Investigating Mechanisms of Intracellular Scaling Cell size varies widely among different organisms as well as within the same organism in different tissue types and during development, placing variable metabolic and functional demands on organelles and internal structures. A fundamental question is how essential subcellular components such as the nucleus, mitotic spindle and chromosomes are regulated to accommodate cell size differences. Xenopus frogs offer two physiological contexts in which we can investigate this question. First, we can compare Xenopus laevis to the smaller, related species Xenopus tropicalis, which lays smaller eggs and has proportionally smaller cells throughout development. Second, we can compare different stages of Xenopus laevis embryogenesis, as the ~1 millimeter diameter egg rapidly cleaves to form smaller blastomeres, which by the 15th division are reduced to 40 microns across. A unique aspect of our approach is to prepare cytoplasmic extracts from eggs and embryos that recapitulate organelle scaling in vitro, which we can use to identify molecular differences that underlie size changes. Our first specific aim focuses on the mitotic spindle, and we take advantage of computer simulations to identify parameters of microtubule dynamics and organization that could contribute to spindle size and morphology changes between species and during development.
This aim also develops novel methods to examine extrinsic scaling mechanisms by physically confining spindle assembly reactions inside different sized droplets, which will reveal whether there are size thresholds that scale the spindle externally or alter assembly pathways.
Aim 2 investigates how the size of mitotic chromosomes is altered during development to coordinate with spindle length so that complete segregation occurs.
In Aim 3, we begin addressing the importance of organelle scaling by examining the consequences of altering nuclear size during development in Xenopus laevis. These experiments will provide insight into how scaling occurs and contributes to intracellular morphogenesis and cell division, processes essential for viability and development, and defective in human diseases including cancer.

Public Health Relevance

Size varies widely in biology at many levels: the animal, the cells that make up the animal, and the contents of the cells, but we don't understand how proper scaling occurs so that everything fits and functions properly. This proposal investigates how cells regulate the size of their internal structures, including the chromosomes that contain the genome and the mitotic spindle that separates the chromosomes to daughter cells, and addresses the importance of internal size regulation in the developing frog embryo. Since defects in cell division and size regulation are often observed in cancer, our research may shed light on the underlying mechanisms and lead to new therapies.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
1R01GM098766-01A1
Application #
8295911
Study Section
Nuclear and Cytoplasmic Structure/Function and Dynamics Study Section (NCSD)
Program Officer
Hagan, Ann A
Project Start
2012-08-01
Project End
2016-07-31
Budget Start
2012-08-01
Budget End
2013-07-31
Support Year
1
Fiscal Year
2012
Total Cost
$463,717
Indirect Cost
$167,412
Name
University of California Berkeley
Department
Biochemistry
Type
Schools of Arts and Sciences
DUNS #
124726725
City
Berkeley
State
CA
Country
United States
Zip Code
94704
Heald, Rebecca; Cohen-Fix, Orna (2014) Morphology and function of membrane-bound organelles. Curr Opin Cell Biol 26:79-86
Helmke, Kara J; Heald, Rebecca (2014) TPX2 levels modulate meiotic spindle size and architecture in Xenopus egg extracts. J Cell Biol 206:385-93
Good, Matthew C; Vahey, Michael D; Skandarajah, Arunan et al. (2013) Cytoplasmic volume modulates spindle size during embryogenesis. Science 342:856-60
Whitehead, Evan; Heald, Rebecca; Wilbur, Jeremy D (2013) N-terminal phosphorylation of p60 katanin directly regulates microtubule severing. J Mol Biol 425:214-21
Wilbur, Jeremy D; Heald, Rebecca (2013) Mitotic spindle scaling during Xenopus development by kif2a and importin *. Elife 2:e00290