Phosphorylation of luminal region of the SUN-domain protein Mps3 promotes nuclear envelope localization during meiosis

During meiosis, protein ensembles in the nuclear envelope (NE) containing SUN- and KASH-domain proteins, called linker nucleocytoskeleton and cytoskeleton (LINC) complex, promote the chromosome motion. Yeast SUN-domain protein, Mps3, forms multiple meiosis-specific ensembles on NE, which show dynamic localisation for chromosome motion; however, the mechanism by which these Mps3 ensembles are formed during meiosis remains largely unknown. Here, we showed that the cyclin-dependent protein kinase (CDK) and Dbf4-dependent Cdc7 protein kinase (DDK) regulate meiosis-specific dynamics of Mps3 on NE, particularly by mediating the resolution of Mps3 clusters and telomere clustering. We also found that the luminal region of Mps3 juxtaposed to the inner nuclear membrane is required for meiosis-specific localisation of Mps3 on NE. Negative charges introduced by meiosis-specific phosphorylation in the luminal region of Mps3 alter its interaction with negatively charged lipids by electric repulsion in reconstituted liposomes. Phospho-mimetic substitution in the luminal region suppresses the localisation of Mps3 via the inactivation of CDK or DDK. Our study revealed multi-layered phosphorylation-dependent regulation of the localisation of Mps3 on NE for meiotic chromosome motion and NE remodelling.

Chromosome oscillation promotes Aurora A–dependent Hec1 phosphorylation and mitotic fidelity

Most cancer cells show chromosomal instability, a condition where chromosome missegregation occurs frequently. We found that chromosome oscillation, an iterative chromosome motion during metaphase, is attenuated in cancer cell lines. We also found that metaphase phosphorylation of Hec1 at serine 55, which is mainly dependent on Aurora A on the spindle, is reduced in cancer cell lines. The Aurora A–dependent Hec1-S55 phosphorylation level was regulated by the chromosome oscillation amplitude and vice versa: Hec1-S55 and -S69 phosphorylation by Aurora A is required for efficient chromosome oscillation. Furthermore, enhancement of chromosome oscillation reduced the number of erroneous kinetochore–microtubule attachments and chromosome missegregation, whereas inhibition of Aurora A during metaphase increased such errors. We propose that Aurora A–mediated metaphase Hec1-S55 phosphorylation through chromosome oscillation, together with Hec1-S69 phosphorylation, ensures mitotic fidelity by eliminating erroneous kinetochore–microtubule attachments. Attenuated chromosome oscillation and the resulting reduced Hec1-S55 phosphorylation may be a cause of CIN in cancer cell lines.

SWR1-independent association of H2A.Z to the LINC complex promotes meiotic chromosome motion

ABSTRACTThe H2A.Z histone variant is deposited into chromatin by the SWR1 complex affecting multiple aspects of meiosis. Here we describe a SWR1-independent localization of H2A.Z at meiotic telomeres and the centrosome. We demonstrate that H2A.Z colocalizes and interacts with Mps3, the SUN component of the LINC complex that spans the nuclear envelope and links meiotic telomeres to the cytoskeleton promoting meiotic chromosome movement. H2A.Z also interacts with the meiosis-specific Ndj1 protein that anchors telomeres to the nuclear periphery via Mps3. Telomeric localization of H2A.Z depends on Ndj1 and the N-terminal domain of Mps3. Although telomeric attachment to the nuclear envelope is maintained in the absence of H2A.Z, the distribution of Mps3 is altered. The velocity of chromosome movement during meiotic prophase I is reduced in the htz1Δ mutant lacking H2A.Z, but it is unaffected in swr1Δ cells. We reveal that H2A.Z is an additional LINC-associated factor that contributes to promote telomere-driven chromosome motion critical for error-free gametogenesis.

Structural view of the yeast Dam1 complex, a ring-shaped molecular coupler for the dynamic microtubule end

In a dividing eukaryotic cell, proper chromosome segregation requires the dynamic yet persistent attachment of kinetochores to spindle microtubules. In the budding yeast Saccharomyces cerevisiae, this function is especially crucial because each kinetochore is attached to a single microtubule; consequently, loss of attachment could lead to unrecoverable chromosome loss. The highly specialized heterodecameric Dam1 protein complex achieves this coupling by assembling into a microtubule-encircling ring that glides near the end of the dynamic microtubule to mediate chromosome motion. In recent years, we have learned a great deal about the structural properties of the Dam1 heterodecamer, its mechanism of self-assembly into rings, and its tethering to the kinetochore by the elongated Ndc80 complex. The most remarkable progress has resulted from defining the fine structures of helical bundles within Dam1 heterodecamer. In this review, we critically analyze structural observations collected by diverse approaches with the goal of obtaining a unified view of Dam1 ring architecture. A considerable consistency between different studies supports a coherent model of the circular core of the Dam1 ring. However, there are persistent uncertainties about the composition of ring protrusions and flexible extensions, as well as their roles in mediating ring core assembly and interactions with the Ndc80 complex and microtubule.

Central-spindle microtubules are strongly coupled to chromosomes during both anaphase A and anaphase B

Spindle microtubules, whose dynamics vary over time and at different locations, cooperatively drive chromosome segregation. Measurements of microtubule dynamics and spindle ultrastructure can provide insight into the behaviors of microtubules, helping elucidate the mechanism of chromosome segregation. Much work has focused on the dynamics and organization of kinetochore microtubules, that is, on the region between chromosomes and poles. In comparison, microtubules in the central-spindle region, between segregating chromosomes, have been less thoroughly characterized. Here, we report measurements of the movement of central-spindle microtubules during chromosome segregation in human mitotic spindles and Caenorhabditis elegans mitotic and female meiotic spindles. We found that these central-spindle microtubules slide apart at the same speed as chromosomes, even as chromosomes move toward spindle poles. In these systems, damaging central-spindle microtubules by laser ablation caused an immediate and complete cessation of chromosome motion, suggesting a strong coupling between central-spindle microtubules and chromosomes. Electron tomographic reconstruction revealed that the analyzed anaphase spindles all contain microtubules with both ends between segregating chromosomes. Our results provide new dynamical, functional, and ultrastructural characterizations of central-spindle microtubules during chromosome segregation in diverse spindles and suggest that central-spindle microtubules and chromosomes are strongly coupled in anaphase.

Spatiotemporal control of mitotic exit during anaphase by an aurora B-Cdk1 crosstalk

According to the prevailing ‘clock’ model, chromosome decondensation and nuclear envelope reformation when cells exit mitosis are byproducts of Cdk1 inactivation at the metaphase-anaphase transition, controlled by the spindle assembly checkpoint. However, mitotic exit was recently shown to be a function of chromosome separation during anaphase, assisted by a midzone Aurora B phosphorylation gradient - the ‘ruler’ model. Here we found that Cdk1 remains active during anaphase due to ongoing APC/CCdc20- and APC/CCdh1-mediated degradation of B-type Cyclins in Drosophila and human cells. Failure to degrade B-type Cyclins during anaphase prevented mitotic exit in a Cdk1-dependent manner. Cyclin B1-Cdk1 localized at the spindle midzone in an Aurora B-dependent manner, with incompletely separated chromosomes showing the highest Cdk1 activity. Slowing down anaphase chromosome motion delayed Cyclin B1 degradation and mitotic exit in an Aurora B-dependent manner. Thus, a crosstalk between molecular ‘rulers’ and ‘clocks’ licenses mitotic exit only after proper chromosome separation.

A New Role for Telomerase in Promoting Meiotic Homolog Pairing Fidelity

ABSTRACTCytoskeletal forces acting upon telomeres promote active chromosome motion needed to pair homologous chromosomes during meiosis. The necessary components that allow this force to be applied to telomeres is still unclear, as are the roles of this motion and whether motion is needed primarily for increasing collisions of homologous regions, testing homolog pairing fidelity, or some other role. Here, we show a novel role for telomerase, previously known to be responsible for telomeric end replication, in anchoring telomeres to the nuclear envelope (NE) to provide proper transmission of cytoskeletal forces during meiosis. Reduction in telomerase function in Saccharomyces cerevisiae results in a dramatic decrease in the frequency of high velocity “pulls” resulting in earlier homolog synapsis and increased recombination. These observations are consistent with a model in which telomeric cytoskeletal engagement ensures homolog pairing fidelity by pulling apart improperly associated regions whereas general chromosomal motion aids in increasing homologous contacts.