Cell size varies greatly among different cell types and organisms, and especially during the reductive divisions that characterize early development. A fundamental question is how organelle size is appropriately regulated relative to cell size. The nucleus is one organelle that exhibits exquisite size scaling both during development and between species. The normal relationship between nuclear and cell size is often abrogated in cancers and other disease states, yet mechanisms that regulate nuclear size are largely unknown and may directly contribute to cancer progression. How steady-state nuclear size is determined is poorly understood. This knowledge gap prevents us from understanding how nuclear size impacts chromatin organization, gene expression, and cell function. The long-term goal is to elucidate mechanisms of nuclear size regulation to understand how nuclear size impacts cell and nuclear function and sub-nuclear organization. The objective of this application is to identify the molecular mechanisms that regulate nuclear expansion and shrinking and to demonstrate how these mechanisms control nuclear size in vivo. The central hypothesis is that steady-state nuclear size is determined by balanced nuclear growth and shrinking activities, which will be tested by pursuing the following three specific aims. 1) Identify mechanisms that regulate nuclear expansion: Nuclei reconstituted in egg extracts from two different size Xenopus frog species exhibit differential nuclear growth rates. Through biochemical characterization of these extracts and microscopy, the contribution of nuclear import cargos to interspecies differences in nuclear expansion will be demonstrated. 2) Identify mechanisms that regulate nuclear shrinking: Early stage Xenopus embryos contain larger cells and nuclei than later stage embryos, and large nuclei isolated from early stage embryos become smaller when incubated in cell extract from late stage embryos. Live time-lapse microscopy will be used to characterize the dynamics of this novel activity and biochemical approaches will identify factors responsible for nuclear shrinking. 3) Demonstrate the in vivo activities of nuclear scaling factors: Some factors that control nuclear size are known and others will be identified in Aims 1 and 2. Nuclear scaling activities will be manipulated in Xenopus embryos by mRNA microinjection and effects on nuclear size and dynamics in vivo will be examined by live cell microscopy. Nuclear scaling factors will also be genetically modulated in budding yeast to determine if their function is conserved. The expected outcome is elucidation of nuclear size control mechanisms, providing the foundation to test the novel hypothesis that nuclear size regulates nuclear organization and function. Organellar scaling is essential to cellular balance, yet mechanisms that maintain size ratios in a cell are largely unknown. This research will thus significantly impact our understanding of how scaling is regulated during biogenesis and growth.
Nuclear size is often deregulated in cancer cells and many cancers are diagnosed and staged based on graded increases in nuclear size. Little is known about the causes or effects of nuclear morphology changes in cancer, so understanding factors and mechanisms of nuclear scaling and how nuclear size impacts cell function will shed light on the contribution of nuclear size to cancer development and progression. Novel approaches and targets for the diagnosis and treatment of cancer might be suggested, and new cancer susceptibility factors could be identified to aid in prevention.
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