Nuclear architecture and nuclear function appear to go hand in hand, as defects in nuclear organization are associated with aging and diseases such as cancer. We have been using budding yeast as a model system to study nuclear architecture. The yeast nucleus differs from that of higher eukaryotes in two aspects: (1) yeast lack lamins, proteins that play a major structural role in shaping the nucleus in metazoans, and (2) the yeast nuclear envelope (NE) remains intact throughout the cell cycle, unlike the NE of higher eukaryotes, which breaks down during mitosis and reassembles after chromosome segregation is completed. Nonetheless, the yeast nucleus shares important features with nuclei of higher eukaryotes: the NE has to expand during the course of the cells cycle, and the nucleus has to acquire and maintain a spherical shape and a volume proportional to cell volume. How the NE expands and what determine nuclear size and shape are questions that remain to be resolved in all systems. Our previous studies identified a number of conditions that lead to abnormal nuclear morphology, most notably elevated phospholipid synthesis (e.g. the spo7 mutant) and a cell cycle delay in mitosis. In both cases the alteration to nuclear shape is confined to the NE region adjacent to the nucleolus, and our studies showed that this confinement is dependent on the conserved polo-like kinase Cdc5. To further understand what affects nuclear morphology we sought additional mutants that have an abnormal nuclear shape. One of these turned out to be in the conserved topoisomerase 2 gene, TOP2. These mutant cells display multiple projection of the nuclear envelope that are not confined to the region adjacent to the nucleolus. Interestingly, only certain top2 mutants confer an abnormal nuclear shape, and this phenotype is correlated with abnormal DNA structure as determined by reduced DAPI staining. This suggests that DNA structure can affect nuclear morphology. This project is carried out in collaboration with Jeff Bachant's lab. To gain a better understanding of the relationship between nuclear and cell size, we have taken two parallel approaches: first, we began to isolate conditional mutants that have an abnormal nuclear/cell volume ratio. This work is ongoing and it is being done in collaboration with the labs of Brenda Andrews and Mike Tyers. Second, we asked what would happen to nuclear shape if cell growth is inhibited. To date, conditions that led to abnormal nuclear shape were done by manipulating the NE, either by increasing the amount of phospholipids or by preventing nuclear division. These conditions could not address whether nuclear size is determined by the availability of nuclear membrane, which in turn may be proportional to cell size. To this end, we examined the consequences of inhibiting cell size expansion on nuclear morphology. Our data suggest that NE expansion is not linked to cell size, and that when cell growth is inhibited the nucleus deforms while maintaining the same nuclear/cell volume ratio. This observation suggests that nuclear volume, and not surface area, is the limiting factor for nuclear size. This work was done in collaboration with Olivier Gadal and Carolyn Larabell. Finally, we found that the spo7 strain exhibits an age dependent defect in mitochondrial morphology. This is also observed in other mutants in lipid synthesis pathways. In collaboration with the Amon and Carman labs we found that as spo7 mutant cell age, their mitochondria fragment and then cluster, losing function in the process. We further observed that these cells can still bud, and least for a few generations, and segregate mitochondria to the bud. Interestingly, the mitochondria that reach the bud are perfectly functional, suggesting that the cell has a quality control mechanism that ensures that daughter cells receive functional mitochondria. This work was done in collaboration with George Carman and Angelika Amon

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12
Fiscal Year
2018
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U.S. National Inst Diabetes/Digst/Kidney
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Cohen-Fix, Orna; Kutay, Ulrike (2016) Editorial overview: The cell nucleus: Plastic, elastic and fantastic. Curr Opin Cell Biol 40:iv-v
Meseroll, Rebecca A; Cohen-Fix, Orna (2016) The Malleable Nature of the Budding Yeast Nuclear Envelope: Flares, Fusion, and Fenestrations. J Cell Physiol 231:2353-60
Cinquin, Bertrand P; Do, Myan; McDermott, Gerry et al. (2014) Putting molecules in their place. J Cell Biochem 115:209-16
Walters, Alison D; May, Christopher K; Dauster, Emma S et al. (2014) The yeast polo kinase Cdc5 regulates the shape of the mitotic nucleus. Curr Biol 24:2861-7
Heald, Rebecca; Cohen-Fix, Orna (2014) Morphology and function of membrane-bound organelles. Curr Opin Cell Biol 26:79-86
Walters, Alison D; Cohen-Fix, Orna (2013) Nuclear division: giving daughters their fair share. Curr Biol 23:R1045-7
Arnone, James T; Walters, Alison D; Cohen-Fix, Orna (2013) The dynamic nature of the nuclear envelope: Lessons from closed mitosis. Nucleus 4:
Witkin, Keren L; Chong, Yolanda; Shao, Sichen et al. (2012) The budding yeast nuclear envelope adjacent to the nucleolus serves as a membrane sink during mitotic delay. Curr Biol 22:1128-33
Cohen-Fix, Orna (2012) Orna Cohen-Fix: playing with nuclear morphology. Interview by Caitlin Sedwick. J Cell Biol 196:550-1
Walters, Alison D; Bommakanti, Ananth; Cohen-Fix, Orna (2012) Shaping the nucleus: factors and forces. J Cell Biochem 113:2813-21

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