EVOLUTION OF NUCLEAR DIVISION STRATEGIES WITHIN THE FISSION YEAST CLADE

EVOLUTION OF NUCLEAR DIVISION STRATEGIES WITHIN THE FISSION YEAST CLADE YAM QIU XIA, CANDICE (B.Sc. (Hons), NTU) A THESIS SUBMITTED FOR THE DOCTOR OF PHILOSPOHY TEMASEK LIFE SCIENCES LABORATORY NATIONAL UNIVERSITY OF SINGAPORE 2013 I DECLARATION I hereby declare that this thesis is my original work and it has been written by me in its entirety. I have duly acknowledged all the sources of information which have been used in the thesis. This thesis has also not been submitted for any degree in any university previously. Yam Qiu Xia, Candice 22 Aug 2013 II Acknowledgements I would like to thank the people around me who have supported and helped me much during my graduate studies. I am very grateful to my supervisor Snezhka Oliferenko for giving me the opportunity to pursue my graduate studies. I would like to thank her for her patience and guidance in mentoring me during this period. She has allowed me to develop my own thoughts about my work and is always there to encourage or help when I need it. These few years as her student has been inspiring and wonderful. I would also like to thank my thesis committee members Prof Mohan Balasubramanian, Dr Jedd Gregory and Dr Wang Yue for their support in allowing me to carry on with this project and giving me advice for my research. I also thank the lab members in Prof Mohan’s and Dr Jedd’s group for their support and friendship. Besides them, I would also like to thank Dr Naweed Naqvi and his lab members. I would like to extend my appreciation to my fellow lab mates; Zhang Dan, Aleksander Vjestica, Gu Ying, He Yue, Yang Jing, Maria Marakova, Sook Keat and Yuen Chyao. The times spent in lab have made us to be good friends which I will treasure dearly. I am also very blessed to have two wonderful attachment students, Khoo Wanxin and Shubham Khetan. I would like to thank them for their friendship and contributions. I am grateful to the imaging department, Dr. Meredith Calvert and Fiona Chia, for helping me with the image processing and microscopy work. Finally, I am grateful to Temasek Life Sciences Laboratory and Temasek Holdings for financial support and the National University of Singapore, NUS, for supporting me to be a graduate student. I thank my mother, Thing Shian, my father, Yam Cheong Beng and my brother , Shaun Yam and the rest of my family and friends for their understanding and support during this period. I also acknowledge Jack Leong, my boyfriend for his patience and advice during this period. Finally, I would like to thank God for his Grace in helping me throughout these years. III TITLE PAGE I DECLARATION II ACKNOWLEGEMENTS III TABLE OF CONTENTS IV SUMMARY VII LIST OF FIGURES IX PUBLICATIONS XII Table of Contents Chapter Introduction Mitosis is a tightly controlled and dynamic process . Varying Forms of Mitosis . Dynamics of the Nuclear Envelope and the Nuclear Pore Complexes . 10 Chromatin dynamics during mitosis . 15 The nuclear lamina 17 Nucleolus 19 Evolution of the nucleus and its constituents 21 S. japonicus as a new model system for study of mitosis . 23 Objectives . 26 Chapter Materials and Methods . 27 1. Drug treatments and Staining Reagents 27 2. Growth media and Conditions 27 3. S. japonicus and S. pombe strains . 28 4. Construction of S. japonicus strains 35 5. Synthetic markers 35 6. S. japonicus transformation . 37 7. Microscopy techniques . 38 8. Image processing and analysis 39 Chapter Results 42 1. S. japonicus and S. pombe exhibit markedly dissimilar progression of nuclear shapes and nuclear pore distribution during mitosis 42 1.1 The nucleus in S. japonicus elongates in a diamond-shaped during mitosis and relocalizes its NPCs towards the poles 42 1.2 Nuclear envelope proteins that are not associated with the NPCs remain evenly distributed around the periphery of the anaphase nucleus in S. japonicus cells . 46 IV 1.3 The mother nucleolus is discarded in between the segregated genomes during anaphase in S. japonicus . 49 2. The Nuclear Envelope Breaks and Reseals during mitosis in S. japonicus 51 2.1 Nuclear integrity is lost during mitosis in S. japonicus 51 2.2 The nuclear envelope ruptures from a single tear during mitosis in S. japonicus . 54 3. Elongating Mitotic spindles in S. japonicus are confined and grossly deformed by the nuclear envelope prior to the loss of nuclear integrity during anaphase. . 57 3.1 The intranuclear mitotic spindle buckles dramatically prior to NE breakage . 57 3.2 lem2 cells severely compromise nuclear structure in S. japonicus and mitotic spindles remain straight during anaphase . 59 3.3 The anaphase NE breakage is a cell-cycle entrained event . 62 4. Increases in the Nuclear Surface Area allows for “Closed” Mitosis . 64 4.1 S. pombe cells increase their nuclear surface area during anaphase by 30% while S. japonicus cells maintain a constant nuclear surface area 64 4.2 Inhibition of growth of NE surface area during mitosis in S. pombe cells leads to spindle breakage and a failure in nuclear division. . 68 5. Nuclear pores co-segregate with chromatin in mitotic S. japonicus cells. 71 5.1 The NPCs migrate polewards at late anaphase concurrent with the segregating chromatids . 71 5.2 Nhp6 preferentially binds the lagging pair of chromatin in mitotic S. japonicus cells . 74 5.3 The rDNA arrays segregates concurrently with nucleolar disassembly . 75 5.4 NPCs are associated with the anaphase chromatin and migrates together with chromatin towards cell ends . 78 6. The LEM domain protein Man1 is responsible for chromatin-NPC association in anaphase S. japonicus cells. 81 6.1 The conserved INM protein Man1 localizes to the NPCs in S. japonicus cells throughout the cell cycle. 81 6.2 Depletion of Man1 leads to abolishment of NPC “sliding” during anaphase in S. japonicus 83 6.3 Chromatin association with the nuclear membrane-bound NPCs is lost during anaphase in man1 cells in S. japonicus 86 6.4 Deletion of either N-terminal or C-terminal domains of Man1 results in a complete lack of NPC sliding . 90 6.5 pom152mutant cells exhibit a similar defect in chromatin-NPC association and NPCs “sliding” 93 The daughter nuclei are not born equal in S. japonicus cells lacking Man1. . 95 7.1 NPCs are not efficiently inherited in daughter nuclei in man1 cells and they are in varied sizes . 95 7.2 The NE breaks randomly along the length of the nucleus forming daughter nuclei of varying sizes in man1cells 98 V 8. Cells lacking Man1 exhibit defects in nucleolar disassembly during mitosis. 101 8.1 The nucleolus is unequally proportioned in the daughter nuclei and the rDNA arrays show improper structuring in man1 cells . 101 8.2 man1 mutant cells lacking either N- or C-terminal domains phenocopies the nucleolar disassembly defect of man1cells . 105 9. Re-association of chromatin with the nuclear periphery in man1 cells restores efficient inheritance of the NPCs, nucleolar remodelling and formation of equally sized daughter nuclei 107 Chapter Discussion 110 Divergence in the mitotic nuclear membrane management in the two fission yeasts species 110 Anaphase-specific perinuclear attachment of chromatin during mitosis in S. japonicus 114 Nucleolar Dispersal in S. japonicus 118 Genomic Similarity does not equate Cell Biology Similarity. 121 S. japonicus and S. pombe, the sister species for comparative cell biology of mitosis . 126 Chapter Conclusions and Perspectives 128 References . 130 VI Summary Evolution of nuclear division strategies within the fission yeast clade Mitosis, an integral eukaryotic process in which the microtubule-based spindle apparatus segregates the identical sets of chromosomes to the two daughter cells, must be carried out with high fidelity and accuracy. Failure to execute this process faithfully may lead to aneuploidy and cell death. The cellular genome in all eukaryotes is surrounded by the nuclear envelope, a membrane barrier made of two lipid bilayers and studded with the nuclear pores. The nuclear envelope isolates the genetic material from the cytoplasm and ensures functional separation between the nuclear-based transcription and RNA processing and cytoplasmic translation. The nucleus and its constituents must be remodelled during mitosis to accommodate the mitotic spindle assembly, chromosome segregation and formation of the daughter nuclei. This can be accomplished through a variety of strategies, with “open” and “closed” modes of mitosis positioned at the opposite ends of the spectrum. In the closed mitosis, the nuclear envelope remains intact throughout the nuclear division. Alternatively, the envelope of the original nucleus breaks down in prophase of mitosis and reassembles around the segregated daughter genomes following the mitotic exit. There is a diverse range of intermediate “variant” mitoses, where the nuclear integrity is lost but at different mitotic stages and through a variety of means. The existence of many variant mitoses in modern eukaryotes may reveal the evolutionary history of mitosis that could reflect the specialized physiology or the increasing cellular and organismal complexity. A fundamental question is how eukaryotic cells adapt the basic mitotic machinery to evolve strikingly different nuclear division strategies. There are four closely related species within the fission yeast clade. A popular model system, Schizosaccaromyces pombe (S. pombe) undergoes “closed” mitosis where nucleocytoplasmic compartmentalization is maintained throughout nuclear division. The early diverging VII Schizosaccharomyces japonicus (S. japonicus) is a relatively under-studied sister species that in early studies was suggested to differ in nuclear membrane remodelling. Here, I have demonstrated that S. japonicus cells undergo a unique form of “semi-open” mitosis where the nuclear envelope dramatically breaks during late anaphase B. I then went on show that the mitotic control of the nuclear surface area may determine the choice between the nuclear envelope breakdown and a fully closed division. In S. pombe, the nuclear membrane expands by approximately 30% during anaphase to allow maintenance of the constant volume during division of the mother nucleus into the two daughters. On the other hand, nuclear membrane does not expand in S. japonicus, necessitating the cell cycle-entrained nuclear envelope breakage to allow formation of the two daughter nuclei. I further showed that the S. japonicus cells utilize the conserved LEM domain protein, Man1, to structure the mitotic nucleus to ensure equal daughter nuclei formation, efficient nuclear pore complexes inheritance and dispersive nucleolar remodelling. My work shows that Man1 performs its function through physically linking the chromatin and the nuclear pore complexes. Interestingly, while the orthologues of this protein are known to structure chromatin at the nuclear periphery during interphase, Man1 in S. japonicus appears to have been repurposed for the mitotic role, in order to accommodate for the semi-open nuclear division mode in S. japonicus. Taken together, in this thesis I discuss my recent work on chromosome and nuclear envelope dynamics in Schizosaccharomyces japonicus and argue that the comparative cell biology studies using two closely related fission yeast species could provide unique insights into physiology and evolution of mitosis. VIII .List of Figures Fig. S. japonicus and S. pombe exhibit markedly dissimilar progression of nuclear shapes and nuclear pore distribution during mitosis Fig. 1.1 The nucleus in S. japonicus elongates in a diamond-shaped during mitosis and relocalizes its NPCs towards the poles Fig. 1.2 Nuclear envelope proteins that are not associated with the NPCs remain evenly distributed around the periphery of the anaphase nucleus in S. japonicus cells Fig. 1.3 The mother nucleolus is discarded in between the segregated genomes during anaphase in S. japonicus Fig. The Nuclear Envelope Breaks and Reseals during mitosis in S. japonicus Fig. 2.1 Nuclear integrity is lost during mitosis in S. japonicus Fig. 2.2 The nuclear envelope ruptures from a single tear during mitosis in S. japonicus Fig. Elongating Mitotic spindles in S. japonicus are confined and grossly deformed by the nuclear envelope prior to the loss of nuclear integrity during anaphase. Fig. 3.1 The intranuclear mitotic spindle buckles dramatically prior to NE breakage Fig. 3.2 lem2 cells severely compromise nuclear structure in S. japonicus and mitotic spindles remain straight during anaphase Fig. 3.3 The anaphase NE breakage is a cell-cycle entrained event Fig. Increases in the Nuclear surface area allows for “Closed” mitosis IX Fig. 4.1 S. pombe cells increase their nuclear surface area during anaphase by 30% while S. japonicus cells maintain a constant nuclear surface area Fig. 4.2 Inhibition of growth of NE surface area during mitosis in S. pombe cells leads to spindle breakage and a failure in nuclear division. Fig. Nuclear pores co-segregate with chromatin in mitotic S. japonicus cells. Fig. 5.1 The NPCs migrate polewards at late anaphase concurrent with the segregating chromatids Fig. 5.2 Nhp6 preferentially binds the lagging pair of chromatin in mitotic S. japonicus cells Fig. 5.3 The rDNA arrays segregate concurrently with nucleolar disassembly Fig. 5.4 NPCs are associated with the anaphase chromatin and migrates together with chromatin towards cell ends Fig. The LEM domain protein Man1 is responsible for chromatin-NPC association in anaphase S. japonicus cells. Fig. 6.1 The conserved INM protein Man1 localizes to the NPCs in S. japonicus cells throughout the cell cycle. Fig. 6.2 Depletion of Man1 leads to abolishment of NPC “sliding” during anaphase in S. japonicus Fig. 6.3 Chromatin association with the nuclear membrane-bound NPCs is lost during anaphase in man1 cells in S. japonicus . Fig. 6.4 Deletion of either N-terminal or C-terminal domains of Man1 results in a complete lack of NPC sliding Fig. 6.5 pom152mutant cells exhibit a similar defect in chromatin-NPC association and NPCs “sliding” X of genes as compared to C. reinhardtii (Hallmann et al., 2003 and Ferris et al., 2010). It is therefore proposed that V. carteri is evolved from C. reinhardtii through both the minor rewiring of the regulatory pathways and gene amplication to gain multi-cellularity (Nishii et al., 2010). Other examples include the different mitotic stages of the slime mold P. polycephalum during the different cell cycle stages and semi-open mitosis of the syncytial embryos of D. melanogaster and C.elegans as described earlier. S. japonicus and S. pombe, the sister species for comparative cell biology of mitosis Both S. japonicus and S. pombe are malleable and tractable yeast species, that in our opinion, prove ideal as a dual model system for the study of basic cellular processes, including mitosis. Both species can be manipulated using similar experimental procedures and their their genomes exhibit a considerable degree of similarity with most genes present as clear orthologues (Rhind et al., 2011). Yet, the two sister organisms exhibit dramatic divergence in the mitotic process. As an example, I have observed that the LEM domain protein Man1 retains its core function of tethering chromatin to the nuclear periphery in both species, yet, this function is exploited at the different stages of the cell cycle. Comparing the cytology and understanding how the mechanisms underlying mitotic nuclear dynamics function in both systems will reveal how different cellular physiologies evolve by fine-tuning a common toolbox of molecular players. 126 Figure 10 127 Chapter Conclusions and Perspectives The bulk of my graduate studies have been focused on elucidating the nuclear dynamics during mitosis in the fission yeast S. japonicus, an early diverging member of the fission yeast clade. I have found that S. japonicus breaks and reforms the NE during mitosis, in stark contrast to its well studied relative, S. pombe that undergoes nuclear division without losing nucleocytoplasmic compartmentalization. I have shown that the anaphase NE breakage is necessary to release the compressive force exerted by the non-expanding nuclear membrane on the elongating intranuclear spindle. The nuclear envelope rupture appears to be an active cell cycle dependent mechanism. An exciting direction to pursue these studies would be to understand mechanistically the processes underlying nuclear envelope breakage and reformation. On one hand, that would involve understanding the cell cycle signalling driving these phenomena. On the other hand, it would be of great interest to identify the molecular players at the NE that function at the crux of mitotic remodelling. With respect to the later problem, the evolutionary conserved LEM domain proteins could prove to be interesting candidates for futures studies. I believe that S. japonicus is set to gain popularity as a model organism for the study of basic cellular processes. Arguably, its most attractive attribute is that it presents an opportunity to perform comparative cell biology studies alongside its relative, S. pombe. My work clearly shows that while these yeast species are not highly diverged on the genomic level, they undergo strikingly different mitotic programs, likely by fine-tuning the mechanistic regulation of basic functional modules. One of the obvious directions to take would be to understand how the mechanisms underlying the nuclear membrane expansion at the G2/M 128 boundary diverged in the two species and whether it is possible to reconstitute the heterologous mode of membrane area control in a different species. My work suggests that the LEM domain protein Man executes a conceptually similar function in promoting perinuclear chromatin association in the two fission yeast species, yet the regulation of this function has diverged over the course of evolution. I showed that in S. japonicus, Man1 plays a direct role in recruiting the chromatin to the nuclear pore complexes during anaphase of mitosis while other reports demonstrated that in S. pombe, its orthologue appears to attach the interphase chromatin. The Man1-mediated mitotic NPC-chromatin attachment in S. japonicus is fundamental to the remodelling of nuclear membrane, the nuclear pore complexes and the nucleolus in this organism. Again, it would be of interest to understand mechanistically how such divergence is achieved. I have also found that unlike organisms with closed nuclear division, S. japonicus disassembles the nucleolus during mitosis, somewhat similarly to higher eukaryotes. Thus, S. japonicus could prove to be a wonderful experimental system to understand mitotic nucleolar dynamics and nucleolar biogenesis following mitosis. Interestingly, the perinuclear attachment of chromatin. including the chromatin ends proximal to the rDNA is important for the regulation of nucleolar disassembly during late anaphase in S. japonicus. Future work will include how the attachment of rDNA ends promotes proper nucleolar dynamics. On a broader level, I hope that my studies illustrate how a comparative cell biology approach using two related species could give insights into evolution and physiology of nuclear division. 129 References Akhtar A, G. S. (2007). "The nuclear envelope and transcriptional control." Nat Rev Genet 8(7): 507-517. Alber F, D. S., Veenhoff LM, Zhang W, Kipper J, Devos D, Suprapto A, Karni-Schmidt O, Williams R, Chait BT, Sali A, Rout MP. (2007). "The molecular architecture of the nuclear pore complex." Nature 450(7170): 695-701. Anderson, D. J. and M. W. Hetzer (2008). "The life cycle of the metazoan nuclear envelope." Curr Opin Cell Biol 20(4): 386-392. Anderson DJ, H. M. (2008). "Shaping the endoplasmic reticulum into the nuclear envelope." J Cell Sci. Anderson, D. J. H., M. (2007). "Nuclear envelope formation by chromatin-mediated reorganization of the endoplasmic reticulum. ." Nature Cell Biol 9. Anderson, D. J. H., M. W. (2008). "Reshaping of the endoplasmic reticulum limits the rate for nuclear envelope formation." J. Cell Biol 182. Angela Taddei, Heiko Schober,3 and Susan M. Gasser3 (2010). "The Budding Yeast Nucleus." Cold Spring Harb Perspect Biol. Aoki, K., H. Hayashi, K. Furuya, M. Sato, T. Takagi, M. Osumi, A. Kimura and H. Niki (2011). "Breakage of the nuclear envelope by an extending mitotic nucleus occurs during anaphase in Schizosaccharomyces japonicus." Genes Cells 16(9): 911-926. Aoki, K., R. Nakajima, K. Furuya and H. Niki (2010). "Novel episomal vectors and a highly efficient transformation procedure for the fission yeast Schizosaccharomyces japonicus." Yeast 27(12): 1049-1060. Aoki, K., Y. Shiwa, H. Takada, H. Yoshikawa and H. Niki (2013). "Regulation of nuclear envelope dynamics via APC/C is necessary for the progression of semi-open mitosis in Schizosaccharomyces japonicus." Genes Cells. Arnone JT, W. A., Cohen-Fix O. (2013). "The dynamic nature of the nuclear envelope: Lessons from closed mitosis." Nucleus 12(4): 4. 130 Asakawa, H., T. Kojidani, C. Mori, H. Osakada, M. Sato, D. Q. Ding, Y. Hiraoka and T. Haraguchi (2010). "Virtual breakdown of the nuclear envelope in fission yeast meiosis." Curr Biol 20(21): 1919-1925. Awaya J, O. T., Ohno H, Omura S. (1975). "Substitution of cellular fatty acids in yeast cells by the antibiotic cerulenin and exogenous fatty acids." Biochim Biophys Acta. . Beaudouin J, G. D., Daigle N, Eils R, Ellenberg J. ( 2002 ). "Nuclear envelope breakdown proceeds by microtubule-induced tearing of the lamina." Cell 108(1): 83-96. Bengtsson, L. (2007). "What MAN1 does to the Smads." FEBS Journal 274(6): 1374-1382. Brachner, A. (2005). "LEM2 is a novel MAN1-related inner nuclear membrane protein associated with A-type lamins." J Cell Sci. 118. Brachner, A. and R. Foisner (2011). "Evolvement of LEM proteins as chromatin tethers at the nuclear periphery." Biochem Soc Trans 39(6): 1735-1741. Brickner, S. A. a. J. H. (2010). "A role for DNA sequence in controlling the spatial organization of the genome." Nucleus. Carman, G. M., and Han, G.S. (2006.). "Roles of phosphatidate phosphatase enzymes in lipid metabolism. ." Trends Biochem. Sci. . Castagnetti, S., Oliferenko, S., and Nurse, P. (2010). "Fission yeast cells undergo nuclear division in the absence of spindle microtubules." PLoS Biology. Chase D, S. C., Ashcroft N, Kosinski M, Longo D, Ferris DK, Golden A. (2000). "The polo-like kinase PLK-1 is required for nuclear envelope breakdown and the completion of meiosis in Caenorhabditis elegans." Genesis. 26(1): 26-41. Choi HS, S. W., Morgan JM, Han GS, Xu Z, Karanasios E, Siniossoglou S, Carman GM. (2011). "Phosphorylation of phosphatidate phosphatase regulates its membrane association and physiological functions in Saccharomyces cerevisiae: identification of SER(602), THR(723), AND SER(744) as the sites phosphorylated by CDC28 (CDK1)encoded cyclin-dependent kinase." J Biol Chem. 131 Clarke PR, Z. C. (2008). "Spatial and temporal coordination of mitosis by Ran GTPase." Nat Rev Mol Cell Biol 9(6): 464-477. Clemente-Blanco A, M.-S. M., Schneider DA, Machín F, Jarmuz A, Tschochner H, Aragón L. ( 2009 ). "Cdc14 inhibits transcription by RNA polymerase I during anaphase." Nature. Cockell, M. M. (1999). "The nucleolus : Nucleolar Space for RENT." Current Biology 9. D'Angelo (2006). "The role of the nuclear envelope in cellular organization." Cell Mol Life Sci 63(3): 16. D'Angelo MA, H. M. (2008). "Structure, dynamics and function of nuclear pore complexes." Trends Cell Biol. 18: 10. Damien D’Amours, F. S. a. A. A. (2004). "Cdc14 and Condensin Control the Dissolution of Cohesin-Independent Chromosome Linkages at Repeated DNA." Cell 117. De Souza CP, O. A., Hashmi SB, Osmani SA. (2004). "Partial nuclear pore complex disassembly during closed mitosis in Aspergillus nidulans." Curr Biol 14(22): 1973-1984. De Souza, C. P. and S. A. Osmani (2009). "Double duty for nuclear proteins--the price of more open forms of mitosis." Trends Genet 25(12): 545-554. Dimario, P. J. (2004). "Cell and molecular biology of nucleolar assembly and disassembly." International Review of Cytolog 239. Drechsler (2012). "Exotic mitotic mechanisms." Open Biology 2(12). Dultz E, Z. E., Wurzenberger C, Braun M, Rabut G, Sironi L, Ellenberg J. (2008). "Systematic kinetic analysis of mitotic dis- and reassembly of the nuclear pore in living cells." J Cell Biol 180(5): 857-865. Ellenberg, J., E.D. Siggia, J.E. Moreira, C.L. Smith, J.F. Presley, H.J. Worman, and J. Lippincott-Schwartz. (1997.). "Nuclear membrane dynamics and reassembly in living cells: targeting of an inner nuclear membrane protein in interphase and mitosis. ." J. Cell Biol. 138(1193-1206). 132 Fahrenkrog B, A. U. (2003). "The nuclear pore complex: nucleocytoplasmic transport and beyond." Nat Rev Mol Cell Biol 4(10): 757-766. Finlan LE, S. D., Thomson I, Boyle S, Kerr E, Perry P, Ylstra B, Chubb JR, Bickmore WA. (2008). "Recruitment to the nuclear periphery can alter expression of genes in human cells." PLoS Genet 4(3). Foisner, R. (2003). "Cell cycle dynamics of the nuclear envelope." ScientificWorldJournal 3: 1-20. Franz C, W. R., Yavuz S, Santarella R, Gentzel M, Askjaer P, Galy V, Hetzer M, Mattaj IW, Antonin W. ( 2007 ). "MEL-28/ELYS is required for the recruitment of nucleoporins to chromatin and postmitotic nuclear pore complex assembly." EMBO Rep. 8(2). Frenkiel-Krispin D, M. B., Aebi U, Medalia O. (2010). "Structural analysis of a metazoan nuclear pore complex reveals a fused concentric ring architecture." J Mol Biol 395(3): 578586. Fry AM, O. R. L., Sabir SR, Bayliss R. (2012). "Cell cycle regulation by the NEK family of protein kinases." J Cell Sci. 125(19): 4423-4433. Fujita, I., Y. Nishihara, M. Tanaka, H. Tsujii, Y. Chikashige, Y. Watanabe, M. Saito, F. Ishikawa, Y. Hiraoka and J. Kanoh (2012). "Telomere-nuclear envelope dissociation promoted by Rap1 phosphorylation ensures faithful chromosome segregation." Curr Biol 22(20): 1932-1937. Furuya, K., K. Aoki and H. Niki (2012). "Construction of an insertion marker collection of Sz. japonicus (IMACS) for genetic mapping and a fosmid library covering its genome." Yeast 29(6): 241-249. Furuya, K. and H. Niki (2009). "Isolation of heterothallic haploid and auxotrophic mutants of Schizosaccharomyces japonicus." Yeast 26(4): 221-233. Furuya, K. and H. Niki (2012). "Hyphal differentiation induced via a DNA damage checkpoint-dependent pathway engaged in crosstalk with nutrient stress signaling in Schizosaccharomyces japonicus." Curr Genet 58(5-6): 291-303. 133 Gabriel, M. (1984 ). "Karyokinesis and Septum Formation during the Regeneration of Incomplete Cell Walls in Protoplasts of Schizosaccharomyces japonicus var. versatilis: a Time-lapse Microcinematographic Study " Microbiology 130(3). Gomez-Cavazos, J. S. and M. W. Hetzer (2012). "Outfits for different occasions: tissuespecific roles of Nuclear Envelope proteins." Curr Opin Cell Biol 24(6): 775-783. Gong D, F. J. J. (2010). "The roles of cyclin A2, B1, and B2 in early and late mitotic events." Mol Biol Cell 21(18). Gonzalez (2012). "Fission yeast Lem2 and Man1 perform fundamental functions of the animal cell nuclear lamina." Nucleus 3(1). Gruenbaum (2003). "The nuclear lamina and its functions in the nucleus." Int Rev Cytol 226: 62. Gruenbaum, Y., A. Margalit, R. D. Goldman, D. K. Shumaker and K. L. Wilson (2005). "The nuclear lamina comes of age." Nat Rev Mol Cell Biol 6(1): 21-31. Grund SE, F. T., Cabal GG, Antúnez O, Pérez-Ortín JE, Hurt E. ( 2008 ). "The inner nuclear membrane protein Src1 associates with subtelomeric genes and alters their regulated gene expression." J Cell Biol. 8(182). Gurskaya, N. G., Verkhusha, V.V., Shcheglov, A.S., Staroverov, D.B., Chepurnykh, T.V., Fradkov, A.F., Lukyanov, S., and Lukyanov, K.A. (2006 ). "Engineering of a monomeric green-to-red photoactivatable fluorescent protein induced by blue light. ." Nat Biotechnol 24. Guttinger, S., E. Laurell and U. Kutay (2009). "Orchestrating nuclear envelope disassembly and reassembly during mitosis." Nat Rev Mol Cell Biol 10(3): 178-191. Hennekes H, P. M., Weber K, Nigg EA. (1993). "Phosphorylation on protein kinase C sites inhibits nuclear import of lamin B2." J Cell Biol 120(6): 1293-1304. Hernandez-Verdun, D. (2005). "Nucleolus in the Spotlight " Cell cycle 4(1). Hernandez-Verdun, D. (2011). "Assembly and disassembly of the nucleolus during the cell cycle." Nucleus 2(3). 134 Hetzer (2010). "The nuclear envelope." Cold Spring Harb Perspect Biol 2(3). Hetzer, M. W., T. C. Walther and I. W. Mattaj (2005). "Pushing the envelope: structure, function, and dynamics of the nuclear periphery." Annu Rev Cell Dev Biol 21: 347-380. Hochegger, H., S. Takeda and T. Hunt (2008). "Cyclin-dependent kinases and cell-cycle transitions: does one fit all?" Nat Rev Mol Cell Biol 9(11): 910-916. Huber (2009). "Overlapping Functions of Nuclear Envelope Proteins NET25 (Lem2) and Emerin in Regulation of Extracellular Signal-Regulated Kinase Signaling in Myoblast Differentiation▿." Mol Cell Biol 10(1128). James M Holaska, K. L. W. a. M. M. (2002). "The nuclear envelope, lamins and nuclear assembly." Current opinion in cell biology 14. Karen (2006). "The spindle checkpoint " JCS 119. Karim Mekhail, J. S., Steven P. Gygi, Danesh Moazed (2009). "Role for perinuclear chromosome tethering in maintenance of genome stability." Nature. Khaitovich, P., W. Enard, M. Lachmann and S. Paabo (2006). "Evolution of primate gene expression." Nat Rev Genet 7(9): 693-702. Kumeta, M. (2012). "Nucleocytoplasmic Shuttling of Cytoskeletal Proteins: Molecular Mechanism and Biological Significance." International Journal of Cell Biology: 12. Kutay, U. and M. W. Hetzer (2008). "Reorganization of the nuclear envelope during open mitosis." Curr Opin Cell Biol 20(6): 669-677. Laurell E, B. K., Krupina K, Theerthagiri G, Bodenmiller B, Horvath P, Aebersold R, Antonin W, Kutay U. (2011). "Phosphorylation of Nup98 by multiple kinases is crucial for NPC disassembly during mitotic entry." Cell 144(4): 539-550. Leena Ukil, C. P. D. S., Hui-Lin Liu, Stephen A. Osmani (2009). "Nucleolar Separation from Chromosomes during Aspergillus nidulans Mitosis Can Occur Without Spindle Forces." Mol Biol Cell 20(8). 135 Leung AK, G. D., Miller G, Lyon C, Lam YW, Lleres D, Daigle N, Zomerdijk J, Ellenberg J, Lamond AI. (2004). "Quantitative kinetic analysis of nucleolar breakdown and reassembly during mitosis in live human cells." J Cell Biol. Lim H W G, H. G., Torii Y, Hirata A, Miller J, Sazer S. (2007). "Vesicle-like biomechanics governs important aspects of nuclear geometry in fission yeast." PLoS One. Loiodice I, A. A., Rabut G, Van Overbeek M, Ellenberg J, Sibarita JB, Doye V. 2004 (2004). " The entire Nup107–160 complex, including three new members, is targeted as one entity to kinetochores in mitosis. ." Mol Biol Cell 15: 3333-3344. Lu KP, H. T. (1995). "Evidence for a NIMA-like mitotic pathway in vertebrate cells." Cell 81(3): 413-424. Mall M, W. T., Gorjánácz M, Davidson IF, Nga Ly-Hartig TB, Ellenberg J, Mattaj IW. (2012). "Mitotic lamin disassembly is triggered by lipid-mediated signaling." J Cell Biol. 198(6): 981-990. Mans (2004). "Comparative Genomics, Evolution and Origins of the Nuclear Envelope and Nuclear Pore Complex." Cellk cycle 3(12). Mansharamani, M. (2005). "Direct Binding of Nuclear Membrane Protein MAN1 to Emerin in Vitro and Two Modes of Binding to Barrier-to-Autointegration Factor*." JBC 280. Martins SB, E. T., Steen RL, Jahnsen T, Skålhegg B S, Collas P. (2000). "HA95 is a protein of the chromatin and nuclear matrix regulating nuclear envelope dynamics." Cell Sci 113(21). Mayer, C. and I. Grummt (2005). "Cellular stress and nucleolar function." Cell Cycle 4(8): 1036-1038. Mekhail, K., J. Seebacher, S. P. Gygi and D. Moazed (2008). "Role for perinuclear chromosome tethering in maintenance of genome stability." Nature 456(7222): 667-670. Mewborn SK, P. M., Abuisneineh F, Fahrenbach JP, Zhang Y, MacLeod H, Dellefave L, Pytel P, Selig S, Labno CM, Reddy K, Singh H, McNally E. (2010). "Altered chromosomal positioning, compaction, and gene expression with a lamin A/C gene mutation." PLoS One. 136 Moazed, K. M. D. (May 2010). "The nuclear envelope in genome organization, expression and stability." Nature Reviews Molecular Cell Biology Mühlhäusser P, K. U. (2007). "An in vitro nuclear disassembly system reveals a role for the RanGTPase system and microtubule-dependent steps in nuclear envelope breakdown." J Cell Biol 178(4): 595-610. Mühlhäusser P, K. U. (2007). "An in vitro nuclear disassembly system reveals a role for the RanGTPase system and microtubule-dependent steps in nuclear envelope breakdown." J Cell Biol 178: 4. Nabeshima K, N. T., Straight AF, Murray A, Chikashige Y, Yamashita YM, Hiraoka Y, Yanagida M. (1998). "Dynamics of centromeres during metaphase-anaphase transition in fission yeast: Dis1 is implicated in force balance in metaphase bipolar spindle." Mol Biol Cell. Nicholas Rhind, Z. C., Moran Yassour, Dawn A Thompson, Brian J Haas, Naomi Habib, Ilan Wapinski, Sushmita Roy, Michael F. Lin, David I Heiman, Sarah K Young, Kanji Furuya, (2011). "Comparative Functional Genomics of the Fission Yeasts." Science 332. Okamoto, S., K. Furuya, S. Nozaki, K. Aoki and H. Niki (2013). "Synchronous activation of cell division by light or temperature stimuli in the dimorphic yeast Schizosaccharomyces japonicus." Eukaryot Cell. Olson, M. O. J. (2000). "The nucleolus: an old factory with unexpected capabilites." Trends Cell Biol. 10. Osmani AH, D. J., Liu HL, Nile A, Osmani SA. 2006. (2006). "Systematic deletion and mitotic localization of the nuclear pore complex proteins of Aspergillus nidulans." Mol Biol Cell 17: 4946-4961. Patterson, K. (2004). "The Functions of Klarsicht and Nuclear Lamin in Developmentally Regulated Nuclear Migrations of Photoreceptor Cells in the Drosophila Eye." Mol Biol Cell 15(2). Paul Nurse, P. T. (1980 ). "Regulatory Genes Controlling Mitosis in the Fission Yeast SCHIZOSACCHAROMYCES POMBE." Genetics 96(3). 137 Paull TT, J. R. (1995). "DNA looping by Saccharomyces cerevisiae high mobility group proteins NHP6A/B. Consequences for nucleoprotein complex assembly and chromatin condensation." J Biol Chem. Pemberton LF, P. B. (2005). "Mechanisms of receptor-mediated nuclear import and nuclear export." Traffic 6(3): 187-198. Portier N, A. A., Maddox PS, Green RA, Dammermann A, Desai A, Oegema K. (2007). "A microtubule-independent role for centrosomes and aurora a in nuclear envelope breakdown." Dev Cell 12(4): 515-529. Puckelwartz MJ, D. F., McNally EM (2011). "Gene expression, chromosome position and lamin A/C mutations." Nucleus 2(3). Puhka M, V. H., Joensuu M, Jokitalo E ( 2007.). " Endoplasmic reticulum remains continuous and undergoes sheet-to-tubule transformation during cell division in mammalian cells." J Cell Biol 179: 895-909. Quimby BB, D. M. (2003). "The small GTPase Ran: interpreting the signs." Curr Opin Cell Biol 15(3): 338-344. Reddy KL, Z. J., Bertolino E, Singh H. (2008). "Transcriptional repression mediated by repositioning of genes to the nuclear lamina." Nature 452(7184). Robinow, C. F. and Hyams, J. S. (1989). General cytology of fission yeast. In Molecular Biology of Fission Yeast (ed. A. Nasim, P. Young and B. F. Johnson). pp. 273-330. Academic Press, San Diego Rothbauer, U., Zolghadr, K., Muyldermans, S., Schepers, A., Cardoso, M.C., and Leonhardt, H. (2008). A versatile nanotrap for biochemical and functional studies with flourescent fusion proteins. Mol. Cell. Proteomics 7, 282-289 Saitoh S, T. K., Nabeshima K, Yamashita Y, Nakaseko Y, Hirata A, Yanagida M. (1996). "Aberrant mitosis in fission yeast mutants defective in fatty acid synthetase and acetyl CoA carboxylase." J Cell Biol. Salina D, B. K., Eckley DM, Schroer TA, Rattner JB, Burke B. (2002). "Cytoplasmic dynein as a facilitator of nuclear envelope breakdown." Cell 11(108): 97-107. 138 Schneider, A. (1873) Untersuchungen über Plathelminthen. Jahrb. Oberhess. Ges. Naturwiss. Heilk. 14, 69–81 (in German). Schreiber, K. H. and B. K. Kennedy (2013). "When lamins go bad: nuclear structure and disease." Cell 152(6): 1365-1375. Shaw, P. (2005). "The Nucleolus Playing by different rules." Cell cycle 4(1). Shu H, W. T., Siretskiy A, Gruissem W, Hennig L. (2012). "Distinct modes of DNA accessibility in plant chromatin." Nat Commun 3(1281). Singh, M., C. R. Hunt, R. K. Pandita, R. Kumar, C. R. Yang, N. Horikoshi, R. Bachoo, S. Serag, M. D. Story, J. W. Shay, S. N. Powell, A. Gupta, J. Jeffery, S. Pandita, B. P. Chen, D. Deckbar, M. Lobrich, Q. Yang, K. K. Khanna, H. J. Worman and T. K. Pandita (2013). "Lamin A/C depletion enhances DNA damage-induced stalled replication fork arrest." Mol Cell Biol 33(6): 1210-1222. Snyder, D. G. a. M. (1991). "Segregation of the Nucleolus During Mitosis in Budding and Fission Yeast." Cell motility and the cytoskeleton 20. Steinberg G, P.-M. J. (2008). "Ustilago maydis, a new fungal model system for cell biology." Trends Cell Biol. 18(2): 61-67. Straight, A. F., Belmont, A.S., Robinett, C.C., and Murray, A.W. (1996). "GFP tagging of budding yeast chromosomes reveals that protein-protein interactions can mediate sister chromatid cohesion. ." Curr Biol Tanaka (1986). "Mitosis in the fission yeast Schizosaccharomyces pombe as revealed by freeze=substitution electron microscopy." J Cell Science 80: 253-268. Thadani R, U. F., Heeger S. (2012). "Condensin, chromatin crossbarring and chromosome condensation." Current Biology. Tran EJ, W. S. ( 2006). "Dynamic nuclear pore complexes: life on the edge." cell 125(6): 1041-1053. 139 Ulbert, S., W. Antonin, M. Platani and I. W. Mattaj (2006). "The inner nuclear membrane protein Lem2 is critical for normal nuclear envelope morphology." FEBS Lett 580(27): 64356441. Vagnarelli P, E. W. (2012). "Repo-Man-PP1: a link between chromatin remodelling and nuclear envelope reassembly." Nucleus 3(138-42). Visintin, R., E. S. Hwang and A. Amon (1999). "Cfi1 prevents premature exit from mitosis by anchoring Cdc14 phosphatase in the nucleolus." Nature 398(6730): 818-823. Vjestica A, O. S. (2012 ). "Nuclear geometry: mitotic nucleus flares out when arrested." Curr Biol. Wagner N, K. G. (2007). "LEM-Domain proteins: new insights into lamin-interacting proteins." Int Rev Cytol 26(1). Wente SR, R. M. (2010 ). "The nuclear pore complex and nuclear transport." Cold Spring Harb Perspect Biol. 10. Wilson, K. L. (2010). "Lamin-binding Proteins " Cold Spring Harb Perspect Biol 2(4). Witkin KL, C. Y., Shao S, Webster MT, Lahiri S, Walters AD, Lee B, Koh JL, Prinz WA, Andrews BJ, Cohen-Fix O. ( 2012). "The budding yeast nuclear envelope adjacent to the nucleolus serves as a membrane sink during mitotic delay." Curr Biology. Worman, H. J. (2012). "Nuclear lamins and laminopathies." J Pathol 226(2): 316-325. Yewdell, W. T., P. Colombi, T. Makhnevych and C. P. Lusk (2011). "Lumenal interactions in nuclear pore complex assembly and stability." Mol Biol Cell 22(8): 13751388. Yoder, T. J., C. G. Pearson, K. Bloom and T. N. Davis (2003). "The Saccharomyces cerevisiae spindle pole body is a dynamic structure." Mol Biol Cell 14(8): 3494-3505. Zhang, D., and Oliferenko, S. (2012). " Remodeling the nuclear membrane during closed mitosis." Curr Opin Cell Biol. 140 Zhang D, V. A., Oliferenko S. (2010). "The cortical ER network limits the permissive zone for actomyosin ring assembly." Current Biology 8(20). Zheng L, S. C., Magidson V, Khodjakov A, Oliferenko S. ( 2007). "The spindle pole bodies facilitate nuclear envelope division during closed mitosis in fission yeast." PLoS Biol. Zheng, L., C. Schwartz, V. Magidson, A. Khodjakov and S. Oliferenko (2007). "The spindle pole bodies facilitate nuclear envelope division during closed mitosis in fission yeast." PLoS Biol 5(7): e170. Zullo JM, D. I., Piqué-Regi R, Gaffney DJ, Epstein CB, Spooner CJ, Luperchio TR, Bernstein BE, Pritchard JK, Reddy KL, Singh H (2012). "DNA sequence-dependent compartmentalization and silencing of chromatin at the nuclear lamina." Cell 147(7). 141 [...]... nuclear envelope and migrates into the cytoplasm of the daughter cell, leaving behind the mother nuclear envelope (Steinberg et al., 2008) In another illustration, in the syncytial embryonic divisions of the Drosophila melanogaster (D melanogaster), the nuclear envelope is broken at the poles during prometaphase together with disassembly of the NPCs while the rest of the nuclear membrane remains intact... semi-open mitosis of the dimorphic plant pathogen, U maydis, microtubules nucleated from the SPB proximal to the mother cell together with dyenin pushes the chromosomes into the daughter bud, extracting the SPBs from the nuclear periphery and stripping off the NE The mitotic apparatus migrates into the daughter nuclei and a mitotic spindle is formed inside the daughter In addition, the fission yeast Schizosaccharomyces... organisms where the nuclear envelope do not break, the MTOC is found situated within the 8 NE during mitosis In contrast, in organisms with closed mitosis such as the popular model systems fission yeast S pombe and the budding yeast Saccharomyces cerevisiae (S cerevisiae), the mitotic microtubule-organizing centers (MTOCs) known as the spindle pole bodies (SPBs), anchored within the plane of the nuclear envelope,... disassembly of nuclear complexes such as the nuclear pore complexes 6 and restructuring of nuclear- related components such as chromatin, the nucleolus, the timing of spindle formation, among many other aspects However, only in closed mitosis is nuclear integrity preserved; in all other forms, the compartmentalization between the nucleus and cytoplasm is compromised During the classical open mitosis, the nuclear. .. the nucleus isolating the nuclear genome from the rest of the cellular organelles Although organisms in the eukaryotic kingdom differ greatly, the basic nuclear structure and various nuclear pore associated proteins appear conserved throughout evolution Amongst some of the most evolutionary conserved nucleoporins are Gle2, Nup98, Nup96 and Nsp1 suggesting that their functions are integral for the nuclear. .. components In the Introduction, I shall introduce the main regulatory pathways involved in mitosis Next, I shall discuss the main differences between the different modes of mitosis Following that, I will describe the dynamics of the various nuclear- associated components, namely the nuclear envelope including the nuclear pores complexes, chromatin, nuclear lamina and the nucleolus with respect to the different... division, there are different degrees of nuclear disassembly, all resulting in the loss of nuclear integrity at some point in mitosis The filamentous ascomycete, A nidulans, loses nuclear integrity by partially permeabilizing the NPCs While the overall nuclear envelope structure remains, 14 nucleoporins disperse from the nuclear pore complexes, promoting the nuclear entry of the Cdk1/cyclin B complex (Osmani... supporting the nuclear structure, in the absence of a functional nuclear lamina Nucleolus The nucleolus is a well defined but not membrane bound nuclear subcompartment that is the site of ribosome biogenesis (Shaw et al., 2005) The rDNA transcription by the RNA polymerase I occurs inside the innermost compartment, the fibrillar centre that contains the rDNA, also known as the nucleolar organizing region The. .. cerevisiae, the Kap121 nuclear transport pathway is altered during mitosis through the binding of Nup53 to Kap121 (Dasso et al., 2013) In S pombe, the cytoplasmic SPB is relocalized to the nuclear periphery to facilitate intranuclear spindle formation at the start of mitosis However, the spatial localization of the MTOCs is not the only factor influencing the choice between different mitotic mechanisms In the. .. time In these cases, semi-open mitosis first take place where the chromosomes from a single nucleus remain confined within the same NE until they are captured by their respective centrosomes This provides a safety measure to prevent crossattachment with centrosomes from other nearby nuclei Dynamics of the Nuclear Envelope and the Nuclear Pore Complexes As one of the most prominent organelles in the eukaryotic . I EVOLUTION OF NUCLEAR DIVISION STRATEGIES WITHIN THE FISSION YEAST CLADE YAM QIU XIA, CANDICE (B.Sc. (Hons), NTU) A THESIS SUBMITTED FOR THE DOCTOR OF PHILOSPOHY. mitosis positioned at the opposite ends of the spectrum. In the closed mitosis, the nuclear envelope remains intact throughout the nuclear division. Alternatively, the envelope of the original nucleus. Varying Forms of Mitosis 6 Dynamics of the Nuclear Envelope and the Nuclear Pore Complexes 10 Chromatin dynamics during mitosis 15 The nuclear lamina 17 Nucleolus 19 Evolution of the nucleus