This goal of this proposal is to develop imaging tools capable of imaging the functional genome by mapping the three dimensional binding sites and clustering of transcription factors and histone modifiers. We will use single molecule particle tracking within the entire nucleus of the cell in real time. Initially this will be done in culture ES cells, and ultimately in living animals. There will be three locations involved: Albert Einstein College of Medicine, the University of California at Berkeley and the Janelia Research Campus of the HHMI, where the Transcription Imaging Consortium integrates the efforts of the investigators of this proposal. The reagents will be developed at Einstein and Berkeley and the microscope technology that has been developed and used predominantly at Janelia, will inform further modifications in building similar microscopes at Berkeley and Einstein. Genes of interest will be marked to image promoter-enhancer interactions in cells, tissues and organisms with high resolution. The microscopes employed and developed for these applications will be the multifocal microscope, the lattice light sheet microscope, the adaptive optics microscope and the high-speed three-color super registration microscope. Importantly, we will evaluate the levels of phototoxicity for the imaging protocols on each microscope and develop approaches to minimize it. Microscopes developed will be made available to the Nucleome community at the Einstein and Berkeley sites and at Janelia through a resource sharing facility, the Advanced Imaging Center, supported by the HHMI and the Moore Foundation with the explicit purpose of disseminating the use of the technology. No funds are requested for the Janelia component of this proposal. All funds will be for development of microscopes and reagents that will be at Berkeley and Einstein.

Public Health Relevance

We intend to develop new tools to image into living cells and tissues of animals in order to see the innermost workings of the cell, in particular the process by which genes are regulated. These tools, which include newer and more powerful microscopes and probes that emit light when imaged, will report on the mechanisms that turn genes on and off. These mechanisms are ultimately at the basis of all disease and malfunctioning of the organism.

Agency
National Institute of Health (NIH)
Institute
National Institute of Biomedical Imaging and Bioengineering (NIBIB)
Type
Research Project--Cooperative Agreements (U01)
Project #
5U01EB021236-03
Application #
9302783
Study Section
Special Emphasis Panel (ZRG1)
Program Officer
Zullo, Steven J
Project Start
2015-09-30
Project End
2020-06-30
Budget Start
2017-07-01
Budget End
2018-06-30
Support Year
3
Fiscal Year
2017
Total Cost
Indirect Cost
Name
Albert Einstein College of Medicine, Inc
Department
Type
DUNS #
079783367
City
Bronx
State
NY
Country
United States
Zip Code
10461
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Chong, Shasha; Dugast-Darzacq, Claire; Liu, Zhe et al. (2018) Imaging dynamic and selective low-complexity domain interactions that control gene transcription. Science 361:
Limi, Saima; Senecal, Adrien; Coleman, Robert et al. (2018) Transcriptional burst fraction and size dynamics during lens fiber cell differentiation and detailed insights into the denucleation process. J Biol Chem 293:13176-13190
Tsai, Albert; Singer, Robert H; Crocker, Justin (2018) Transvection Goes Live-Visualizing Enhancer-Promoter Communication between Chromosomes. Mol Cell 70:195-196
Tsai, Albert; Muthusamy, Anand K; Alves, Mariana Rp et al. (2017) Nuclear microenvironments modulate transcription from low-affinity enhancers. Elife 6:

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