Asymmetric localization and function of cell-fate determinants: a fly’s view Allison J Bardin, Roland Le Borgne and Franc¸ois Schweisguth1 One mechanism to generate daughter cells with distinct fates is the asymmetric inheritance of regulatory proteins, leading to differential gene regulation in the daughter cells. This mode of cell division is termed ‘asymmetric cell division.’ The nervous system of the fly employs asymmetric cell division, both in the central nervous system, to generate neural precursors, neurons and glial cells; and in the peripheral nervous system, to create sensory organs that are composed of multiple cell types. These cell lineages are excellent models to examine the gene expression program that leads to fate acquisition, the cell-fate determinants that control these programs and how these determinants, in turn, are distributed through cell polarity machinery. Addresses Ecole Normale Supe´rieure, CNRS UMR 8542, 46, rue d’Ulm, 75230 Paris Cedex 05, France 1 e-mail: [email protected]

Current Opinion in Neurobiology 2004, 14:6–14 This review comes from a themed issue on Development Edited by Christopher A Walsh and Barry Dickson 0959-4388/$ – see front matter ß 2003 Elsevier Ltd. All rights reserved. DOI 10.1016/j.conb.2003.12.002

Abbreviations CNS central nervous system Dlg discs-large protein GFP green fluorescent protein GMC ganglion mother cell Lgl lethal(2) giant larvae protein MRLC myosin regulatory light chain N Notch NB neuroblast NICD Notch intracellular domain PDZ PSD95/Discs large/ZO1 domain PNS peripheral nervous system SOP sensory organ precursor

Introduction Diverse organisms, from bacteria to humans, have evolved means by which to create multiple cell types through a mitotic cell division. One method of generating sister cells with distinct identities is the asymmetric segregation of cell fate determinants, resulting in an ‘asymmetric cell division’ [1]. The nervous system of the fruit fly has been an invaluable model for understanding the mechanisms underlying asymmetric cell division. This review focuses on the mechanisms by Current Opinion in Neurobiology 2004, 14:6–14

which cell fates are acquired within the neuroblast lineage of the central nervous system (CNS), as well as the sensory organ precursor lineage of the peripheral nervous system (PNS) in Drosophila melanogaster. In addition, the possible function of vertebrate homologs of cell fate determinants in flies is discussed. Within the nervous system of D. melanogaster, the neuroblast (NB) and sensory organ precursors (SOP) undergo asymmetric cell division. Embryonic neuroblasts (NBs) delaminate basally from the surface epithelium and then undergo an asymmetric cell division, giving rise to a smaller, basal ganglion mother cell (GMC) and a larger, apical NB. NBs continue to divide in this manner, while each GMC divides once, asymmetrically, to form two different daughter neurons (or two glial cells, in some lineages). Thus, both NB and its daughter, the GMC, undergo an asymmetric cell division (Figure 1a). The adult bristle sensory organs are also produced by asymmetric cell divisions (Figure 1b). The sensory organ precursor (SOP), or pI cell, divides within the plane of the epithelium, yielding a posterior pIIa cell and an anterior pIIb cell. The pIIa cell divides to form the posterior socket cell and the anterior hair cell, whereas, the pIIb cell produces the neuron, the sheath cell and a glial cell that is eliminated via apoptosis (Figure 1b) [2–4]. The ability to create distinct daughter cells is crucial, both for maintaining the stem-cell-like identity of the NB in the NB lineage and for generating a complete sensory organ, composed of multiple cell types that are derived from the SOP. The generation of daughter cells with different cell fates involves three levels of control (Figure 1c): (i) regulated programs of gene expression, controlling cell fate; (ii) unequally-segregated fate determinants that, in turn, regulate programs of gene expression in only one of the two daughter cells; (iii) cell polarity machinery that coordinates asymmetric localization of fate determinants, with respect to the cell division plane. This review discusses current knowledge about each of these three processes in both the NB and SOP asymmetric cell divisions in Drosophila. We also review current knowledge of possible asymmetric cell divisions within the developing CNS in vertebrates. The large amount of data concerning the homologous cell polarity machinery in Caenorhabditis elegans is not discussed, as it is beyond the scope of this review (for a review, see [5–7]).

Specifying distinct daughter cell fates Several mechanisms can enable one daughter cell to activate a program of gene expression that is required www.sciencedirect.com

Asymmetric localization and function of cell-fate determinants Bardin, Le Borgne and Schweisguth 7

Figure 1

(a)

(b) pI

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Current Opinion in Neurobiology

The asymmetric cell divisions and their control. (a) The neuroblast lineage. The neuroblast (NB) divides in a stem-cell-like fashion, to give rise to an NB and ganglion mother cell (GMC), which then divides to generate two distinct neurons. The fate determinant Prospero (green) asymmetrically segregates to the GMC during the NB division. (b) The adult sensory organ precursor lineage. The pI (primary precursor or SOP) divides, giving rise to the pIIa and pIIb cells, which are differentiated by inheritance of Numb and Neur (both shown in green). The indicated divisions result in the adult bristle, consisting of the socket (so), shaft (sf), sheath (st), and neuron (n). The glial cell is eliminated by apoptosis. Numb and Neur are thought to bias all resulting cell-fate decisions. (c) Creating daughter cells with different cell fates. Panel one depicts the asymmetric activation and/or repression of cell-fate target genes. Panel two illustrates one mechanism for this, through the asymmetric segregation of a regulatory protein (green), during the previous mother cell division. In panel three, the mother cell establishes cortical domains, which guide the regulatory protein to one of the two daughter cells.

for a particular cell fate, while the other daughter cell does not. The NB division accomplishes this through asymmetric inheritance of the transcription factor Prospero (Pros) (Figure 1a). Division of the SOP, however, uses a different mechanism that enables the activation of the transcription factor Suppressor of Hairless (Su(H)) in one daughter cell and the translational repression of the transcription factor Tramtrack69 in the other daughter cell. Asymmetric transcription factor segregation

Pros is a homeodomain transcription factor that accumulates in the nuclei of GMC, but not NB. In pros mutant embryos, GMCs fail to express normal GMC marker proteins (such as Even-skipped) and their progeny neurons display axonal defects [8–12], indicating that Pros www.sciencedirect.com

regulates the GMC-specific program of gene expression. Differential expression of target genes is controlled through the asymmetric segregation of Pros to daughter cells during NB division [10–12]; however, the direct transcriptional targets of Pros that are required for GMC cell fate are not well characterized. Asymmetric transcription complex assembly

Cell fate within the SOP lineage is, primarily, regulated through the asymmetric control of transcriptional complexes, bound to DNA via Su(H). Su(H) is a DNAbinding protein that associates with either co-repressor or co-activator complexes. The processed form of the transmembrane receptor Notch (N) is a member of a potent co-activator complex for Su(H). Following the SOP division, N is activated in the posterior cell by Current Opinion in Neurobiology 2004, 14:6–14

8 Development

Figure 2

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specific proteolysis of Ttk69 in the pIIb also plays a role in generating asymmetry [19]. How N signaling inactivates Musashi and what cell fate-specific target genes respond to Ttk69 are not known.

Neuralized Delta Sanpodo

Having discussed the downstream events allowing binary cell fate decisions through asymmetric control of transcriptional programs, we now focus on the upstream events that generate the asymmetry. In the NB division, localization of Pros into GMCs is dependent on unequal segregation of adaptor molecules. In the SOP, cell-to-cell N signaling is directionally biased by regulatory proteins that either inhibit (Numb) or promote (Neuralized) N signaling. Prospero and its regulation in the NB division

Current Opinion in Neurobiology

A model of the Notch pathway and its regulators in asymmetric cell division. This is a compilation of data from the SOP and the MP2 NB lineages. In the signal-sending cell, Neur promotes Delta (Dl) endocytosis, which is important for activation of Notch (N) in the signal-receiving cell [43,46]. N activity leads to Su(H)-dependent transcription, important for cell fate [14–16]. Numb, inherited by the signal-sending cell, inhibits cell surface localization of Sanpodo [27,40]. In the signal-receiving cell, Sanpodo promotes N activation at the cell surface [40]. Numb also promotes localization of a-adaptin to the signal-sending cell, where it inhibits N signalling [37]. It is unclear whether a-adaptin functions by promoting N endocytosis or whether it blocks Sanpodo surface accumulation.

the transmembrane ligands Delta (Dl) and Serrate (Ser). This is thought to result in the release of the N IntraCellular Domain (NICD; reviewed in [13]) in the pIIa cell, thereby leading to the formation of a transcriptional activator complex specifying the pIIa fate (Figure 2) [14–16]. The direct Su(H) targets that are repressed in the pIIb cell and activated (or derepressed) in the pIIa cell remain to be identified.

Asymmetric segregation of Pros is the result of two redundant mechanisms: pros mRNA localization through the Staufen RNA binding protein [20,21], and localization of Pros protein through its anchor protein, Miranda [10–12,22,23]. In contrast to Pros protein localization, that of pros mRNA does not play an essential role in cell fate determination, except when Pros protein is limiting [20,21]. In addition to localizing Pros protein, Miranda regulates pros mRNA localization by anchoring Staufen to the basal cortex of the dividing NB [24–26]. Null alleles of miranda result in symmetric segregation of Pros and pros mRNA into daughter cells and, therefore, inappropriate GMC cell-fate specification [22,25]. Regulation of Su(H)-dependent transcription in the SOP

In the SOP lineage, activation of N switches on Su(H)dependent transcriptional programs. How, then, does Nsignaling occur in one daughter cell but not in the other? To date, two proteins are known to enable ‘directional’ N-signaling, ensuring that it occurs only in the pIIa cell and not in the pIIb cell : Numb and Neuralized. Numb

Asymmetric translational repressor activity

An additional level of cell fate regulation in the SOP division comes from the asymmetric translational repression of the ‘tramtrack’ (ttk69) mRNA that encodes a transcriptional repressor. Loss of ttk69 activity leads to a pIIa-to-pIIb cell fate transfomation, suggesting that ttk69 is important for specifying the pIIa cell [17]. Ttk69 protein is present in the pIIa cell but not in the pIIb cell, although mRNA levels appear to be equal in the two cells [18]. Asymmetry is generated when a translational repressor protein, Musashi, binds to and inhibits the translation of the ttk69 mRNA, specifically, in the pIIb cell. Musashi protein is present in both pIIa and pIIb, however, its ability to inhibit ttk69 translation is blocked in pIIa cells by an N signaling-dependent mechanism that is not well understood [18]. Additionally, it seems that Current Opinion in Neurobiology 2004, 14:6–14

Loss of numb function results in transformation of the pIIb cell into a second pIIa cell. During the pI division, Numb protein segregates preferentially to the anterior daughter cell, which, therefore, adopts the pIIb fate [27]. In the embryo, Numb localization is, in part, dependent on an interacting protein; ‘Partner of Numb’ (Pon) [28]. Numb inhibits N receptor signaling in the anterior daughter cell, whereas NICD activates Su(H) in the posterior daughter cell [29–31]. Numb also segregates in an asymmetric manner in the NB lineage [27,32]; however, within most NB lineages, loss of numb function does not effect the GMC fate, but plays a role in specifiying neuron identity at the GMC division [27,33–35]. An exception to this is the MP2 NB lineage, which undergoes a single, terminal division that is regulated by Numb and N signaling [30,32]. www.sciencedirect.com

Asymmetric localization and function of cell-fate determinants Bardin, Le Borgne and Schweisguth 9

Recent work has begun to elucidate the way that Numb impinges on N signaling. Berdnik et al. [36,37] have found that the a-adaptin, a protein involved in endocytosis, associates with Numb and is asymmetrically segregated to the pIIb cell in a numb-dependent manner (Figure 2). Given the ability of Numb to associate with the NICD [31,38,39], Numb may downregulate the N receptor through a-adaptin-dependent endocytosis (Figure 2) [37]. This model predicts that Notch would be endocytosed specifically in the pIIb cell; but this has not yet been demonstrated. Recently, a second model has been proposed for Numb function. Sanpodo (Spdo) is a transmembrane protein that is specifically expressed in asymmetrically dividing cells and co-immunoprecipitates with Numb and Notch. Genetic analysis indicates that spdo is required for N signaling, generating two neurons during GMC cell divisions. spdo acts downstream of Delta and antagonistically to numb. Strikingly, the subcellular localization of Spdo appears to be regulated by Numb. Spdo is found at the cell surface in the absence of Numb (i.e. in the cells that do not inherit Numb or in numb-mutant cells), whereas, in the presence of Numb (i.e. in the cells that inherit Numb), Spdo accumulates in uncharacterized intracellular structures that might correspond to endocytic compartments [40]. This would, therefore, suggest that one function of Numb is to inhibit Spdo activity by mediating its endocytosis, and that Spdo cell surface membrane localization is essential for N signaling. Whether Numb regulates the localization of N signaling components other than Spdo is, at present, unclear, as is the mechanism by which Spdo acts to aid N signaling. Neuralized

Regulation of N-signaling within the SOP is also controlled by Neuralized (Neur), which is essential for specification of the pIIa cell [41,42,43]. Neur is a ring-finger containing E3 ubiquitin ligase that targets Dl for endocytosis [44–47]. Similar to Numb, Neur is asymmetrically segregated to the pIIb cell during the pI division, suggesting that Neur acts in the pIIb cell to promote Nsignaling in the pIIa cell (Figure 2). Clonal analysis has, indeed, demonstrated a non-autonomous role for neur in promoting Dl endocytosis and N-signaling [43,46], although a cell-autonomous role in Dl endocytosis has also been proposed [41,42]. By what mechanism Dl endocytosis promotes N signaling is not yet understood. Additionally, a direct cortical anchor of Neur has yet to be identified. It will be of interest to determine whether Neur is similarly asymmetrically localized in the NB lineage and regulates cell fate therein. Through their asymmetric inheritance, Pros, Numb and Neur, thus enable acquisition of distinct daughter fates. We now examine how the localization of these regulators and their anchors is coordinated, by cell polarity machinery, with the cell division axis. www.sciencedirect.com

Segregating cell fate determinants Despite the different downstream events regulated by Numb, Neur, and Pros, conserved mechanisms exist that co-regulate their localization and link it to spindle orientation, allowing asymmetry to be generated upon cell division. The emerging theme is that the coordinated effort of several protein complexes and the actin cytoskeleton leads to the establishment of cortical domains that are crucial for segregation of cell fate determinants, spindle alignment and daughter cell size. The apical complex of the NB

Within the NB, two conserved protein subcomplexes act together with the actin cytoskeleton to divide the cell cortex into apical and a basal domains. One of the subcomplexes is composed of the GoLoCo motif-containing protein, ‘Partner of Inscuteable’ (Pins) and its associated G-protein a subunit (Gai) [48–51]. A second subcomplex consists of the atypical protein kinase C (DaPKC) and two PDZ (PSD95/Discs large/ZO1 domain)-containing proteins, DmPar6 and Bazooka (Baz) [52–55] (see [7] for review). An adaptor protein, Inscuteable (Insc), forms the ‘apical complex’ by linking these two subcomplexes. The apical localization, and in many cases, the stability of each member of this complex is interdependent on the other ‘apical complex’ members in the NB. The G-protein b and g complex (Gbg), which competes with Pins for Gai binding, is also essential for apical complex localization and stability [51,56]. The loss of any component of the apical complex results in loss of metaphase localization of the Pon-Numb and Miranda-Pros crescents to the basal cortex, although crescent formation can occur later in telophase, through an unknown mechanism of ‘telophase rescue’. The apical complex also coordinates the alignment of the mitotic spindle with cell fate determinants and regulates spindle asymmetry, thereby controlling daughter cell size [11,49,50,51,52–54,56–58]. Lgl as a target of DaPKC

Elegant studies by Betschinger and co-workers [59], recently, have shed light on the mechanism used by the apical complex to direct localization of cell-fate regulators to the basal cortex (Figure 3). This mechanism involves lethal(2) giant larvae (lgl), a gene required for asymmetric segregation of cell-fate determinants [60,61]. Lgl interacts with DaPKC and DmPar6, and is a direct phosphorylation target of DaPKC [59]. In NBs, Lgl localizes at the cortex and its cortical localization depends on Discs-large (Dlg), a protein that is also required for the basal localization of cell-fate determinants [60,61]. In cells expressing either a version of DaPKC that is no longer restricted to the apical cortex, or a non-phosphorylatable Lgl, Miranda is no longer restricted to the basal cortex but is either cytoplasmic (former) or found around the entire cell cortex (latter). These results have led to a model, whereby, phosphorylation of Lgl by DaPKC at the apical cortex inactivates Current Opinion in Neurobiology 2004, 14:6–14

10 Development

Figure 3

Apical aPKC Lgl-P

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model, whereby Lgl antagonizes the myosin II-dependent exclusion of cell-fate determinants from the cortex [62]; however, additional roles for Lgl in NBs cannot be excluded. Indeed, the homologue of Lgl in Saccharomyces cerevisiae is involved in exocytic trafficking [64], suggesting that another possible function of Lgl in NBs might be the regulation of the anchoring of vesicular cargos at the basal cortex. Role of the actin cytoskeleton

Miranda

Basal Current Opinion in Neurobiology

Regulation of Lgl and basal component localization in the NB. Recent work has suggested a model for the way that apically localized components lead to basal cell-fate regulator localization. aPKC phosphorylates and inactivates Lgl at the apical surface (where aPKC is localized with other apical complex components), thus, resulting in asymmetric Lgl activity at the basal cortex [59]. Lgl, in turn, negatively regulates the localization of Myosin II, resulting in MyoII localization to the apical cortex [62]. Cortical MyoII (presumably myosin filaments) blocks Miranda localization [60,61]. Additionally, phosphorylation of MyoII by Rho kinase is required for Miranda and other basal component localization. How Miranda is inhibited by MyoII and the direct binding partner of Miranda, Pros, Numb and Pon at the basal cortex is yet unknown.

Lgl, leaving the remaining basally located Lgl as the active pool (Figure 3) [59]. These studies have, therefore, established Lgl as a major target of the ‘apical complex’ in the localization of basal determinants. Inhibition of Myosin function by Lgl

In a more recent study, Barros and co-workers [62] have proposed that one function of Lgl in NBs is to inhibit Myosin activity. Lgl was known to bind to myosin II [63] and to antagonize myosin II function [60,61]. Importantly, myosin II function is required for the basal localization of cell-fate determinants. A reduction in levels of myosin regulatory light chain (MRLC) or drug-mediated inhibition of Rho kinase, a known myosin II activator, disrupts asymmetric localization of Miranda at the cortex, but Baz remains properly localized at the apical cortex [62]. The effect of the Rho kinase inhibitor is suppressed by the expression of a phosphomimetic form of MRLC, indicating that the effect of Rho kinase inhibition results from the reduced level of MRLC phosphorylation. Moreover, myosin II localizes, predominantly, at the apical cortex at metaphase, in opposition to Miranda, suggesting that myosin II antagonizes Miranda localization. The apical localization of myosin II depends on the DaPKC-mediated inhibition of Lgl, because loss of lgl function results in uniform cortical localization of myosin II, whereas, expression of a non-phosphorylatable form of Lgl, mimicking ‘active Lgl’, results in the cytoplasmic accumulation of myosin II. These results, then, suggest a Current Opinion in Neurobiology 2004, 14:6–14

Disruption of cytoskeletal structures, through drug treatments, has defined the requirements for cortical localization of the cell-fate determinants. The localization of Numb, Pon, Neur, Pros and Miranda, during mitosis into basal (NB) or anterior crescents (SOP), is dependent on the actin cytoskeleton, but independent of polymerized microtubules [25,28,43,65–67]. The function of the actin cytoskeleton in localizing cell-fate determinants is not well understood. First, actin-dependent myosin motors might transport cell-fate determinant to the basal cortex. For instance, the myosin VI Jaguar has been shown to co-immunoprecipitate with Miranda and to be required for Miranda crescent formation, raising the possibility that myosin VI-dependent transport localizes Miranda to the basal cortex [68]. Consistent with this hypothesis, treatment of cells with the myosin motor inhibitor butanedione-2-monoxime (BDM) abolishes crescent formation of Pon, Miranda and Neur [28,43, 60,61]. However, more recent studies have indicated that non-muscle myosins are not inhibited by BDM [69]. Instead, BDM appears to block actin polymerization [70]. Second, active myosin at the apical cortex has now been proposed to modify the actin cytoskeleton to exclude Miranda binding [62]. Future studies should aim to better define the way that the actin cytoskeleton positively and/or negatively regulates the cortical localization of cell-fate determinants. Regulation of cortical domains within the SOP

In opposition to the NB, the SOP cell divides planar to the epithelium and establishes separate cortical domains of Pins–Gai (at the anterior cortex) and Baz–DaPKC– DmPar6 (at the posterior cortex) [51,71]. Localization of these two complexes in separate domains is dependent on the lack of insc expression in the SOP [71]. Pon–Numb colocalizes with Pins–Gai at the anterior cortex [27,51,71]. Interestingly, Dlg binds to Pins and is co-localized to the anterior cortex [71]. Dlg–Pins–Gai and Baz, as in the NB, are required for proper localization of Numb [51,71]; likewise, segregation of Neur to the pIIb is also dependent on Dlg and Pins [43]. The role played by Lgl in the SOP for asymmetric segregation of Numb and cell fate is somewhat controversial. Ohshiro and co-workers [60] have reported that, as in the NB, lglts3 mutant SOPs possess delocalized Numb crescents, as well as bristle phenotypes, consistent with a pIIa-to-pIIb cell fate transformation that might be associated with Numb miswww.sciencedirect.com

Asymmetric localization and function of cell-fate determinants Bardin, Le Borgne and Schweisguth 11

segregation. Justice et al. [72] have recently reported a contrasting result. In mitotic clones of a complete loss of function lgl allele, Numb crescent formation at the anterior cortex is unperturbed. Additionally, pIIb-to-pIIa cell fate transformations were observed and genetic studies suggest that Lgl acts upstream of NICD in the pIIa vs pIIb fate decision and, in contrast to the NB, downstream or in parallel to numb. The mechanism of Lgl action is an important issue to resolve, to understand how the cell-fate determinants are localized within the SOP.

Unequal segregation of Numb and asymmetric cell division in Vertebrates The fundamental importance of asymmetric cell division in the development of the nervous system in invertebrates raises the question of whether unequal segregation of fate determinants also contributes to the generation of various cell types in the vertebrate nervous system. The molecular conservation of the fate determinants Numb and Neur is consistent with this suggestion [38,73–76]. By contrast, no Miranda homologs have been identified in sequenced vertebrate genomes, and there is no evidence for unequal distribution of Pros homologs at mitosis. The identification of Neur as a cell-fate determinant in the fly is too recent to have permitted an analysis of the role of Neur homologs in asymmetric divisions in vertebrates; but various studies have examined the possible role of Numb in asymmetric cell division. In the developing cortex of mammals, neuroepithelial cells are polarized along their apical–basal axis and have been shown to divide parallel (horizontally), perpendicular (vertically) or with an oblique angle, relative to the lumen of the tube. Results from live imaging analyses suggested that horizontal divisions of progenitor cells were proliferative (symmetrical), thus, augmenting the pool of neural progenitor cells, whereas, vertical divisions were differentiative (asymmetrical), generating one apical cell that remained in the proliferative zone and one basal cell that behaved as a newly born neuron [77]. A recent double knockout analysis of numb and numb-like mutant mouse embryos has indicated that Numb and Numb-like play an essential (yet partly redundant) role in maintaining neural progenitor cell populations [78]. Mouse Numb has been shown to localize at the apical cortex of dividing neuroepithelial cells, although Numb has also been suggested to preferentially accumulate into the neuronal daughter (i.e. the presumed ‘basal’ daughter [79]), bind mouse Notch and inhibit Notch signaling [38,80]. Therefore, a model is suggested where orientation of the division regulates the equal versus unequal partitioning of Numb, hence the fate asymmetry in the choice between neural progenitor and neuron. Consistent with this model, vertical divisions have been observed in fixed tissue studies of chick and rat retina [81,82], and chick cortex [39]; furthermore, unequal partitioning of Numb is correlated with a fate difference, following www.sciencedirect.com

ex vivo divisions of isolated progenitor cells from mouse cortex [79]. Finally, a recent time-lapse analysis of rat retina explants has suggested that vertical divisions tend to produce cells with distinct identities, while horizontal divisions exhibit the opposite tendency [83]. Despite these exciting observations, direct in vivo evidence that unequal segregation of Numb in vertical division regulates a fate difference between the two daughter cells is still lacking. And more significantly, this simple model has been challenged by the following observations. First, the number of vertical divisions throughout neurogenesis appears to be too low to account for the large number of emerging neurons [77,81]. Second, no correlation was observed between the orientation of the division, Numb segregation and the expression of an early neural marker in the chick retina [82]. Third, a recent time-lapse analysis of divisions in living zebrafish embryos has indicated that most neurons are born from divisions that produce two neurons and that no correlation exists between the apical–basal orientation of the division and the fate of the daughter cells [84]. To what extent these contradictory conclusions reflect differences in model organisms, regional identity within the nervous system, experimental approaches and/or marker used, remains to be examined. Additionally, interpretation of some of the published data is complicated by two confounding points. First, as initially pointed out by Huttner and Brand in 1997 [85], unequal segregation of apical Numb might also result from oblique cleavage in which the apical domain is unequally bisected. Thus, divisions that are described as horizontal could be asymmetric, in terms of Numb partitioning. Second, the axis of polarity used for unequal partitioning of Numb (or other fate determinants) is not necessarily the apical–basal axis. For instance, the polarization machinery uses the anterior–posterior polarity of the SOP to localize Numb and Neur at the anterior pole. Interestingly, a recent in vivo imaging analysis of division in the zebrafish retina has suggested that the orientation of progenitor cell divisions is regulated by planar central–peripheral polarity cues [86]. This study nicely illustrates the importance of green fluorescent protein (GFP) imaging in the study of asymmetric cell division. Clearly, combining live imaging in whole organisms and tissue explants with GFP probes, revealing both orientation of division and, most importantly, unequal partitioning of fate determinants, should help to establish the role of Numb unequal partitioning in generating cell-fate diversity.

Perspectives During the past ten years, genetic studies of asymmetric cell division in Drosophila have dramatically increased our understanding of the molecular basis of asymmetric cell division. This review has discussed several mechanisms by which distinct daughter cell fates arise in the nervous system of Drosophila. Although the role of vertebrate Current Opinion in Neurobiology 2004, 14:6–14

12 Development

homologs of fate determinants in flies is either debated (Numb) or unknown (Neur), Drosophila will certainly continue to contribute to our understanding. Nevertheless, despite the importance of Drosophila as a model system, we must bear in mind that acquisition of distinct cell fates during asymmetric cell division might be acquired through mechanisms other than those that have been identified, so far, in the developing Drosophila nervous system. For instance, microtubule-based directional transport [87] and/or asymmetric mRNA/protein degradation [88] have been shown to contribute to unequal partitioning of fate determinants in molluscs and worms. Future studies on asymmetric cell divisions in various systems, including vertebrates, should further our understanding of how stem cells are maintained, how distinct cell types arise and differentiate, and how these processes can become deregulated, leading to pathologies such as cancers.

Acknowledgements We thank C. Barros and A. Brand for sharing unpublished data, and C. Goridis for critical reading.

References and recommended reading Papers of particular interest, published within the annual period of review, have been highlighted as:  of special interest  of outstanding interest

12. Spana EP, Doe CQ: The Prospero transcription factor is asymmetrically localized to the cell cortex during neuroblast mitosis in Drosophila. Development 1995, 121:3187-3195. 13. Fortini ME: Gamma-secretase-mediated proteolysis in cell-surface-receptor signalling. Nat Rev Mol Cell Biol 2002, 3:673-684. 14. Hartenstein V, Posakony JW: A dual function of the Notch gene in Drosophila sensillum development. Dev Biol 1990, 142:13-30. 15. Schweisguth F, Posakony JW: Antagonistic activities of Suppressor of Hairless and Hairless control alternative cell fates in the Drosophila adult epidermis. Development 1994, 120:1433-1441. 16. Zeng C, Younger-Shepherd S, Jan LY, Jan YN: Delta and Serrate are redundant Notch ligands required for asymmetric cell divisions within the Drosophila sensory organ lineage. Genes Dev 1998, 12:1086-1091. 17. Guo M, Bier E, Jan LY, Jan YN: tramtrack acts downstream of numb to specify distinct daughter cell fates during asymmetric cell divisions in the Drosophila PNS. Neuron 1995, 14:913-925. 18. Okabe M, Imai T, Kurusu M, Hiromi Y, Okano H: Translational repression determines a neuronal potential in Drosophila asymmetric cell division. Nature 2001, 411:94-98. 19. Pi H, Wu HJ, Chien CT: A dual function of phyllopod in Drosophila external sensory organ development: cell fate specification of sensory organ precursor and its progeny. Development 2001, 128:2699-2710. 20. Li P, Yang X, Wasser M, Cai Y, Chia W: Inscuteable and Staufen mediate asymmetric localization and segregation of prospero RNA during Drosophila neuroblast cell divisions. Cell 1997, 90:437-447. 21. Broadus J, Fuerstenberg S, Doe CQ: Staufen-dependent localization of prospero mRNA contributes to neuroblast daughter-cell fate. Nature 1998, 391:792-795.

1.

Horvitz HR, Herskowitz I: Mechanisms of asymmetric cell division: two Bs or not two Bs, that is the question. Cell 1992, 68:237-255.

2.

Reddy GV, Rodrigues V: A glial cell arises from an additional division within the mechanosensory lineage during development of the microchaete on the Drosophila notum. Development 1999, 126:4617-4622.

23. Shen CP, Jan LY, Jan YN: Miranda is required for the asymmetric localization of Prospero during mitosis in Drosophila. Cell 1997, 90:449-458.

3.

Gho M, Bellaiche Y, Schweisguth F: Revisiting the Drosophila microchaete lineage: a novel intrinsically asymmetric cell division generates a glial cell. Development 1999, 126:3573-3584.

24. Schuldt AJ, Adams JH, Davidson CM, Micklem DR, Haseloff J, St Johnston D, Brand AH: Miranda mediates asymmetric protein and RNA localization in the developing nervous system. Genes Dev 1998, 12:1847-1857.

4.

Fichelson P, Gho M: The glial cell undergoes apoptosis in the microchaete lineage of Drosophila. Development 2003, 130:123-133.

5.

Lyczak R, Gomes JE, Bowerman B: Heads or tails: cell polarity and axis formation in the early Caenorhabditis elegans embryo. Dev Cell 2002, 3:157-166.

25. Shen CP, Knoblich JA, Chan YM, Jiang MM, Jan LY, Jan YN: Miranda as a multidomain adapter linking apically localized Inscuteable and basally localized Staufen and Prospero during asymmetric cell division in Drosophila. Genes Dev 1998, 12:1837-1846.

22. Ikeshima-Kataoka H, Skeath JB, Nabeshima Y, Doe CQ, Matsuzaki F: Miranda directs Prospero to a daughter cell during Drosophila asymmetric divisions. Nature 1997, 390:625-629.

26. Matsuzaki F, Ohshiro T, Ikeshima-Kataoka H, Izumi H: miranda localizes staufen and prospero asymmetrically in mitotic neuroblasts and epithelial cells in early Drosophila embryogenesis. Development 1998, 125:4089-4098.

6.

Pellettieri J, Seydoux G: Anterior-posterior polarity in C. elegans and Drosophila – PARallels and differences. Science 2002, 298:1946-1950.

7.

Henrique D, Schweisguth F: Cell polarity: the ups and downs of the Par6/aPKC complex. Curr Opin Genet Dev 2003, 13:341-350.

8.

Doe CQ, Chu-LaGraff Q, Wright DM, Scott MP: The prospero gene specifies cell fates in the Drosophila central nervous system. Cell 1991, 65:451-464.

28. Lu B, Rothenberg M, Jan LY, Jan YN: Partner of Numb colocalizes with Numb during mitosis and directs Numb asymmetric localization in Drosophila neural and muscle progenitors. Cell 1998, 95:225-235.

9.

Vaessin H, Grell E, Wolff E, Bier E, Jan LY, Jan YN: Prospero is expressed in neuronal precursors and encodes a nuclear protein that is involved in the control of axonal outgrowth in Drosophila. Cell 1991, 67:941-953.

29. Frise E, Knoblich JA, Younger-Shepherd S, Jan LY, Jan YN: The Drosophila Numb protein inhibits signaling of the Notch receptor during cell-cell interaction in sensory organ lineage. Proc Natl Acad Sci USA 1996, 93:11925-11932.

10. Hirata J, Nakagoshi H, Nabeshima Y, Matsuzaki F: Asymmetric segregation of the homeodomain protein Prospero during Drosophila development. Nature 1995, 377:627-630. 11. Knoblich JA, Jan LY, Jan YN: Asymmetric segregation of Numb and Prospero during cell division. Nature 1995, 377:624-627. Current Opinion in Neurobiology 2004, 14:6–14

27. Rhyu MS, Jan LY, Jan YN: Asymmetric distribution of numb protein during division of the sensory organ precursor cell confers distinct fates to daughter cells. Cell 1994, 76:477-491.

30. Spana EP, Doe CQ: Numb antagonizes Notch signaling to specify sibling neuron cell fates. Neuron 1996, 17:21-26. 31. Guo M, Jan LY, Jan YN: Control of daughter cell fates during asymmetric division: interaction of Numb and Notch. Neuron 1996, 17:27-41. www.sciencedirect.com

Asymmetric localization and function of cell-fate determinants Bardin, Le Borgne and Schweisguth 13

32. Spana EP, Kopczynski C, Goodman CS, Doe CQ: Asymmetric localization of numb autonomously determines sibling neuron identity in the Drosophila CNS. Development 1995, 121:3489-3494. 33. Uemura T, Shepherd S, Ackerman L, Jan LY, Jan YN: Numb, a gene required in determination of cell fate during sensory organ formation in Drosophila embryos. Cell 1989, 58:349-360. 34. Buescher M, Yeo SL, Udolph G, Zavortink M, Yang X, Tear G, Chia W: Binary sibling neuronal cell fate decisions in the Drosophila embryonic central nervous system are nonstochastic and require inscuteable-mediated asymmetry of ganglion mother cells. Genes Dev 1998, 12:1858-1870. 35. Lear BC, Skeath JB, Patel NH: Neural cell fate in rca1 and cycA mutants: the roles of intrinsic and extrinsic factors in asymmetric division in the Drosophila central nervous system. Mech Dev 1999, 88:207-219. 36. Santolini E, Puri C, Salcini AE, Gagliani MC, Pelicci PG, Tacchetti C, Di Fiore PP: Numb is an endocytic protein. J Cell Biol 2000, 151:1345-1352. 37. Berdnik D, Torok T, Gonzalez-Gaitan M, Knoblich JA: The endocytic protein alpha-Adaptin is required for numbmediated asymmetric cell division in Drosophila. Dev Cell 2002, 3:221-231. 38. Zhong W, Feder JN, Jiang MM, Jan LY, Jan YN: Asymmetric localization of a mammalian numb homolog during mouse cortical neurogenesis. Neuron 1996, 17:43-53. 39. Wakamatsu Y, Maynard TM, Jones SU, Weston JA: Numb localizes in the basal cortex of mitotic avian neuroepithelial cells and modulates neuronal differentiation by binding to NOTCH-1. Neuron 1999, 23:71-81. 40. O’Connor-Giles KM, Skeath JB: Numb inhibits membrane  localization of Sanpodo, a four-pass transmembrane protein, to promote asymmetric divisions in Drosophila. Dev Cell 2003, 5:231-243. This paper re-characterizes the sanpodo gene and demonstrates that it encodes a transmembrane protein that is required for N signaling, that functions upstream of the S3 cleavage of N but downstream of Dl. Spdo localizes in a numb-dependent manner to intracellular foci, presumed to be endocytic compartments in the dMP2 cell (that inherits Numb) and at the cell surface in the vMP2 cell (that has activated N). Importantly, Sanpodo is shown to associate both with Numb and N in co-immunoprecipitation experiments. The authors propose that Numb acts by inactivating Sanpodo in the dMP2 cell through endocytic downregulation of surface Sanpodo, while in the vMP2 cell, the lack of Numb leaves Sanpodo at the surface where it is required for N signaling. 41. Lai EC, Rubin GM: neuralized functions cell-autonomously to regulate a subset of notch-dependent processes during adult Drosophila development. Dev Biol 2001, 231:217-233. 42. Yeh E, Zhou L, Rudzik N, Boulianne GL: Neuralized functions cell autonomously to regulate Drosophila sense organ development. EMBO J 2000, 19:4827-4837. 43. Le Borgne R, Schweisguth F: Unequal segregation of Neuralized  biases Notch activation during asymmetric cell division. Dev Cell 2003, 5:139-148. The E3 ubiquitin ligase, Neuralized, which was previously known to be important for SOP cell-fate decisions and for ubiquitination and endocytosis of Dl, is shown, in this paper, to be asymmetrically segregated to the pIIb cell. A novel assay is employed to investigate Dl trafficking in living cells; it shows enhanced endocytosis of Dl in the pIIb cell, which is greatly reduced in neur mutant clones. Furthermore, the authors provide evidence that neur acts non-cell autonomously to promote pIIa cell fate. 44. Deblandre GA, Lai EC, Kintner C: Xenopus neuralized is a ubiquitin ligase that interacts with XDelta1 and regulates Notch signaling. Dev Cell 2001, 1:795-806. 45. Lai EC, Deblandre GA, Kintner C, Rubin GM: Drosophila neuralized is a ubiquitin ligase that promotes the internalization and degradation of delta. Dev Cell 2001, 1:783-794. 46. Pavlopoulos E, Pitsouli C, Klueg KM, Muskavitch MA, Moschonas NK, Delidakis C: neuralized Encodes a peripheral membrane protein involved in delta signaling and endocytosis. Dev Cell 2001, 1:807-816. www.sciencedirect.com

47. Yeh E, Dermer M, Commisso C, Zhou L, McGlade CJ, Boulianne GL: Neuralized functions as an E3 ubiquitin ligase during Drosophila development. Curr Biol 2001, 11:1675-1679. 48. Parmentier ML, Woods D, Greig S, Phan PG, Radovic A, Bryant P, O’Kane CJ: Rapsynoid/partner of inscuteable controls asymmetric division of larval neuroblasts in Drosophila. J Neurosci 2000, 20:RC84. 49. Schaefer M, Shevchenko A, Knoblich JA: A protein complex containing Inscuteable and the Galpha-binding protein Pins orients asymmetric cell divisions in Drosophila. Curr Biol 2000, 10:353-362. 50. Yu F, Morin X, Cai Y, Yang X, Chia W: Analysis of partner of inscuteable, a novel player of Drosophila asymmetric divisions, reveals two distinct steps in inscuteable apical localization. Cell 2000, 100:399-409. 51. Schaefer M, Petronczki M, Dorner D, Forte M, Knoblich JA: Heterotrimeric G proteins direct two modes of asymmetric cell division in the Drosophila nervous system. Cell 2001, 107:183-194. 52. Schober M, Schaefer M, Knoblich JA: Bazooka recruits Inscuteable to orient asymmetric cell divisions in Drosophila neuroblasts. Nature 1999, 402:548-551. 53. Wodarz A, Ramrath A, Kuchinke U, Knust E: Bazooka provides an apical cue for Inscuteable localization in Drosophila neuroblasts. Nature 1999, 402:544-547. 54. Wodarz A, Ramrath A, Grimm A, Knust E: Drosophila atypical protein kinase C associates with Bazooka and controls polarity of epithelia and neuroblasts. J Cell Biol 2000, 150:1361-1374. 55. Petronczki M, Knoblich JA: DmPAR-6 directs epithelial polarity and asymmetric cell division of neuroblasts in Drosophila. Nat Cell Biol 2001, 3:43-49. 56. Yu F, Cai Y, Kaushik R, Yang X, Chia W: Distinct roles of Galphai  and Gbeta13F subunits of the heterotrimeric G-protein complex in the mediation of Drosophila neuroblast asymmetric divisions. J Cell Biol 2003, 162:623-633. See annotation for [58]. 57. Cai Y, Yu F, Lin S, Chia W, Yang X: Apical complex genes control  mitotic spindle geometry and relative size of daughter cells in Drosophila neuroblast and pI asymmetric divisions. Cell 2003, 112:51-62. See annotation for [58]. 58. Fuse N, Hisata K, Katzen AL, Matsuzaki F: Heterotrimeric g  proteins regulate daughter cell size asymmetry in Drosophila neuroblast divisions. Curr Biol 2003, 13:947-954. Together, these four elegant studies establish that the activity of the Gai and Gb13F genes is essential for the basal localization of cell-fate determinants in dividing NBs and SOPs. The GDP-bound form of Gai binds to Pins, and Gai is shown to localize at the apical cortex of NBs. Moreover, loss-of-function alleles of Gai give similar phenotypes to loss of pins function. Furthermore, the subcomplexes Gai–Pins and Baz–Par6– aPKC each act redundantly in NBs to regulate the geometry of the mitotic spindle, hence the daughter cell-size difference. Consistent with this, daughter cells have equal size in the Gb13F mutant embryos and Gb13F is required for apical localization of the Gai–Pins and Baz–Par6–aPKC subcomplexes. 59. Betschinger J, Mechtler K, Knoblich JA: The Par complex directs  asymmetric cell division by phosphorylating the cytoskeletal protein Lgl. Nature 2003, 422:326-330. In this study, Lgl is found to co-immunoprecipitate in a complex with Par6 and aPKC, but not with Baz. Lgl is shown to be phosphorylated by aPKC in vitro and in vivo. Mutations of the phosphorylation sites abolish the aPKC-dependent delocalization of Lgl from the cortex in S2 cells. Importantly, overexpression of non-phosphorylatable Lgl in NB, but not wildtype Lgl, causes Miranda to be distributed around the entire cell cortex. Similarly, expression of a version of aPKC that cannot anchor to the apical cortex causes Miranda to be cytoplasmic. The authors propose a model in which apically localized aPKC phosphorylates Lgl and inactivates it locally, resulting in localization of Miranda to the basal cortex, where Lgl remains unphosphorylated and active. 60. Ohshiro T, Yagami T, Zhang C, Matsuzaki F: Role of cortical tumour-suppressor proteins in asymmetric division of Drosophila neuroblast. Nature 2000, 408:593-596. Current Opinion in Neurobiology 2004, 14:6–14

14 Development

61. Peng CY, Manning L, Albertson R, Doe CQ: The tumoursuppressor genes lgl and dlg regulate basal protein targeting in Drosophila neuroblasts. Nature 2000, 408:596-600. 62. Barros CS, Phelps CB, Brand AH: Drosophila non-muscle myosin  II promotes the asymmetric segregation of cell fate determinants by cortical exclusion rather than acive transport. Dev Cell 2003, in press. Here, the role that myosin II plays in the basal localization of Pros and Miranda in the NB is characterized. In embryos with strongly reduced maternal and zygotic MRLC function, Miranda no longer localized at the cortex. Additionally, the authors demonstrate that Rho kinase phosphorylation of MyoII is necessary for its function and that Lgl inhibits MyoII function. Because MyoII localized to the apical pole of the NB, the authors propose a model of asymmetric localization, based on cortical exclusion rather than transport. In this model, MyoII would block basal component localization through its localization to the apical pole, which is controlled by Lgl. 63. Strand D, Jakobs R, Merdes G, Neumann B, Kalmes A, Heid HW, Husmann I, Mechler BM: The Drosophila lethal(2)giant larvae tumor suppressor protein forms homo-oligomers and is associated with nonmuscle myosin II heavy chain. J Cell Biol 1994, 127:1361-1373. 64. Lehman K, Rossi G, Adamo JE, Brennwald P: Yeast homologs of tomosyn and lethal giant larvae function in exocytosis and are associated with the plasma membrane SNARE, Sec9. J Cell Biol 1999, 146:125-140. 65. Broadus J, Doe CQ: Extrinsic cues, intrinsic cues and microfilaments regulate asymmetric protein localization in Drosophila neuroblasts. Curr Biol 1997, 7:827-835. 66. Knoblich JA, Jan LY, Jan YN: The N terminus of the Drosophila Numb protein directs membrane association and actindependent asymmetric localization. Proc Natl Acad Sci USA 1997, 94:13005-13010. 67. Berdnik D, Knoblich JA: Drosophila Aurora-A is required for centrosome maturation and actin-dependent asymmetric protein localization during mitosis. Curr Biol 2002, 12:640-647. 68. Petritsch C, Tavosanis G, Turck CW, Jan LY, Jan YN: The Drosophila myosin VI Jaguar is required for basal protein targeting and correct spindle orientation in mitotic neuroblasts. Dev Cell 2003, 4:273-281. 69. Cheung A, Dantzig JA, Hollingworth S, Baylor SM, Goldman YE, Mitchison TJ, Straight AF: A small-molecule inhibitor of skeletal muscle myosin II. Nat Cell Biol 2002, 4:83-88. 70. Yarrow JC, Lechler T, Li R, Mitchison TJ: Rapid de-localisation of actin leading edge components with BDM treatment. BMC Cell Biol 2003, 4. 71. Bellaiche Y, Radovic A, Woods DF, Hough CD, Parmentier ML, O’Kane CJ, Bryant PJ, Schweisguth F: The partner of Inscuteable/Discs-large complex is required to establish planar polarity during asymmetric cell division in Drosophila. Cell 2001, 106:355-366. 72. Justice N, Roegiers F, Jan LY, Jan YN: Lethal giant larvae acts  together with numb in notch inhibition and cell fate specification in the Drosophila adult sensory organ precursor lineage. Curr Biol 2003, 13:778-783. Clonal analysis, using a null allele of lgl, indicates that Lgl is not required for Numb asymmetric localization in the SOP division, but is required to promote the pIIb cell fate. Lgl acts upstream of activated N and functions either downstream or parallel to Numb to block N signaling. 73. Verdi JM, Schmandt R, Bashirullah A, Jacob S, Salvino R, Craig CG, Program AE, Lipshitz HD, McGlade CJ: Mammalian NUMB is an

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evolutionarily conserved signaling adapter protein that specifies cell fate. Curr Biol 1996, 6:1134-1145. 74. Zhong W, Jiang MM, Weinmaster G, Jan LY, Jan YN: Differential expression of mammalian Numb, Numblike and Notch1 suggests distinct roles during mouse cortical neurogenesis. Development 1997, 124:1887-1897. 75. Timmusk T, Palm K, Belluardo N, Mudo G, Neuman T: Dendritic localization of mammalian neuralized mRNA encoding a protein with transcription repression activities. Mol Cell Neurosci 2002, 20:649-668. 76. Pavlopoulos E, Kokkinaki M, Koutelou E, Mitsiadis TA, Prinos P, Delidakis C, Kilpatrick MW, Tsipouras P, Moschonas NK: Cloning, chromosomal organization and expression analysis of Neurl, the mouse homolog of Drosophila melanogaster neuralized gene. Biochim Biophys Acta 2002, 1574:375-382. 77. Chenn A, McConnell SK: Cleavage orientation and the asymmetric inheritance of Notch1 immunoreactivity in mammalian neurogenesis. Cell 1995, 82:631-641. 78. Petersen PH, Zou K, Hwang JK, Jan YN, Zhong W: Progenitor cell maintenance requires numb and numblike during mouse neurogenesis. Nature 2002, 419:929-934. 79. Shen Q, Zhong W, Jan YN, Temple S: Asymmetric Numb distribution is critical for asymmetric cell division of mouse cerebral cortical stem cells and neuroblasts. Development 2002, 129:4843-4853. 80. Sestan N, Artavanis-Tsakonas S, Rakic P: Contact-dependent inhibition of cortical neurite growth mediated by notch signaling. Science 1999, 286:741-746. 81. Cayouette M, Whitmore AV, Jeffery G, Raff M: Asymmetric segregation of Numb in retinal development and the influence of the pigmented epithelium. J Neurosci 2001, 21:5643-5651. 82. Silva AO, Ercole CE, McLoon SC: Plane of cell cleavage and numb distribution during cell division relative to cell differentiation in the developing retina. J Neurosci 2002, 22:7518-7525. 83. Cayouette M, Raff M: The orientation of cell division influences cell-fate choice in the developing mammalian retina. Development 2003, 130:2329-2339. 84. Lyons DA, Guy AT, Clarke JD: Monitoring neural progenitor fate through multiple rounds of division in an intact vertebrate brain. Development 2003, 130:3427-3436. 85. Huttner WB, Brand M: Asymmetric division and polarity of neuroepithelial cells. Curr Opin Neurobiol 1997, 7:29-39. 86. Das T, Payer B, Cayouette M, Harris WA: In vivo time-lapse  imaging of cell divisions during neurogenesis in the developing zebrafish retina. Neuron 2003, 37:597-609. Live imaging, using two-photon microscopy of developing zebrafish retina reveals that divisions are planar and that the orientation of divisions changes over time in both zebrafish and rat, from the central-peripheral axis in early neurogenesis to circumferential axis later in neurogenesis. This study raises the possibility that planar polarity cues might regulate asymmetric divisions during CNS development. 87. Lambert JD, Nagy LM: Asymmetric inheritance of centrosomally localized mRNAs during embryonic cleavages. Nature 2002, 420:682-686. 88. DeRenzo C, Reese KJ, Seydoux G: Exclusion of germ plasm proteins from somatic lineages by cullin-dependent degradation. Nature 2003, 424:685-689.

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