The information stored in an organism's genome directs its development and behavior, but the regulatory networks that control the expression of that information are only beginning to be understood in metazoans. Determining precisely when and where genes, particularly transcription factor genes, are expressed is central to gaining a comprehensive understanding of these networks. The nematode C. elegans offers unique advantages for determining gene expression patterns and in turn the regulatory networks that control them. These advantages include a fixed cell lineage, a total somatic cell number of less than 1,000, a compact, fully sequenced genome and a transparent body throughout the life cycle. Exploiting these advantages in the past grant periods, we have developed 4D imaging technology that automatically determines expression patterns of individual genes in each cell with high temporal resolution over the first half of embryogenesis. We have applied this technology to determine the embryonic expression patterns of some 200 transcription factor genes, revealing a wide variety of expression patterns that suggest roles for these genes in specifying cell identity. More recently we have devised methods to collect timed series data for FACS sorted embryonic cells to provide coarser spatiotemporal estimates of embryonic expression of all genes in the genome. We propose in the coming grant period to extend and complete the catalog of expression patterns for transcription factors using the 4D technology. We will build a recently developed diSPIM microscope and use that to assay expression through embryogenesis. We will complete the construction of strains with GFP tagged transcription factors and use these to determine the detailed expression patterns for the bulk of transcription factors in the genome. To complement this we will perform RNA-seq. on FACS sorted tissues and cell populations from roughly synchronized embryos to measure the dynamics of gene expression for all genes in the genome. We will also explore technology to provide expression data on individual cells of the embryo. These data sets, combined with ChIP-seq. and other data sets will be used to build regulatory networks active in each cell throughout embryogenesis. All the data and strains will be made available to the community through our website, Worm Base and the Caenorhaditis Genetics Center. Knowledge of the regulatory networks of this simple metazoan will have direct implications for understanding regulatory networks in humans both in health and in disease.

Public Health Relevance

We have developed and are applying technology to determine the expression of each of the individual transcription factor genes at the single cell level throughout embryogenesis of the model organism C. elegans. In addition we will measure transcript abundance of all genes, but with coarser time and spatial resolution, using RNA-seq. These datasets combined with information about transcription factor binding sites being acquired in other projects will provide unique insights into the regulatory networks that control information flow from the genome to direct development.

Agency
National Institute of Health (NIH)
Type
Research Project (R01)
Project #
2R01GM072675-10
Application #
8759127
Study Section
Development - 1 Study Section (DEV1)
Program Officer
Sledjeski, Darren D
Project Start
Project End
Budget Start
Budget End
Support Year
10
Fiscal Year
2014
Total Cost
Indirect Cost
Name
University of Washington
Department
Genetics
Type
Schools of Medicine
DUNS #
City
Seattle
State
WA
Country
United States
Zip Code
98195
Aydin, Zafer; Murray, John I; Waterston, Robert H et al. (2010) Using machine learning to speed up manual image annotation: application to a 3D imaging protocol for measuring single cell gene expression in the developing C. elegans embryo. BMC Bioinformatics 11:84
Zhao, Zhongying; Flibotte, Stephane; Murray, John I et al. (2010) New tools for investigating the comparative biology of Caenorhabditis briggsae and C. elegans. Genetics 184:853-63
Zhao, Zhongying; Boyle, Thomas J; Liu, Zongzhi et al. (2010) A negative regulatory loop between microRNA and Hox gene controls posterior identities in Caenorhabditis elegans. PLoS Genet 6:e1001089
Bao, Zhirong; Zhao, Zhongying; Boyle, Thomas J et al. (2008) Control of cell cycle timing during C. elegans embryogenesis. Dev Biol 318:65-72
Murray, John Isaac; Bao, Zhirong; Boyle, Thomas J et al. (2008) Automated analysis of embryonic gene expression with cellular resolution in C. elegans. Nat Methods 5:703-9
Zhao, Zhongying; Boyle, Thomas J; Bao, Zhirong et al. (2008) Comparative analysis of embryonic cell lineage between Caenorhabditis briggsae and Caenorhabditis elegans. Dev Biol 314:93-9