Circadian clocks allow organisms to anticipate daily environmental cycles, and regulate physiology and behavior to optimize the timing of resource allocation for improved fitness. Because virtually all aspects of human physiology and behavior are linked to the clock, defects in the circadian system, either through mutation or by living against the clock (e.g. shift work) are associated with a wide range of diseases, including metabolic syndrome, which affects up to 40% of adults over the age of 50. As a consequence of clock regulation of metabolism, many drugs are more effective or more toxic depending on the time of administration. A key strength of our work is our ability to utilize a single-celled model organism, Neurospora crassa, to test novel hypotheses concerning the core mechanisms by which rhythmic gene expression is controlled by the circadian clock, and then apply this information to develop new therapies in human circadian disorders. Learn more about our specific projects below.
CIRCADIAN CLOCK CONTROL OF mRNA TRANSLATION AND TRANSLATION FIDELITY
CIRCADIAN CLOCK CONTROL OF RIBOSOME HETEROGENEITY
A longstanding view in molecular biology is that all of the ribosomes in a cell function identically. Our exciting preliminary data, and those of others, challenge this view, and instead support the idea that heterologous ribosomes globally modulate translation, or more controversially, favor translation of specific mRNAs. In N. crassa, there are 84 expressed cytoplasmic ribosomal protein (r-protein) encoding genes, including 8 r-protein variants that share ~20% identity and >40% similarity with core r-proteins, similar to mammalian cells. To our surprise, we found rhythmic mRNA accumulation for 75% of the r-protein genes, with peaks in mRNA levels occurring at different times of the day. Based on these data, we considered that the composition of some ribosomes changes temporally, and that the existence of day- and night-specific ribosomes might, in combination with clock control of translation initiation and elongation, explain the observed rhythms in protein synthesis from non-rhythmic mRNAs. Using quantitative mass spectroscopy we identified several cytoplasmic r-proteins that cycled in abundance in ribosomes in WT, but not in clock mutant cells, with abundance peaks at different times of day. Our objective is to capitalize on these novel findings to test the hypothesis that the clock regulates r-protein composition, modification, and/or interactions with accessory proteins, and that these changes affect the choice of which mRNAs are rhythmically translated and the accuracy of translation. This work challenges existing paradigms of gene regulation, for both the clock and other inputs. Come work with us to be at the forefront of ribosome biology!
Most of the focus on understanding circadian clock control of gene expression has been at the level of transcription. However, in many systems, there are examples of specific proteins that show a circadian rhythm in levels, while levels of the associated mRNA are relatively constant throughout the day. This suggests that the clock also regulates mRNA translation, but which proteins cycle in abundance in cells over the day, and the mechanisms of clock control of translation, are not well understood. We discovered that the circadian clock regulates the activity of the highly conserved eukaryotic elongation factor 2 (eEF2) and eukaryotic initiation factor 2α subunit (eIF2α). The peak in eIFF2α and eEF2 activities occur at night, coincident with increased overall mRNA translation and energy levels. Using genome-wide approaches (ribosome profiling coupled with RNA-seq), we found that the clock regulates translation of specific mRNAs, in part through mRNA sequestration in P-bodies and stress granules . We have several exciting projects, including clock control of translation fidelity through clock control of tRNA synthetase levels, that combine genetics, molecular biology, genomics, bioinformatics, and biochemistry to attack this problem, and work closely with our colleague and lab neighbor Dr. Mathew Sachs, an expert in mRNA translation in fungi.
Circadian clock-based treatments for jet lag and aging
Fungi serve as important models for circadian clock and aging research. We are using Neurospora crassa to rapidly screen compounds that increase circadian amplitude in aged cultures (these compounds represent potential therapeutics for aging), and compounds that rapidly reset the phase of the clock (these compounds are potential treatments for jet lag). We have identified several compounds that significantly increase circadian amplitude and/or can phase shift the clock. We have several projects to test these compounds in mammals and humans, and to determine their mechanism of action.
CIRCADIAN TRANSCRIPTION FACTOR NETWORKS REGULATE RHYTHMIC PHASE
It is estimated that at least 40% of adults over the age of 50 in the USA suffer from metabolic syndrome, a combination of medical disorders that increases the risk for type 2 diabetes, cardiovascular disease, stroke, and cancer. The prevalence of metabolic syndrome is not only rising in parallel with poor food choices, but also with eating at inappropriate times. Humans now live in a modern world in which our cycles of feeding/fasting and activity/sleep no longer match the light/dark cycle. When food is not properly metabolized due to consumption at the wrong time of day (e.g. by shift workers), downstream processes dependent on that metabolism are altered. This altered metabolism results in inappropriate storage of nutrients, such as fat, and a predisposition for metabolic disease. Substantial data in eukaryotic model systems has shown that the biological clock directly regulates aspects of metabolism through control of rhythmic gene expression. Importantly, mice with a liver-specific deletion of the core clock component Bmal1 exhibit abnormal glucose homeostasis. Despite the clear connections between the clock and metabolism, very little is known about how the clock regulates metabolic homeostasis. We are combining predictive mathematical modeling (in collaboration with Dr. James Galagan at Boston University) with experimental data to determine the role of a circadian transcriptional network on control of the phase of expression of clock-controlled genes (ccgs) involved in cellular metabolism. This information has the potential to uncover new ways to treat, for example, metabolic disease associated with shift-work by manipulating the phase of peak expression of metabolic genes to match the nighttime activity cycle. We are also interested in using this information in exciting new projects to develop synthetic promoters to drive gene expression to peak at any specific phase of the day. If you are interested in synthetic biology and combining modeling with experimental science, please email firstname.lastname@example.org.
Circadian clocks throughout the body play a major role in human health and performance by providing for the local coordination of tissue- or cell-specific processes. Importantly, alterations in these circadian regulated processes occur in certain types of cancer. Furthermore, environmental or genetic deregulation of circadian rhythms increases risk for developing cancer and for poor clinical prognosis in response to treatment. Hence, “chronotherapeutic” strategies have had a significant positive impact on the treatment of many types of cancer by optimizing the specific timing of drug administration to improve the efficacy and reduce the toxicity of chemotherapy. However, circadian biology has not been applied to the development of chronotherapeutic strategies for the treatment of glioblastoma multiforme (GBM), and clinical outcomes for this common primary brain tumor have shown limited improvement over the past 30 years. Because the p38 mitogen activated protein kinase (MAPK) pathway plays a role in the highly invasive properties of GBM, and our data shows that p38 MAPK activity is arrhythmic in glioblastoma cells, as compared to its circadian regulation in astroglial cells, our goal, in collaboration with Dr. David Earnest and Gerard Toussaint at Texas A&M, is to examine the time-of-day-specific effects of VX-745, a potent and highly specific p38 MAPK inhibitor, in arresting glioma cell invasiveness in an in vivo model system to better understand the mechanism by which circadian regulation of p38 MAPK activity regulates glioma cell invasion, and to develop novel chronotherapeutic strategies for GBM treatment with p38 inhibitors to improve their efficacy and patient outcomes. Join us to learn more about the role of the clock in human health!