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Spindle Pole Body (spindle + pole_body)
Selected AbstractsCut1/separase C-terminus affects spindle pole body positioning in interphase of fission yeast: pointed nuclear formationGENES TO CELLS, Issue 11 2002Takahiro Nakamura Background: The separase-securin complex is required for anaphase. Separase activated by securin destruction cleaves the cohesin subunit Scc1/Rad21 enriched in kinetochores. Fission yeast Cut1/separase resides in interphase cytoplasm and mobilizes to the spindle and the spindle pole bodies (SPBs) in mitosis, while Cut2/securin remains in the nucleus from interphase to metaphase, and temporarily locates at the short spindle. Results: We here report a novel SPB-led dynamic nuclear movement in fission yeast, when the Cut1 C-terminal fragment is over-expressed. The tip of the pointed nucleus contained both SPB and centromeric DNA, and rapidly moved along the bundled cytoplasmic microtubules. The same pointed nucleus was produced when the human separase C-fragment was over-expressed. The pointed nuclear formation did not require the protease site of separase, but required the conserved C-terminus and a microtubule- and kinetochore-binding protein Mtc1/Alp14, a homologue of frog XMAP215 and budding yeast Stu2. The movement-inducing C-fragment should be cytoplasmic, as the pointed nucleus was abolished when the fragment contained the NLS (nuclear localization signal). Conclusions: Overproduced separase C-fragment abolishes correct SPB-positioning in interphase. Resulting pointed nuclear formation (alternatively called ,pigtail movement') requires cytoplasmic microtubules and Mtc1/Alp14. [source] Localization of gene products using a chromosomally tagged GFP-fusion library in the fission yeast Schizosaccharomyces pombeGENES TO CELLS, Issue 2 2009Aki Hayashi We constructed a library of chromosomally-tagged green fluorescent protein (GFP) fusions in the fission yeast Schizosaccharomyces pombe. This library contains 1058 strains. In each strain, the coding sequence of GFP is integrated at the 3,-end of a particular chromosomal ORF such that the full-length GFP fusion construct is expressed under the control of the original promoter. Integration of the GFP coding sequence at the authentic chromosomal location of each gene was confirmed by PCR. Microscopic screening of these strains detected sufficient levels of GFP signal in 710 strains and allowed assignment of these GFP-fusion gene products with their intracellular localization: 374 proteins were localized in the nucleus, 65 proteins in the nucleolus, 34 proteins at the nuclear periphery, 27 proteins at the plasma membrane and cytoplasmic membranous structures, 24 proteins at the spindle pole body and microtubules, 92 proteins at cytoplasmic structures, and 94 proteins were uniformly distributed throughout the cytoplasm. [source] Modulation of Alp4 function in Schizosaccharomyces pombe induces novel phenotypes that imply distinct functions for nuclear and cytoplasmic ,-tubulin complexesGENES TO CELLS, Issue 4 2006Hirohisa Masuda The ,-tubulin complex acts as a nucleation unit for microtubule assembly. It remains unknown, however, how spatial and temporal regulation of the complex activity affects microtubule-mediated cellular processes. Alp4 is one of the essential components of the S. pombe,-tubulin complex. We show here that overproduction of a carboxy-terminal form of Alp4 (Alp4C) and its derivatives tagged to a nuclear localization signal or to a nuclear export signal affect localization of ,-tubulin complexes and induces novel phenotypes that reflect distinct functions of nuclear and cytoplasmic ,-tubulin complexes. Nuclear Alp4C induces a Wee1-dependent G2 delay, reduces the levels of the ,-tubulin complex at the spindle pole body, and results in defects in mitotic progression including spindle assembly, cytoplasmic microtubule disassembly, and chromosome segregation. In contrast, cytoplasmic Alp4C induces oscillatory nuclear movement and affects levels of cell polarity markers, Bud6 and Tip1, at the cell ends. These results demonstrate that regulation of nuclear ,-tubulin complex activity is essential for cell cycle progression through the G2/M boundary and M phase, whereas regulation of cytoplasmic ,-tubulin complex activity is important for nuclear positioning and cell polarity control during interphase. [source] The carboxy-terminus of Alp4 alters microtubule dynamics to induce oscillatory nuclear movement led by the spindle pole body in Schizosaccharomyces pombeGENES TO CELLS, Issue 4 2006Hirohisa Masuda Alp4 is an essential component of the S. pombe,-tubulin complex. Overproduction of the carboxy-terminus of Alp4 induces oscillatory nuclear movement led by the spindle pole body (SPB). The movement is not dependent on cytoplasmic dynein dhc1, or kinesin-related proteins pkl1 and klp2. Rates of SPB movement correlate with elongation rates of microtubules (MTs) extending backwards from the moving SPB (backward-extending MTs), showing that pushing forces exerted by backward-extending MTs move the nucleus via the SPB. These backward-extending MTs are more stable than those of control cells and, thus, are able to push the SPB further towards the cell end, inducing nuclear oscillation with larger amplitudes than in control cells. SPB movement is biased towards the new end of the cell where levels of the CLIP170 homolog Tip1 increase, suggesting that the movement is related to MT-mediated cell polarity control. These results demonstrate that the carboxy-terminus of Alp4 alters MT dynamics and induces nuclear oscillation by modulating a nuclear positioning mechanism based on the balance of MT pushing forces, and suggest that regulation of ,-tubulin complex activity is important for controlling MT dynamics and nuclear positioning. [source] Cut1/separase C-terminus affects spindle pole body positioning in interphase of fission yeast: pointed nuclear formationGENES TO CELLS, Issue 11 2002Takahiro Nakamura Background: The separase-securin complex is required for anaphase. Separase activated by securin destruction cleaves the cohesin subunit Scc1/Rad21 enriched in kinetochores. Fission yeast Cut1/separase resides in interphase cytoplasm and mobilizes to the spindle and the spindle pole bodies (SPBs) in mitosis, while Cut2/securin remains in the nucleus from interphase to metaphase, and temporarily locates at the short spindle. Results: We here report a novel SPB-led dynamic nuclear movement in fission yeast, when the Cut1 C-terminal fragment is over-expressed. The tip of the pointed nucleus contained both SPB and centromeric DNA, and rapidly moved along the bundled cytoplasmic microtubules. The same pointed nucleus was produced when the human separase C-fragment was over-expressed. The pointed nuclear formation did not require the protease site of separase, but required the conserved C-terminus and a microtubule- and kinetochore-binding protein Mtc1/Alp14, a homologue of frog XMAP215 and budding yeast Stu2. The movement-inducing C-fragment should be cytoplasmic, as the pointed nucleus was abolished when the fragment contained the NLS (nuclear localization signal). Conclusions: Overproduced separase C-fragment abolishes correct SPB-positioning in interphase. Resulting pointed nuclear formation (alternatively called ,pigtail movement') requires cytoplasmic microtubules and Mtc1/Alp14. [source] Calcineurin is implicated in the regulation of the septation initiation network in fission yeastGENES TO CELLS, Issue 10 2002Yabin Lu Background: In fission yeast, calcineurin has been implicated in cytokinesis because calcineurin-deleted cells form multiple septa and cell separation is impeded. However, this mechanism remains unclear. Results: We screened for mutations that confer syn-thetic lethality with calcineurin deletion and isolated a mutant, its10-1/cdc7-i10, a novel allele of the cdc7+ gene involved in the septation initiation network (SIN). The mutation created a termination codon, resulting in the truncation of Cdc7 by 162 amino acids, which is not localized in the spindle pole body. Following treatment with the immune suppressive drug FK506, cdc7-i10 and the original cdc7-24 mutant cells showed highly elongated multinuclear morphology with few visible septa, closely resembling the phenotype at the restrictive temperature. Other SIN mutants, cdc11, spg1, sid2 and mob1 showed similar phenotypes following FK506 treatment. Consistent with this, expression of the constitutively active calcineurin suppressed the growth defects and septum initiation deficiency of these SIN mutants at the restrictive temperature. Moreover, electron microscopy revealed that calcineurin-deleted cells had very thick multiple septa which were partially and ectopically formed. Conclusion: These results suggest that calcineurin is involved in the regulation of the SIN pathway, and is required for the proper formation and maturation of the septum in fission yeast. [source] The ,-tubulin complex protein Alp4 provides a link between the metaphase checkpoint and cytokinesis in fission yeastGENES TO CELLS, Issue 4 2002Leah Vardy Background:, The progression of cytokinesis requires cyclin B destruction by the anaphase promoting complex (APC/C) and, in fission yeast, activation of the septation initiation network (SIN) is also essential. The ,-tubulin complex (,-TuC) localizes to the centrosome throughout the cell cycle and is directly involved in the organization of the mitotic spindle. Results:, We have previously shown that the mutant defective in alp4+ (Spc97/GCP2) displays bipolar spindle defects due to a failure in the recruitment of the ,-TuC on to the spindle pole body (SPB, the centrosome equivalent). Here we show that in these mutants the Mad2 checkpoint is activated, yet septation proceeds due to the untimely activation of the SIN. The Sid1 kinase, the downstream effector of the SIN, is recruited prematurely to both, instead of only one, of the SPBs, which triggers septation despite the presence of monopolar spindles. Remarkably, cyclin B levels, which would normally have declined, remain high at the SPB in septated mutant cells. Conclusions:, We propose a novel role of the ,-TuC in inhibiting activation of the SIN until cyclin B is destroyed. Given the ubiquitous existence of the ,-TuC, this mechanism may be conserved throughout evolution and function to couple cytokinesis to mitotic exit. [source] Dynamic distribution of BIMGPP1 in living hyphae of Aspergillus indicates a novel role in septum formationMOLECULAR MICROBIOLOGY, Issue 5 2002H. Fox Summary Mutation of bimG, the major protein phosphatase 1 gene in Aspergillus nidulans, causes multiple cell cycle and hyphal growth defects that are associated with overphosphorylation of subcellular components. We have used functional translational fusions with the green fluorescent protein (GFP) to show that BIMG has at least four discrete locations within growing hyphae. Three of these locations, the hyphal tip, the spindle pole body and the nucleus, correlate with previously known requirements for bimGPP1 in mitosis and hyphal growth and are highly dynamic. BIMG-GFP in the hyphal tip seemed to be associated with the plasma membrane and formed a collar of fluorescence within the apical dome. The distribution of nuclear BIMG-GFP varied depending on nutritional conditions; on poor medium, it concentrated more in the nucleolus than in the nucleoplasm, whereas on rich medium, it was more evenly distributed between the two nuclear regions. The association of BIMG-GFP with developing septa was transient, and we present evidence that BIMG phosphatase plays a direct role in septum formation, distinct from its role in mitosis. We conclude that, by being physically present at several sites, the BIMG phosphatase has roles in multiple cellular processes. [source] | ||||||||||