The SUN Rises on Meiotic Chromosome Dynamics

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1 The SUN Rises on Meiotic Chromosome Dynamics Yasushi Hiraoka 1,2, * and Abby F. Dernburg 3,4, * 1 Graduate School of Frontier Biosciences, Osaka University, 1-3 Yamadaoka, Suita , Japan 2 Kobe Advanced ICT Research Center, National Institute of Information and Communications Technology, Iwaoka, Iwaoka-cho, Nishi-ku, Kobe , Japan 3 Howard Hughes Medical Institute and Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA , USA 4 Life Sciences Division, Lawrence Berkeley National Laboratory, Berkeley, CA 94720, USA *Correspondence: hiraoka@fbs.osaka-u.ac.jp (Y.H.), afdernburg@lbl.gov (A.F.D.) DOI /j.devcel Recent studies in diverse eukaryotes have implicated a family of nuclear envelope proteins containing SUN domains as key components of meiotic nuclear organization and chromosome dynamics. In many cases, these transmembrane proteins are also known to contribute to centrosome or spindle pole body function in mitotically dividing cells. During meiotic prophase, the apparent role of these SUN-domain proteins, together with their partner KASH-domain proteins, is to connect chromosomes through the intact nuclear envelope to force-generating mechanisms in the cytoplasm. A Family of SUN Proteins The SUN domain was named for homologous sequences shared by Sad1 and UNC-84 proteins (Malone et al., 1999). Sad1 was originally identified as an essential component of the spindle pole body (SPB), which organizes the mitotic spindle in the fission yeast Schizosaccharomyces pombe (Hagan and Yanagida, 1995). UNC-84 is a nuclear membrane protein required for nuclear migration and positioning in the nematode Caenorhabditis elegans, mutations in which produce an uncoordinated phenotype due to neurological defects (Malone et al., 1999). The C. elegans genome contains another gene identified initially by its homology to unc-84, called sun-1 or matefin/mtf- 1 (from the Hebrew word for envelope ; Fridkin et al., 2004; Malone et al., 2003). Both Sad1 and SUN-1 play major roles in meiotic chromosome dynamics, in addition to their essential roles in mitotically dividing cells, as discussed below. Evolutionary analysis has indicated that proteins containing SUN domains are among the most broadly conserved nuclear envelope (NE) proteins and are therefore likely to be among the most ancient in origin (Mans et al., 2004). The eukaryotic SUNdomain proteins are highly diverse (Jaspersen et al., 2006; Mans et al., 2004; Figure 1). Most members contain a single C-terminal SUN domain, but other architectures, including duplication of the domain, are also found. Figure 1A shows the architecture of those proteins containing a single C-terminal SUN domain and at least one predicted transmembrane domain. Molecular phylogeny and sequence alignment of their SUN domains are shown in Figure 1B and Figure S1 (available online). The nomenclature used for these proteins has not been systematic, even in cases where the orthologous relationships are clear. In mammals, at least five genes encoding SUN-domain proteins can be identified, and multiple isoforms of their transcripts have been documented. Three of these genes in mice have been previously named Sun1, Sun2 (Malone et al., 1999), and Sun3 (Crisp et al., 2006), but their human orthologs are known as Unc84A, Unc84B, and SUNC1, respectively. Two other SUN-domain genes were originally designated as rat sperm-associated antigen 4 (SPAG4) and SPAG4-like (SPAG4L). In this review, we use these existing names and also refer to these genes and their orthologs in mouse and human genomes as Sun4 and Sun5, respectively. The genome of Drosophila melanogaster contains two SUN-domain proteins, known as Klaroid and Giacomo, which play key roles in nuclear migration in the developing eye and sperm development, respectively. Arabidopsis thaliana also appears to have two SUN-domain proteins, which we here call SUN1 and SUN2. No obvious homologs had been found in the budding yeast Saccharomyces cerevisiae until Jaspersen and colleagues identified Mps3 as a SUN-domain protein (Jaspersen et al., 2006). Mps3 (also called Nep98) is essential for mitosis. It localizes prominently to the SPB, which is embedded in the NE, and can be more faintly detected along the NE, where it anchors telomeres and sites of DNA breaks and suppresses their recombination and repair, respectively (Antoniacci et al., 2007; Bupp et al., 2007; Jaspersen et al., 2002; Nishikawa et al., 2003; Oza et al., 2009; Schober et al., 2009). Like Sad1 in S. pombe, this SPB protein is conscripted for a role in chromosome dynamics during meiosis, although the details are different between budding and fission yeast, as described below. Mechanical Linkers of the Nucleus to the Cytoskeleton SUN-domain proteins are thought to be restricted to the inner membrane layer of the NE. They do not have obvious nuclear localization signals, so it seems most likely that this compartmentalization is determined by their interaction with components in the nucleoplasm. It is thought that their N-terminal regions reside on the nuclear face of the inner nuclear membrane (INM) and mediate such interactions. In some cases, these nucleoplasmic domains interact with nuclear lamin proteins, which form a filamentous meshwork, or lamina, underlying the INM (Haque et al., 2006). However, SUN-domain proteins are more widespread than lamins; for example, they are apparently ubiquitous in fungi, which lack lamins (Mans et al., 2004). Furthermore, SUN-domain protein can retain localization to the NE when their lamin interactions are disrupted (Hasan et al., 2006). Together, these observations indicate that the nuclear anchors for SUNdomain proteins must include components other than lamins. 598 Developmental Cell 17, November 17, 2009 ª2009 Elsevier Inc.

2 Figure 1. The SUN-Domain Protein Family (A) Primary structure of 18 SUN-domain proteins in S. pombe (Sp), C. elegans (Ce), human (Hs), mouse (Mm), D. melanogaster (Dm), A. thaliana (At), and S. cerevisiae (Sc). UniProt protein ID: SpSad1, Q09825; CeUNC-84, Q20745; CeSUN-1, Q20924; HsSUN1, O94901; HsSUN2, Q9UH99; MmSUN1, Q6B4H0; MmSUN2, Q6B4H2; HsSUN3, Q8TAQ9; HsSUN4, Q9NPE6; HsSUN5, Q8TC36; MmSUN3, Q5SS91; MmSUN4, Q9JJF2; MmSUN5, Q9DA32; Klaroid, A1Z6Q1; Giacomo, A5HL83. International Nucleotide Sequence Databases accession number: AtSUN1, BX830640; AtSUN2, BX822923; ScMps3, DQ SUN domains are indicated by green boxes. The number at the right of each bar indicates the total number of amino acid residues. Transmembrane sequences (TMs, magenta) were predicted by when obvious TMs were not predicted by this program, hydrophobic stretches were indicated by a method of Kyte and Doolittle (1982) in GENETYX-MAC (pinkish gray in CeSUN-1 and Klaroid). (B) An unrooted phylogenic tree of the 18 SUN domains generated by a nearest junction method in ClustalW. Members whose functions in meiosis have been demonstrated are shown in red. Meiotic functions have no obvious relationship with phylogeny. The C-terminal SUN domain (or domains) resides within the lumenal space between the inner and outer nuclear membranes, as indicated by immunoelectron microscopy (Hodzic et al., 2004). In animals, evidence indicates that the SUN domain mediates interactions with transmembrane proteins containing a weakly conserved motif termed the KASH domain, for the prototypical Klarsicht (Drosophila), ANC-1 (C. elegans), and Syne (mammals) homology (Apel et al., 2000; Mosley-Bishop et al., 1999; Starr and Fischer, 2005; Starr and Han, 2002). The KASH domain is a short (30 aa) C-terminal motif that lies adjacent to a transmembrane domain. The localization of some KASH-domain-containing proteins to the NE has been shown to depend on their physical interaction with SUNdomain proteins, and this attribute may be shared by all family members. Fungal proteins that interact with SUN-domain proteins, including fission yeast Kms1 and budding yeast Csm4, lack recognizable KASH domains and may therefore carry an analogous motif that can interact with the SUN domain; we will refer to these here as KASH-domain proteins for simplicity. Computational analysis indicates that the SUN domain is derived from a carbohydrate-binding motif known as a discoidin domain (Mans et al., 2004), suggesting that SUN domains might interact with glycans. However, no evidence has emerged to indicate that KASH-domain proteins are glycosylated. The N-terminal regions of KASH-domain proteins are highly variable and often enormous and are thought to protrude into the cytoplasm. They typically contain cytoskeletal linker motifs such as spectrin repeats or HOOK domains. Genetic and biochemical studies have indicated that these proteins interact with various cytoskeletal structures (reviewed in Starr and Fischer, 2005; Tzur et al., 2006; Wilhelmsen et al., 2006). The emergent view is that SUN-domain and KASH-domain proteins spanning the NE can link structures within the nucleus, such as the lamina and/or the chromosomes, to cytoskeletal components. Although SUN-KASH pairs were originally identified based on roles in nuclear migration and positioning (reviewed in Starr and Fischer, 2005; Tzur et al., 2006; Wilhelmsen et al., 2006; Burke and Roux, 2009 [this issue of Developmental Cell]), it is becoming clear that they contribute to other functions, particularly chromosome organization and dynamics. Partner Switching among SUN- and KASH-Domain Proteins During neuronal development in mice, Syne proteins, also known as nesprins, play key roles in anchoring myoblast nuclei at the Developmental Cell 17, November 17, 2009 ª2009 Elsevier Inc. 599

3 neuromuscular junction (Zhang et al., 2007). Recent work has shown that two SUN-domain proteins, Sun1 and Sun2, play partially redundant roles in this anchoring mechanism (Lei et al., 2009), suggesting that nesprins may be able to interact with multiple partners. However, how the SUN-KASH interactions might be regulated is currently unknown. In C. elegans, the SUN-domain protein UNC-84 is required for nuclear migration and positioning during development (Malone et al., 1999). This function is mediated by microtubules through interaction with the UNC-83 KASH-domain protein (McGee et al., 2006; Starr et al., 2001). UNC-84 also interacts with at least two other KASH-domain proteins: ANC-1, which is required to anchor the nucleus to actin filaments (Starr and Han, 2002), and KDP-1, which is involved in cell-cycle progression (McGee et al., 2009). KDP-1 requires SUN-1 for proper localization in the germline but also localizes to the NE in tissues where SUN- 1 is absent, suggesting the possibility of other SUN partners. SUN-1 also interacts with the KASH-domain protein ZYG-12 in embryonic and germline nuclei, where it plays key roles in centrosome attachment, nuclear positioning, and meiotic chromosome dynamics (Malone et al., 2003; Penkner et al., 2007; Zhou et al., 2009). These data indicate that a particular SUN domain can interact with multiple KASH partners (also reviewed in Tzur et al., 2006). This ability to switch partners may enrich the versatility of SUN and KASH proteins to contribute to distinct functions in different cell types. Active Players Driving Chromosome Movements The original biological functions attributed to SUN-KASHdomain pairs suggested that they mediate motion of the whole nucleus by interacting with dynamic cytoskeletal networks. Consistent with that notion, mammalian Sun1 and Sun2 were shown to be extremely immobile within the NE of HeLa cells (Lu et al., 2008). However, studies of meiotic nuclei have led to a much more dynamic view of SUN-KASH-domain proteins. Recent studies have extended the known roles for SUN-KASH proteins by showing that these bridges can link on the nucleoplasmic face to specific chromosomal loci to orchestrate chromosome organization and dynamics. Such intranuclear movements of chromosomes were first documented during meiotic prophase in the fission yeast S. pombe. During fission yeast meiosis, telomeres establish interactions with the NE that lead to their clustering adjacent to the SPB (Chikashige et al., 1994; reviewed in Hiraoka, 1998). Telomeres are also associated with the NE in vegetative cells, but their motion becomes much more dramatic during meiotic prophase as a consequence of the meiosisspecific expression of at least two proteins, Bqt1 and Bqt2 (Chikashige et al., 2006). Dispersed, NE-associated telomeres first move to cluster at the SPB, after which the entire nucleus is dragged by microtubules and associated motors back and forth along the length of the conjoined cells. The SPB and associated telomere cluster remain at the leading edge of this horsetail nucleus as it oscillates back and forth for most of meiotic prophase, which typically lasts about 2 hr. It is likely that the oscillation of the entire nucleus facilitates chromosome alignment to promote pairing of homologous chromosomes and at the same time agitates chromosomes to interrupt transient nonhomologous interactions. Meiotic telomere attachment to the SPB, clustering, and motion require two proteins that were initially identified based on their contributions to mitotic SPB function: Sad1, a founding member of the SUN family, and Kms1, which interacts with Sad1 and contains a C-terminal region resembling a KASH domain (Chikashige et al., 2006; Starr and Fischer, 2005). Upon cells entry into meiosis, Sad1-Kms1 complexes, which normally localize exclusively at the SPB (Miki et al., 2004), relocalize with telomeres at dispersed positions along the NE (Chikashige et al., 2006; Y. Chikashige and Y.H., unpublished data for Kms1). Eventually, these dispersed telomeres coalesce at the SPB (Chikashige et al., 2006; reviewed in Chikashige et al., 2007). Kms1 interacts with dynein microtubule motors in the cytoplasm (Goto et al., 2001; Miki et al., 2004). Notably, the Kms1-Sad1 complex is able to capture telomeres and tether them to the SPB in the absence of cytoplasmic dynein (Yamamoto et al., 1999), but the interaction of Kms1 with dynein is essential to drive oscillatory chromosome and nuclear motion (Chikashige et al., 2007). Taken together, these findings provided the first hint that SUN-KASH proteins can act as more than just a static linker between nuclear and cytoplasmic components they can move within nuclear membranes and establish de novo interactions with cytoskeletal components to actively choreograph chromosome dynamics (Figure 2). Interestingly, it has been demonstrated that in S. pombe, mutations that disrupt meiotic telomere attachment to the SPB (e.g., taz1, bqt1, and bqt4) lead to defects not only in homologous pairing but also in meiotic spindle formation (Tomita and Cooper, 2007; Chikashige et al., 2009). This may reflect an inability of SPB components to properly coalesce and to balance spindle forces if their connection to chromosomes is disrupted during meiotic prophase. Intranuclear Movements of Meiotic Chromosomes in Other Organisms As these discoveries were made in S. pombe, analogous stories began to emerge from studies of meiosis in other organisms. Classic cytological evidence dating from the early 1900s from widely divergent organisms revealed that chromosome ends associate with the nuclear membrane and frequently cluster, forming a meiosis-specific arrangement called the bouquet (reviewed in Scherthan, 2001; Zickler and Kleckner, 1998). This configuration occurs during early meiosis, roughly coincident with the establishment of homologous chromosome pairing and synapsis. Indeed, it is a reasonable assumption that largescale movements must be universally important during early meiosis, since when meiosis initiates, each chromosome is generally at some distance from its homologous partner. Large-scale movement of the entire nucleus during meiotic prophase has not been reported outside of S. pombe. However, intranuclear chromosome movement has been directly observed during meiotic prophase in other organisms, e.g., in rat spermatocytes (Parvinen and Söderström, 1976) and in budding yeast (Scherthan et al., 2007; Trelles-Sticken et al., 2005). The extent to which such movements are driven by active, motorbased mechanisms versus polymer diffusion is an area of active investigation. Recent discoveries in several organisms have revealed roles for SUN-domain proteins in meiotic organization and movement of chromosomes. These findings have come from studies in mammals, the nematode C. elegans, and the budding yeast S. cerevisiae. Evidence has recently emerged that these events 600 Developmental Cell 17, November 17, 2009 ª2009 Elsevier Inc.

4 Figure 2. SUN-KASH-Mediated Chromosome and Nuclear Movements (A C) Intranuclear movement of telomeres (A and B) and movement of the entire nucleus (C) in S. pombe. (D) Enlarged view showing components connecting telomeres to microtubules. Components in other organisms are summarized in Table 1. SUN and KASH proteins form a complex in the nuclear membranes. Telomeres or pairing centers are connected to the SUN protein upon appearance of meiosis-specific chromosomal connectors. The KASH protein interacts with cytoskeletal networks, microtubules, or actin filaments. SUN-KASH movements are mediated by actin in S. cerevisiae, whereas they are mediated by microtubules in S. pombe and C. elegans. involve a common cast of players: NE proteins containing SUN or KASH domains, meiotically expressed chromosomal connector proteins, and cytoskeletal proteins (Figure 2; summarized in Table 1). Mammals Electron microscopy of mammalian spermatocytes has shown that sites of chromosome attachment to the NE are marked by a densely staining structure known as an attachment plaque (Esponda and Giménez-Martín, 1972; Woollam et al., 1967). Immunolocalization of Sun1 and Sun2 from rat and mouse revealed that these proteins concentrate at sites of telomere attachment to the NE during meiotic prophase in spermatocytes, whereas they are distributed more uniformly along the NE in somatic cells (Ding et al., 2007; Schmitt et al., 2007). Immunoelectron microscopy confirmed that Sun2 is associated with the attachment plaque structure in rats (Schmitt et al., 2007). Sun1 was also shown by immunofluorescence to localize with telomeres during early meiosis in mouse spermatocytes. Although a function for Sun2 during meiosis has not yet been demonstrated, deletion of Sun1 in mice results in normal development but complete sterility in both males and females due to meiotic failure (Ding et al., 2007). Careful analysis of Sun1 knockout mice revealed defects in telomere-nuclear membrane attachment and homologous chromosome pairing. Thus, although Sun1 and Sun2 appear to have partially redundant functions in mammalian somatic cells (Lei et al., 2009), Sun1 is clearly essential for meiotic chromosome dynamics in the mouse. Although these studies of localization were carried out in fixed spermatocytes, it can be inferred that the protein moves within the NE during meiosis, because Sun1 and/or Sun2 relocalize from a diffuse distribution along the envelope to discrete foci (Ding et al., 2007; Schmitt et al., 2007). We do not yet know which specific cytoskeletal elements might be required for telomere movements during mammalian meiosis, although colchicine treatment has been shown to disrupt homologous chromosome synapsis (Tepperberg et al., 1999). Also unknown is the mechanism by which telomeres associate with Sun1 at the nuclear periphery. By analogy to S. pombe, itislikelythatmeiosis-specificlinker proteins are involved in establishing this nuclear organization. S. cerevisiae In the budding yeast S. cerevisiae, clustering of telomeres in meiosis is also observed during the pairing and recombination Developmental Cell 17, November 17, 2009 ª2009 Elsevier Inc. 601

5 Table 1. Components Connecting Chromosomes to the Cytoskeleton S. pombe C. elegans S. cerevisiae Mouse KASH-domain protein Kms1 ZYG-12 Csm4 unidentified SUN-domain protein Sad1 SUN-1 Mps3 Sun1, Sun2 Meiosis-specific connector Bqt1, Bqt2 HIM-8, ZIM proteins Ndj1 unidentified Chromosomal loci tethered telomeres pairing centers telomeres telomeres of homologous chromosomes (Rockmill and Roeder, 1998; Trelles-Sticken et al., 2000; Wu and Burgess, 2006). However, it appears that telomere attachment and clustering play a relatively minor role relative to what has been observed in fission yeast. Mutations that disrupt telomere attachment or movement result in defects in the distribution of meiotic recombination events and low levels of chromosome nondisjunction, but the progression of meiosis and the ability of homologs to pair and synapse are only subtly affected. In this organism, large-scale translocation of the entire nucleus is not observed, but continuous deformation of the nuclear surface has been reported (Hayashi et al., 1998). Recent studies have shown that intranuclear movements of telomeres are coupled with protrusions of the nuclear surface and that this motion is mediated through actin filaments (Conrad et al., 2008; Koszul et al., 2008; Scherthan et al., 2007; Trelles-Sticken et al., 2005). Some of these experiments have revealed the identity of the molecules that couple telomeres to the actin cytoskeleton. Ndj1 (also called Tam1) is a meiosis-specific protein that associates with telomeres (Chua and Roeder, 1997; Conrad et al., 1997). Loss of Ndj1 causes defects in telomere clustering and homologous recombination during meiosis (Conrad et al., 2008; Kosaka et al., 2008; Scherthan et al., 2007; Trelles-Sticken et al., 2000; Wanat et al., 2008; Wu and Burgess, 2006). In the absence of Ndj1, telomere movements are lost, while the nuclear membrane movements continue, indicating that Ndj1 connects the telomeres to the nuclear membranes. Mps3 is a SUN-domain protein that localizes to the SPB in vegetative cells (Jaspersen et al., 2002; Nishikawa et al., 2003). The N terminus of this protein, thought to be its nucleoplasmic domain, was found to interact directly with Ndj1 by yeast two-hybrid analysis and was subsequently shown to be required for normal meiotic telomere clustering and motion (Conrad et al., 2007). During meiotic prophase, Msp3 redistributes to multiple dynamic patches along the NE, corresponding to telomere attachment sites. Unlike in S. pombe, these patches do not stably coalesce at the SPB, but they do show a tendency to be in the same region of the membrane. In addition to Ndj1, another meiosis-specific protein, Csm4, is also necessary to connect telomeres to the NE. Loss of Csm4 also causes defects in telomere clustering and homologous recombination during meiosis (Conrad et al., 2008; Kosaka et al., 2008; Wanat et al., 2008). In the absence of Csm4, the motion of the nuclear membranes is also markedly reduced, suggesting that Csm4 couples force from actin cables to the nuclear membranes (Koszul et al., 2008). This suggests that Csm4 may be physically associated with the outer nuclear membrane, but high-resolution localization has not yet been reported. Csm4 has a single transmembrane domain in its C-terminal region, but it lacks an obvious KASH motif. However, as mentioned above, known SUN-domain-interacting proteins in fungi lack recognizable KASH domains, raising the possibility that Csm4 may interact directly with Mps3. Alternatively, a KASH-domain protein required for telomere movement during meiosis in S. cerevisiae may yet be discovered. C. elegans During meiosis in the nematode C. elegans, telomeres do not appear to play a key role in chromosome dynamics. Instead, it appears that nuclear reorganization functionally analogous to the meiotic bouquet is mediated by special regions on each chromosome known as homolog recognition regions, or pairing centers. These regions act in cis to promote pairing and synapsis of individual chromosomes (MacQueen et al., 2005; Villeneuve, 1994). The pairing center on each chromosome is bound by a single member of a family of four zinc-finger proteins known as HIM-8 and ZIM-1, -2, and -3. Each of these proteins is essential for proper meiosis, but none of them appears to play a critical role in somatic cells, at least individually. These proteins and their bound chromosome sites interact with the NE during meiotic prophase (Phillips and Dernburg, 2006; Phillips et al., 2005). The sites where they are attached to the NE become transiently enriched for the SUNand KASH-domain proteins SUN-1 and ZYG-12 during early meiotic prophase (Penkner et al., 2007; Sato et al., 2009). In mitotically dividing cells, these proteins are distributed homogenously around the nuclear surface. They play a critical role in tethering centrosomes to the nucleus in embryos (Malone et al., 2003)and in positioning germline nuclei (Zhou et al., 2009). The reorganization of SUN-1 and ZYG-12 during meiosis implies that these proteins are able to move within the NE, and their dynamic behavior has been directly observed through in vivo imaging (Sato et al., 2009; Penkner et al., 2009). Differences in their behavior in somatic versus meiotic cells appear to be controlled by several closely spaced phosphorylation sites within the N-terminal (nucleoplasmic) domain of SUN-1 (Penkner et al., 2009). Reduction or loss of SUN-1 or ZYG-12 results in aberrant synapsis between nonhomologous chromosomes (Penkner et al., 2007; Sato et al., 2009). ZYG-12 interacts with cytoplasmic dynein (Malone et al., 2003). Somewhat surprisingly, loss of dynein delays but does not abrogate pairing at the pairing center regions. Instead, loss of dynein results in failure of polymerization of the synaptonemal complex (SC), a protein structure formed between homologous chromosomes during meiosis. Microtubule-depolymerizing agents, including colchicine, inhibit homolog pairing, indicating that microtubules are required for pairing but that dynein is required at a discrete step, to enable chromosomes paired at their pairing center regions to progress to synapsis. One interpretation of this finding is that dynein allows chromosomes to attain more complete alignment, which in turn allows the SC to polymerize. However, in sun-1 mutants, SC polymerizes between nonaligned chromosomes (Penkner et al., 2007), indicating that alignment is not required for SC polymerization. 602 Developmental Cell 17, November 17, 2009 ª2009 Elsevier Inc.

6 Together, these data suggest that dynein is required for a discrete step in the process of pairing and synapsis that we have termed licensing. We envision that dynein may exert a force that attempts to separate paired chromosomes and that the resulting tension may be the cue that allows SC polymerization to initiate. Dynein, SUN-1, and ZYG-12 were recently shown to play additional roles in nuclear positioning in the C. elegans germline. Microtubules that appear to nucleate at the cell surface contact the NE and interact with ZYG-12 (Zhou et al., 2009). Notably, during the period in which homolog pairing and synapsis are taking place, the positioning of meiotic nuclei within the cells of the gonad becomes less uniformly central (A.F.D., unpublished data), perhaps reflecting an imbalance of forces acting at the NE as SUN-1 and ZYG-12 are conscripted to the sites of pairing centers rather than being distributed throughout the envelope as they are in earlier and later stages. Developmentally Regulated Chromosomal Connectors As described above, the S. pombe meiotic proteins Bqt1 and Bqt2 act to connect telomeres to a SUN-domain protein, Sad1, during telomere clustering. Sad1 is exclusively localized at the SPB but forms foci at the telomere in the presence of the connector proteins Bqt1 and Bqt2. In mice, uniform localization of Sun1 and Sun2 at the NE, as seen in somatic cells, is lost as these proteins accumulate at the telomeres in meiotic prophase. The de novo association of these SUN-domain proteins with telomeres during meiosis suggests the existence of meiosis-specific connector proteins or modifications to ubiquitously expressed proteins. To date, the proteins that act as meiosis-specific connectors between chromosomes and the NE, including Bqt1 and Bqt2, Ndj1, and the ZIM/HIM-8 family, have been unique to specific evolutionary lineages. This diversity seems to correspond to the weak conservation among the N-terminal regions of SUN-domain proteins, which are involved in these connections. This diversity of connector proteins may enable cells to remodel nuclear architecture not only during meiosis but during other developmental stages, by tethering specific chromosome loci to the NE through SUN-domain proteins. Sad1-mediated chromosome movements along the NE have been observed when meiotically arrested cells return to mitotic growth in S. pombe: while scattered centromeres recluster at the SPB during this return-to-growth process, multiple foci of Sad1 colocalized with Kms1 appear at the scattered centromeres (Goto et al., 2001). Thus, chromosome movements mediated by a SUN-KASH complex appear to act on the centromere as well. However, connector proteins that tether the centromere to the SPB during the return-to-growth process have not been identified. In C. elegans, a family of meiotic proteins connects the pairing centers of chromosomes to SUN-1 and ZYG-12. Taken together, these findings lead to the idea that developmentally regulated connector molecules link specific chromosomal loci, such as telomeres, centromeres, or pairing centers, to the SUN- KASH complex to accomplish specific differentiation programs by reorganizing chromosomes and the genes that they contain. An Evolutionary Origin for Chromosome Movements via Nuclear Envelope Attachment The ubiquitous observation of attachment of chromosomal loci to the nuclear membrane in meiosis likely reflects an ancient mechanism linking chromosomes to cytoskeletal structures (Ris, 1975) that is partially preserved in both the NE-associated SPB structure in fungi and also in more primitive protomitotic chromosome segregation mechanisms. Chromosomes in most of the familiar experimental organisms have kinetochores, a specialized structure formed on the centromere that is captured by spindle microtubules to segregate chromosomes. However, in some organisms that undergo a closed mitosis, kinetochorelike structures are formed at a centromere-ne attachment site, and chromosomes are segregated by microtubules that reach the nucleus from outside (Oakley and Dodge, 1974; Ris, 1975). Thus, chromosome movements along the NE may reflect a primitive way to segregate chromosomes during mitosis. This similarity in proposed function between mitotic centromeres and meiotic telomeres is intriguing in light of a recent proposal that eukaryotic centromeres may be derived from telomeres (Villasante et al., 2007). The evolutionary diversity of connector proteins may provide flexibility in tethering and moving different chromosomal loci, through the expression of cognate connector proteins, to reorganize intranuclear chromosome arrangement under developmental regulation. SUPPLEMENTAL DATA Supplemental Data include one figure and can be found with this article online at ACKNOWLEDGMENTS We thank A. Sato and Y. Chikashige for helpful comments on the manuscript. We also thank D. Starr, K. Zhou, and V. Jantsch for sharing information prior to publication. REFERENCES Antoniacci, L.M., Kenna, M.A., and Skibbens, R.V. (2007). The nuclear envelope and spindle pole body-associated Mps3 protein bind telomere regulators and function in telomere clustering. Cell Cycle 6, Apel, E.D., Lewis, R.M., Grady, R.M., and Sanes, J.R. (2000). Syne-1, a dystrophin- and Klarsicht-related protein associated with synaptic nuclei at the neuromuscular junction. J. Biol. 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