Investigating the Effect of Negative Arm Protein Conformations on the Circadian Post-Transcriptional Regulation of Cellulases.
EMSL Project ID
60407
Abstract
Biofuel production using cellulases extracted from fungi has the potential to be a major resource to replace fossil fuels. Regulating the synthesis of these and other enzymes is crucial for the economics of biofuel production from lignocellulose. However, environmental influences on cellular metabolism are known to modulate the amount of cellulases that can be produced, and these influences are poorly understood. Circadian rhythms, the highly conserved, roughly 24-hour, physiological cycles that tune cellular functions to the day/night sequence, are one of the major environmental influences and a thorough understanding of clock regulation over metabolism could be utilized to maximize cellulase production.
Circadian rhythms are timed via a highly regulated transcription-translation based negative feedback loop, or clock. The current paradigm for clock regulation over cellular physiology is that transcriptional activity from the activating complex of the transcription-translation negative feedback loop drives the expression of a host of gene promoters that modulate organismal behavior. However, preliminary data from collaborative work with the JGI and EMSL suggests that circadian regulation is imparted on cellular physiology beyond the level of transcription and that the repressive complex of the feedback loop may play a role in this regulation independent of its modulation of activating complex activity.
The principal goal of this user proposal is to identify the mechanistic underpinnings of repressive complex regulation over cellular physiology. Based on our discovery that the repressive complex proteins are classified as Intrinsically Disordered Proteins (proteins that possess little to no three-dimensional structure), we hypothesize that the intrinsically disordered nature of the repressive complex enables the clock to not only regulate the core circadian feedback loop but also cellular output through the modulation of repressive complex protein conformations. Our aims include the mapping of repressive complex conformational shifts over the circadian day and the analysis of the effect these repressive complex conformational shifts have on circadian regulation over cellular physiology. To carry out all of these aims, we will rely on the technical abilities of EMSL and JGI, cutting-edge BioSANS and cryo-EM/NMR analysis, sequencing capabilities, and protein production pipelines. With this data, we can generate models of circadian regulation of metabolism due to protein conformational shifts that can be immediately tested and validated in vivo.
The long-term goals of our project are to identify the link between time of day and metabolic regulation in an effort to better understand microbial systems to maximize the manufacture of components for energy production. As the ease of biochemical and genetic manipulation in Neurospora crassa (Neurospora) is unparalleled in any other fungal or clock model system, we will address our hypotheses utilizing Neurospora. Timekeeping mechanisms are highly conserved and, in so far as clocks have been described in many phyla within the fungal kingdom, understanding circadian control of metabolism in Neurospora will contribute broadly to understanding metabolic control of cellulases in many important fungal species as well as in plants and bacteria. The success of the overarching goal of this project will provide insights into how to factor clock regulation of metabolism into biofuel manufacture.
Project Details
Project type
FICUS Research
Start Date
2022-10-01
End Date
N/A
Status
Active
Released Data Link
Team
Principal Investigator
Team Members