The circadian clock is an evolutionarily conserved time-keeping mechanism that, through the regulation of rhythmic gene expression, coordinates the physiology of an organism with daily environmental cycles. Because virtually all aspects of human physiology and behavior are linked to the clock, abnormalities in the circadian system are associated with a wide range of diseases such as sleep disorders, cardiovascular disease, metabolic syndrome, and cancer. In addition, the clock controls temporal aspects of drug metabolism and vulnerability to cytotoxic agents. Thus, knowing what genes and proteins are regulated by the clock, and the mechanisms of this regulation, are necessary to understand clock-associated diseases and rhythmic drug metabolism. The primary focus of research on circadian control of gene expression has been at the transcriptional level. However, substantial evidence exists for a role of the clock in regulating rhythmic translation. Despite this, essentialy nothing is known about how translation is controlled by the clock. In the previous funding period, using the model system Neurospora crassa, we discovered that the circadian clock controls the phosphorylation of two highly conserved central regulators of mRNA translation, eukaryotic elongation factor 2 (eEF2), and eukaryotic initiation factor 2? (eIF2?), both peaking in activity a night. In addition, we found that the clock controls the levels of S6 kinase, a key regulator of th highly conserved initiation factor eIF4B. Using high throughput RNA-seq and ribosome profiling in wild type cells, and cells that are defective in rhythmic activity of the translation factors, w will determine if the clock regulates translation of all or just some proteins. Results from these studies will allow us to determine which proteins cycle in abundance, and using molecular genetic techniques, to decipher the mechanism used by the clock to control translation elongation (Aim 1) and initiation (Aim 2). This study will fill major gaps in our knowledge regarding which proteins accumulate with a circadian rhythm and how this regulation is controlled, and thus give us a much deeper understanding of what aspects of cell physiology and metabolism are regulated by the clock.
Circadian clock control of protein synthesis is understudied. Our investigation of clock control of translation initiation and elongation, fueled by the discover that conserved regulatory proteins required for translation are controlled by the clock, has broad implications for understanding how the clock regulates rhythmic protein production, important in rhythmic control of drug metabolism, as well as cancer, heart disease, and metabolic disease associated with shift work and genetic mutations of the clock.
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