Our long term goal is to describe in the language of genetics and biochemistry the feedback cycles and pathways that comprise intracellular circadian systems - how they work, how they are synchronized with the environment, and how time information generated by them is used to regulate the behavior of cells. This proposal focuses on the model system Neurospora, as well as on the mouse and mammalian cell lines, to understand the paradigms underlying circadian control of cell physiology and metabolism. We also continue a longstanding effort aimed at understanding circadian photoreception and photobiology in Neurospora and use this to break new ground on a salient fungal pathogen.
In Specific Aim 1, we will carry out a global analysis and description of the circadian output network in Neurospora, using RNA sequencing, chromatin immunoprecipitation, and bioinformatics to describe the regulatory hierarchy governing circadian regulation of transcription in a cell. Then, using the exceptional background of biochemical genetics available in Neurospora, we will track the metabolic activities carried out by the proteins encoded by clock-controlled genes, and as foci of clock-controlled activities emerge, we will begin to lay the spectrum of clock-controlled processes on the Neurospora metabolic map to see how the clock regulates metabolism and physiology.
In Specific Aim 2, we will exploit recent atomic level structure analysis of the photoreceptive domain to determine the biological significance of photocycle kinetics. We will extend analysis of photobiology to the important fungal pathogen Aspergillus fumigatus where we can envision a way to exploit our understanding of fungal photobiology to enable a new treatment for hundreds of thousands of patients with aspergillosis.
In Specific Aim 3, we will use RNA sequencing to determine the circadian profile of clock-controlled genes in adipocytes of wt and RIP140 knockout cells, and will use chromatin immunoprecipitation of RIP140 in adipocytes to begin to dissect the role of this co-activator/co-repressor in the circadian biology of this important cell type. We will look for physical association of RIP140 with known clock proteins and transcription factors involved with circadian output, and will generate white adipose tissue-specific knockouts of the clock to probe the significance of circadian regulation to fat metabolism in the mouse, hoping to gain insights into diabetes and metabolic syndrome. These projects are complementary and mutually enriching in that they each rely on genetic and molecular techniques to dissect, and ultimately to understand, the response of cells to their environment and the organization of eukaryotic cells as a function of time.

Public Health Relevance

Biological clocks work in all cells of the human body to regulate metabolism. By studying cells of mice and humans, as well as cells of a fungus, we can understand how clock control works, how its malfunction leads to diseases like diabetes and mental illness, while also suggesting a therapy for life-threatening fungal infections.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM083336-25
Application #
8601709
Study Section
Molecular Genetics B Study Section (MGB)
Program Officer
Sesma, Michael A
Project Start
1989-09-01
Project End
2015-12-31
Budget Start
2014-01-01
Budget End
2014-12-31
Support Year
25
Fiscal Year
2014
Total Cost
$644,370
Indirect Cost
$246,611
Name
Dartmouth College
Department
Genetics
Type
Schools of Medicine
DUNS #
041027822
City
Hanover
State
NH
Country
United States
Zip Code
03755
Dasgupta, Arko; Fuller, Kevin K; Dunlap, Jay C et al. (2016) Seeing the world differently: variability in the photosensory mechanisms of two model fungi. Environ Microbiol 18:5-20
Wang, Bin; Zhou, Xiaoying; Loros, Jennifer J et al. (2016) Alternative Use of DNA Binding Domains by the Neurospora White Collar Complex Dictates Circadian Regulation and Light Responses. Mol Cell Biol 36:781-93
Wang, Zheng; Li, Ning; Li, Jigang et al. (2016) The Fast-Evolving phy-2 Gene Modulates Sexual Development in Response to Light in the Model Fungus Neurospora crassa. MBio 7:e02148
Olivares-Yañez, Consuelo; Emerson, Jillian; Kettenbach, Arminja et al. (2016) Modulation of Circadian Gene Expression and Metabolic Compensation by the RCO-1 Corepressor of Neurospora crassa. Genetics 204:163-76
Hurley, Jennifer M; Loros, Jennifer J; Dunlap, Jay C (2016) The circadian system as an organizer of metabolism. Fungal Genet Biol 90:39-43
Hurley, Jennifer; Loros, Jennifer J; Dunlap, Jay C (2015) Dissecting the mechanisms of the clock in Neurospora. Methods Enzymol 551:29-52
Oliveira, Anderson G; Stevani, Cassius V; Waldenmaier, Hans E et al. (2015) Circadian control sheds light on fungal bioluminescence. Curr Biol 25:964-8
Emerson, Jillian M; Bartholomai, Bradley M; Ringelberg, Carol S et al. (2015) period-1 encodes an ATP-dependent RNA helicase that influences nutritional compensation of the Neurospora circadian clock. Proc Natl Acad Sci U S A 112:15707-12
Fuller, Kevin K; Loros, Jennifer J; Dunlap, Jay C (2015) Fungal photobiology: visible light as a signal for stress, space and time. Curr Genet 61:275-88
Fuller, Kevin K; Chen, Shan; Loros, Jennifer J et al. (2015) Development of the CRISPR/Cas9 System for Targeted Gene Disruption in Aspergillus fumigatus. Eukaryot Cell 14:1073-80

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