a mild defect in cell polarization (Hirano et al 1996, Mack et al 1996). Bem4binds several of the Rho class GTPases including Cdc42, Rho1, and Rho3. The

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lack of specificity in Bem4-GTPase interactions has led to speculation that Bem4acts as a Rho-type GTPase escort or sequestration protein with a role analogousto that suggested for GDIs.

CYTOSKELETON AND SECRETORY MACHINERY

Ultimately, the regulatory pathways described above must connect with the cy-toskeleton and secretory machinery; however, proven links remain scarce. Whiledetailed coverage of the cytoskeleton and secretory machinery is beyond the scopeof this review, it is worth touching upon the likely entry points for regulation.

Actin

The ARP (actin-related protein) complex, an actin nucleation activity present in alleukaryotes, is undoubtedly an important target of polarity establishment machinery(Machesky et al 1994). ARP components are required for viability in yeast, andthey appear largely associated with actin patches (Moreau et al 1996, Winteret al 1997). Whether a small amount of ARP complex is also at the bud site tonucleate actin cables is not known. In reconstitution studies withXenopusextracts(Ma et al 1998b), the ARP complex has been defined as an important indirecttarget of Cdc42. Given the recent finding that WASP-type molecules interact withthe ARP complex, the Cdc42 to ARP link is potentially completed in higher cells(Machesky & Insall 1998) and, in fact, several groups have shown that Cdc42-GTP can stimulate actin polymerization in vitro when provided with WASP andthe ARP complex (Machesky et al 1999, Yarar et al 1999, Rohatgi et al 1999). Inyeast, Cdc42 almost certainly does not interact directly with the WASP equivalent,Las17/Bee1 (Li 1996). Even if Las17 is controlled via an intermediary, geneticstudies suggest that the Cdc42 to ARP pathway cannot be so simple.cdc42nullsare lethal, as are nulls in genes encoding Arp complex subunits; however,las17nulls are viable and grow quite well (Li 1997, Karpova et al 1998). The simplestinterpretation is that Cdc42 or another Rho protein controls the ARP complexthrough a Las17-independent pathway.

The ultimate regulation of actin polarization should prove complex. In additionto nucleation, other actin-regulatory activities are undoubtedly controlled in apolarized fashion, including bundling, severing, capping, and depolymerization.The recent work of Pruyne et al (1998) highlights yeast tropomyosins as importantregulators of actin, as well. Finally, several lines of evidence suggest that myosinscontrol the geometry of the actin cytoskeleton, rather than simply responding toit: Myo2 is reported to be a direct Rho3 target (Robinson et al 1999); Myo2 canlocalize independently of actin (Ayscough et al 1997); and actin patch motility islikely powered by myosins (rather than actin treadmilling; Lappalainen & Drubin1997).

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Microtubules

Astral microtubules move the nucleus and orient the spindle within the cell. Or-ganization of astral microtubules must be regulated in a polarized fashion. Suchregulation probably occurs by selective stabilization of dynamic microtubules atspecific locations. Several studies of microtubule organization suggest that, duringbudding, first an astral microtubule is stabilized at the mother-bud neck to pull thenucleus to the neck and then astral microtubules are selectively stabilized at themother and bud cortexes to align the spindle (Carminati & Stearns 1997, Shawet al 1997). Likewise, astral microtubules are stabilized at the tip of the matingprojection. Evidence suggests that microtubule motors act upon these connec-tions to move the nucleus and to align the spindle (Stearns 1997 and referencestherein). Little is known about the molecular nature of the cortical microtubuleattachments. One known protein, Kar9, probably has an important role in micro-tubule attachment at the cortex.kar9 mutants are deficient in spindle orientation,and the Kar9 protein can be seen localized as a spot at the tip of the bud or matingprojection, positions where microtubules are selectively stabilized (Miller & Rose1998). Astral microtubules appear to be stabilized at other locations where Kar9is not present. Therefore, other proteins that affect microtubule interactions withthe cortex or bud neck are presumably present in these locations. Whether Kar9interacts with the plus ends of microtubules directly is not known. Ultimately, thespatial regulation of Kar9, other stabilization proteins, and the microtubule motorsmust be under the control of the polarity establishment machinery. Evidence sug-gests that the polarity establishment machinery controls microtubule organizationindirectly through its effects on actin (Palmer et al 1992). An equally tenable andlargely unexplored possibility is that Cdc42 or one of the Rho proteins controls amicrotubule stabilization pathway more directly.

Secretion

In recent years, much of the machinery involved in bringing secretory vesicles fromthe Golgi to the cell surface has been clarified by Novick and coworkers (Terbushet al 1996, Guo et al 1999). A complex of proteins—the exocyst—in concert withSec4, a Rab GTPase, is largely responsible for the docking of secretory vesicles atthe bud site. Accumulation of vesicles at growth sites exploits at least two targetingmechanisms that likely work additively to make the process highly efficient. First,a peripheral component of the exocyst complex, Sec3, is tightly localized to siteson the plasma membrane where vesicles are fused (Finger et al 1998). Sec3is potentially the membrane docking factor for vesicles. The means by whichSec3 becomes localized to specific sites on the cell cortex is not understood, butsome evidence has been presented raising the possibility that Sec3 localization isindependent of Cdc42—a possibility that demands further investigation. Second,it appears that myosin motors, translocating on actin cables, direct at least some

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types of secretory vesicles to the vicinity of the budding or mating site (Govindanet al 1995, Finger & Novick 1998, Pruyne et al 1998).

FURTHER DIRECTIONS

As reflected in this review, tremendous progress has been made in painting thebroad outlines of the molecular mechanisms used by yeast cells to polarize. Withinthe next five years the genomic information and newest methods available shouldprovide a precise and accurate view of the molecules and pathways employed.Currently, conspicuous gaps exist in our knowledge. In particular, the followingquestions remain unanswered. How are axial or bipolar landmarks linked to theBud1-GTPase module that controls Cdc42? What is the functional linkage be-tween Cdc42 and the other Rho GTPases? Does Cdc42 control the activities ofthese other GTPases in a cascade analogous to the Cdc42-Rac-Rho cascade of somemammalian cells? How do Cdc42 and the other Rho-type GTPases communicatewith the cytoskeleton? In the case of Cdc42 it appears that as-yet-unknown effec-tors shall prove important. Much work remains in defining the linkages betweenthis regulatory machinery and the cytoskeleton in mechanistic detail.

In terms of defining linkages in these pathways, genetic and two-hybrid in-teractions need to be buttressed with biochemical work. Many proteins in thesepathways can be shown to be constituents of large complexes and display multipleinteractions with other players. Which complexes and interactions are important?Interactions must be disrupted in vivo through point mutation to assess their im-portance. As an illustration of this point, two papers show that, for many of thefunctions of Ste20 PAK kinase, surprisingly, the interaction with Cdc42 is entirelydispensible (Peter et al 1996, Leberer et al 1997). Other work has shown thatGic proteins are completely dependent upon Cdc42 interaction for their activities(Brown et al 1997, Chen et al 1997).

Another important area for future studies is real-time analysis of protein dynam-ics in wild-type and mutant situations. With the popularization of green fluorescentprotein, these studies are beginning to occur. For example, one could study thedynamics of Cdc42 localization at the bud or mating site. Then one could examinethe alterations in dynamics when GAP proteins are absent or when the upstreamBud1 module is missing. Quantitative data in this regard would help mold bio-chemical, genetic, and localization data into coherent models describing the invivo workings of these pathways.

Finally, interesting areas remain entirely unexplored, such as how all of thismachinery acts cooperatively to generate a single axis of polarization that resultsin a single bud or single mating projection. Even in cases where axes are ori-ented seemingly at random (e.g. in abud1mutant or in uniform fields of matingpheromone), a single coherent axis forms with essentially the same kinetics asin wild-type cells. Remarkably few if any mutants are known that produce more

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than one axis simultaneously. What is the nature of this cooperativity in molecularterms? This cooperativity problem is general rather than yeast specific. For exam-ple, treatment of fibroblasts with certain growth factors leads to the extension oflamellaepodia; discreet lamellaepodia form even in the presence of uniform fieldsof growth factor. As another example, many chemotactic mammalian cells main-tain a polarized motile morphology in culture even when no localized exogenoussignals are provided.

CLOSING REMARKS

Remarkable progress has been made in deciphering the molecular mechanismsof cell polarization in yeast. The broad outlines of the pathways and the generalprinciples observed will undoubtedly be conserved across the eukaryotic kingdom.In detail, some of the machinery is yeast specific, such as the bud site landmarkproteins, whereas other machinery is highly conserved in function, such as Cdc42controlling axis formation or G protein-coupled receptors guiding chemotropismor chemotaxis. Yeast has always represented the forefront of eukaryotes in termsof experimental accessibility. Completion of the genome and the development ofnovel tools, such as GFP or extremely facile gene replacement, have acceleratedthe pace of yeast research several-fold in the last few years. When a sequel tothis review is written in five years, the landscape will have changed markedly,accompanied by more detail and a deeper understanding.

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Adams AEM, Johnson DI, Longnecker RM,Sloat BF, Pringle JR. 1990.CDC42 andCDC43, two additional genes involved inbudding and the establishment of cell polar-ity in the yeastSaccharomyces cerevisiae. J.Cell Biol. 111:131–42

Amon A. 1998. Controlling cell cycle andcell fate: common strategies in prokaryotesand eukaryotes.Proc. Natl. Acad. Sci. USA95:85–86

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Annual Review of Cell and Developmental Biology Volume 15, 1999

CONTENTSVacuolar Import of Proteins and Organelles from the Cytoplasm, Daniel J. Klionsky, Yoshinori Ohsumi 1

Blue-Light Photoreceptors In Higher Plants, Winslow R. Briggs, Eva Huala 33

Cooperation Between Microtubule- and Actin-Based Motor Proteins, Susan S. Brown 63

Molecular Mechanisms of Neural Crest Formation, Carole LaBonne, Marianne Bronner-Fraser 81

Lymphocyte Survival--The Struggle Against Death, Robert H. Arch, Craig B. Thompson 113

The Road Less Traveled: Emerging Principles of Kinesin Motor Utilization, Lawrence S. B. Goldstein, Alastair Valentine Philp 141

Proteins of the ADF/Cofilin Family: Essential Regulators of Actin Dynamics, James R. Bamburg 185

Visual Transduction in Drosophila, Craig Montell 231

Biochemical Pathways of Caspases Activation During Apoptosis, Imawati Budihardjo, Holt Oliver, Michael Lutter, Xu Luo, Xiaodong Wang

269

Regulation of Nuclear Localization: A Key to a Door, Arie Kaffman, Erin K. O'Shea 291

Actin-Related Proteins, D. A. Schafer, T. A. Schroer 341

Cell Polarity in Yeast, John Chant 365

Vertebrate Endoderm Development, James M. Wells, Douglas A. Melton 393

Neural Induction, Daniel C. Weinstein, Ali Hemmati-Brivanlou 411

SCF and CDC53/Cullin-Based Ubiquitin Ligases, R. J. Deshaies 435Integration of Signaling Networks that Regulate Dictyostelium Differentiation, Laurence Aubry, Richard Firtel 469

When to Switch to Flowering, Gordon G. Simpson, Anthony R. Gendall, Caroline Dean 519

Regulation of Mammalian O2 Homeostasis by Hypoxia-Inducible Factor 1, Gregg L. Semenza 551

Mechanisms of Viral Interference with MHC Class I Antigen Processing and Presentation, Jonathan W. Yewdell, Jack R. Bennink 579

Transport Between the Cell Nucleus and the Cytoplasm, Dirk Görlich, Ulrike Kutay 607

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[PSI+]: An Epigenetic Modulator of Translation Termination Efficiency, Tricia R. Serio, Susan L. Lindquist 661

Adaptors for Clathrin-Mediated Traffic, Tomas Kirchhausen 705

Synaptic Vesicle Biogenesis, Matthew J. Hannah, Anne A. Schmidt, Wieland B. Huttner 733

The Translocon: A Dynamic Gateway at the ER Membrane, Arthur E. Johnson, Michael A. van Waes 799

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