Immediate visualization of recombination events and chromosome segregation defects in fission yeast meiosis

During mitosis, the spindle assembly checkpoint (SAC) inhibits the Cdc20-activated anaphase-promoting complex/cyclosome (APC/CCdc20), which promotes protein degradation, and delays anaphase onset to ensure accurate chromosome segregation. However, the SAC function in meiotic anaphase regulation is poorly understood. Here, we examined the SAC function in fission yeast meiosis. As in mitosis, a SAC factor, Mad2, delayed anaphase onset via Slp1 (fission yeast Cdc20) when chromosomes attach to the spindle improperly. However, when the SAC delayed anaphase I, the interval between meiosis I and II shortened. Furthermore, anaphase onset was advanced and the SAC effect was reduced at meiosis II. The advancement of anaphase onset depended on a meiosis-specific, Cdc20-related factor, Fzr1/Mfr1, which contributed to anaphase cyclin decline and anaphase onset and was inefficiently inhibited by the SAC. Our findings show that impacts of SAC activation are not confined to a single division at meiosis due to meiosis-specific APC/C regulation, which has probably been evolved for execution of two meiotic divisions.

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Cohesins Determine the Attachment Manner of Kinetochores to Spindle Microtubules at Meiosis I in Fission Yeast

Molecular and Cellular Biology ◽

10.1128/mcb.23.11.3965-3973.2003 ◽

2003 ◽

Vol 23 (11) ◽

pp. 3965-3973 ◽

Cited By ~ 76

Author(s):

Shihori Yokobayashi ◽

Masayuki Yamamoto ◽

Yoshinori Watanabe

Keyword(s):

Fission Yeast ◽

Chromosome Segregation ◽

Sister Chromatid ◽

Specific Requirement ◽

Spindle Poles ◽

Chromatid Cohesion ◽

Spindle Microtubules ◽

Yeast Meiosis ◽

Meiosis I ◽

Random Division

ABSTRACT During mitosis, sister kinetochores attach to microtubules that extend to opposite spindle poles (bipolar attachment) and pull the chromatids apart at anaphase (equational segregation). A multisubunit complex called cohesin, including Rad21/Scc1, plays a crucial role in sister chromatid cohesion and equational segregation at mitosis. Meiosis I differs from mitosis in having a reductional pattern of chromosome segregation, in which sister kinetochores are attached to the same spindle (monopolar attachment). During meiosis, Rad21/Scc1 is largely replaced by its meiotic counterpart, Rec8. If Rec8 is inactivated in fission yeast, meiosis I is shifted from reductional to equational division. However, the reason rec8Δ cells undergo equational rather than random division has not been clarified; therefore, it has been unclear whether equational segregation is due to a loss of cohesin in general or to a loss of a specific requirement for Rec8. We report here that the equational segregation at meiosis I depends on substitutive Rad21, which relocates to the centromeres if Rec8 is absent. Moreover, we demonstrate that even if sufficient amounts of Rad21 are transferred to the centromeres at meiosis I, thereby establishing cohesion at the centromeres, rec8Δ cells never recover monopolar attachment but instead secure bipolar attachment. Thus, Rec8 and Rad21 define monopolar and bipolar attachment, respectively, at meiosis I. We conclude that cohesin is a crucial determinant of the attachment manner of kinetochores to the spindle microtubules at meiosis I in fission yeast.

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Live observation of fission yeast meiosis in recombination-deficient mutants

Journal of Cell Science ◽

10.1242/jcs.114.15.2843 ◽

2001 ◽

Vol 114 (15) ◽

pp. 2843-2853 ◽

Cited By ~ 7

Author(s):

Monika Molnar ◽

Jürg Bähler ◽

Jürg Kohli ◽

Yasushi Hiraoka

Keyword(s):

Fission Yeast ◽

Chromosome Segregation ◽

Meiotic Division ◽

Chiasma Formation ◽

Homologous Chromosomes ◽

Yeast Meiosis ◽

Mutant Cells ◽

Random Level ◽

Meiosis I ◽

Chromosome Disjunction

Regular segregation of homologous chromosomes during meiotic divisions is essential for the generation of viable progeny. In recombination-proficient organisms, chromosome disjunction at meiosis I generally occurs by chiasma formation between the homologs (chiasmate meiosis). We have studied meiotic stages in living rec8 and rec7 mutant cells of fission yeast, with special attention to prophase and the first meiotic division. Both rec8 and rec7 are early recombination mutants, and in rec7 mutants, chromosome segregation at meiosis I occurs without any recombination (achiasmate meiosis). Both mutants showed distinct irregularities in nuclear prophase movements. Additionally, rec7 showed an extended first division of variable length and with single chromosomes changing back and forth between the cell poles. Two other early recombination deficient mutants (rec14 and rec15) showed very similar phenotypes to rec7 during the first meiotic division, and the fidelity of achiasmate chromosome segregation slightly exceeded the expected random level. We discuss possible regulatory mechanisms of fission yeast to deal with achiasmate chromosome segregation.

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Immediate visualization of recombination events and chromosome segregation defects in fission yeast meiosis

10.1101/458398 ◽

2018 ◽

Cited By ~ 1

Author(s):

Dmitriy Li ◽

Marianne Rocl ◽

Raif Yuecel ◽

Alexander Lorenz

Keyword(s):

Fission Yeast ◽

Chromosome Segregation ◽

High Throughput ◽

Meiotic Recombination ◽

Recombination Frequency ◽

Genetic Interval ◽

Chromosome Iii ◽

Yeast Meiosis ◽

Chromosome I ◽

High Throughput Screens

AbstractSchizosaccharomyces pombe, also known as fission yeast, is an established model for studying chromosome biological processes. Over the years research employing fission yeast has made important contributions to our knowledge about chromosome segregation during meiosis, as well as meiotic recombination and its regulation. Quantification of meiotic recombination frequency is not a straightforward undertaking, either requiring viable progeny for a genetic plating assay, or relying on laborious Southern blot analysis of recombination intermediates. Neither of these methods lends itself to high-throughput screens to identify novel meiotic factors. Here, we establish visual assays novel to Sz. pombe for characterizing chromosome segregation and meiotic recombination phenotypes. Genes expressing red, yellow, and/or cyan fluorophores from spore-autonomous promoters have been integrated into the fission yeast genomes, either close to the centromere of chromosome I to monitor chromosome segregation, or on the arm of chromosome III to form a genetic interval at which recombination frequency can be determined. The visual recombination assay allows straightforward and immediate assessment of the genetic outcome of a single meiosis by epi-fluorescence microscopy without requiring tetrad dissection. We also demonstrate that the recombination frequency analysis can be automatized by utilizing imaging flow cytometry to enable high-throughput screens. These assays have several advantages over traditional methods for analysing meiotic phenotypes.

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Linear elements are stable structures along the chromosome axis in fission yeast meiosis

Chromosoma ◽

10.1007/s00412-021-00757-w ◽

2021 ◽

Author(s):

Da-Qiao Ding ◽

Atsushi Matsuda ◽

Kasumi Okamasa ◽

Yasushi Hiraoka

Keyword(s):

Fission Yeast ◽

Strand Break ◽

Wide Field ◽

Structured Illumination ◽

Structured Illumination Microscopy ◽

Meiotic Chromosomes ◽

Linear Elements ◽

Yeast Meiosis ◽

Chromosome Axis ◽

Stable Structures

AbstractThe structure of chromosomes dramatically changes upon entering meiosis to ensure the successful progression of meiosis-specific events. During this process, a multilayer proteinaceous structure called a synaptonemal complex (SC) is formed in many eukaryotes. However, in the fission yeast Schizosaccharomyces pombe, linear elements (LinEs), which are structures related to axial elements of the SC, form on the meiotic cohesin-based chromosome axis. The structure of LinEs has been observed using silver-stained electron micrographs or in immunofluorescence-stained spread nuclei. However, the fine structure of LinEs and their dynamics in intact living cells remain to be elucidated. In this study, we performed live cell imaging with wide-field fluorescence microscopy as well as 3D structured illumination microscopy (3D-SIM) of the core components of LinEs (Rec10, Rec25, Rec27, Mug20) and a linE-binding protein Hop1. We found that LinEs form along the chromosome axis and elongate during meiotic prophase. 3D-SIM microscopy revealed that Rec10 localized to meiotic chromosomes in the absence of other LinE proteins, but shaped into LinEs only in the presence of all three other components, the Rec25, Rec27, and Mug20. Elongation of LinEs was impaired in double-strand break-defective rec12− cells. The structure of LinEs persisted after treatment with 1,6-hexanediol and showed slow fluorescence recovery from photobleaching. These results indicate that LinEs are stable structures resembling axial elements of the SC.

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Novel Genes Required for Meiotic Chromosome Segregation Are Identified by a High-Throughput Knockout Screen in Fission Yeast

Current Biology ◽

10.1016/j.cub.2005.07.059 ◽

2005 ◽

Vol 15 (18) ◽

pp. 1663-1669 ◽

Cited By ~ 67

Author(s):

Juraj Gregan ◽

Peter K. Rabitsch ◽

Benjamin Sakem ◽

Ortansa Csutak ◽

Vitaly Latypov ◽

...

Keyword(s):

Fission Yeast ◽

Chromosome Segregation ◽

High Throughput ◽

Meiotic Chromosome ◽

Novel Genes

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Srs2 helicase prevents the formation of toxic DNA damage during late prophase I of yeast meiosis

10.1101/518035 ◽

2019 ◽

Cited By ~ 2

Author(s):

Hiroyuki Sasanuma ◽

Hana Subhan M. Sakurai ◽

Yuko Furihata ◽

Kiran Challa ◽

Lira Palmer ◽

...

Keyword(s):

Dna Damage ◽

Chromosome Segregation ◽

Dna Recombination ◽

Dna Helicase ◽

Aggregate Formation ◽

Late Prophase ◽

Prophase I ◽

Early Protein ◽

Srs2 Helicase ◽

Yeast Meiosis

AbstractProper repair of double-strand breaks (DSBs) is key to ensure proper chromosome segregation. In this study, we found that the deletion of the SRS2 gene, which encodes a DNA helicase necessary for the control of homologous recombination, induces aberrant chromosome segregation during budding yeast meiosis. This abnormal chromosome segregation in srs2 cells accompanies the formation of a novel DNA damage induced during late meiotic prophase-I. The damage may contain long stretches of single-stranded DNAs (ssDNAs), which lead to aggregate formation of a ssDNA binding protein, RPA, and a RecA homolog, Rad51, as well as other recombination proteins inside of the nuclei. The Rad51 aggregate formation in the srs2 mutant depends on the initiation of meiotic recombination and occurs in the absence of chromosome segregation. Importantly, as an early recombination intermediate, we detected a thin bridge of Rad51 between two Rad51 foci or among the foci in the srs2 mutant, which is rarely seen in wild type. These might be cytological manifestation of the connection of two DSB ends and multi-invasion. The DNA damage with Rad51 aggregates in the srs2 mutant is passed through anaphase-I and -II, suggesting the absence of DNA damage-induced cell-cycle arrest after the pachytene stage. We propose that Srs2 helicase resolves early protein-DNA recombination intermediates to suppress the formation of aberrant lethal DNA damage during late prophase-I.

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A microtubule polymerase cooperates with the kinesin-6 motor and a microtubule cross-linker to promote bipolar spindle assembly in the absence of kinesin-5 and kinesin-14 in fission yeast

Molecular Biology of the Cell ◽

10.1091/mbc.e17-08-0497 ◽

2017 ◽

Vol 28 (25) ◽

pp. 3647-3659 ◽

Cited By ~ 23

Author(s):

Masashi Yukawa ◽

Tomoki Kawakami ◽

Masaki Okazaki ◽

Kazunori Kume ◽

Ngang Heok Tang ◽

...

Keyword(s):

Fission Yeast ◽

Chromosome Segregation ◽

Spindle Pole Body ◽

Spindle Assembly ◽

Spindle Pole ◽

Temperature Sensitive ◽

Bipolar Spindle ◽

Pole Body ◽

Cross Linker ◽

Spindle Elongation

Accurate chromosome segregation relies on the bipolar mitotic spindle. In many eukaryotes, spindle formation is driven by the plus-end–directed motor kinesin-5 that generates outward force to establish spindle bipolarity. Its inhibition leads to the emergence of monopolar spindles with mitotic arrest. Intriguingly, simultaneous inactivation of the minus-end–directed motor kinesin-14 restores spindle bipolarity in many systems. Here we show that in fission yeast, three independent pathways contribute to spindle bipolarity in the absence of kinesin-5/Cut7 and kinesin-14/Pkl1. One is kinesin-6/Klp9 that engages with spindle elongation once short bipolar spindles assemble. Klp9 also ensures the medial positioning of anaphase spindles to prevent unequal chromosome segregation. Another is the Alp7/TACC-Alp14/TOG microtubule polymerase complex. Temperature-sensitive alp7cut7pkl1 mutants are arrested with either monopolar or very short spindles. Forced targeting of Alp14 to the spindle pole body is sufficient to render alp7cut7pkl1 triply deleted cells viable and promote spindle assembly, indicating that Alp14-mediated microtubule polymerization from the nuclear face of the spindle pole body could generate outward force in place of Cut7 during early mitosis. The third pathway involves the Ase1/PRC1 microtubule cross-linker that stabilizes antiparallel microtubules. Our study, therefore, unveils multifaceted interplay among kinesin-dependent and -independent pathways leading to mitotic bipolar spindle assembly.

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