Organisms living on Earth cope daily with changes in their environment. An endogenous oscillator, known as the circadian clock, enables organisms to coordinate metabolism, physiology, and development in anticipation of diurnal and seasonal environmental changes and therefore, enhances fitness. The molecular network underlying the circadian system relies on interlocked transcriptional-translational feedback loops, in addition to multiple layers of regulation at the cellular and organismal level. We propose to deploy a combination of genetic, molecular, biochemical, physiological, bioinformatic, and genome-wide approaches to identify new components of the clockwork, and unravel the intricate wiring of the circadian clock in Arabidopsis. TOC1 is a key element of the core-clock of Arabidopsis. Recent data from our laboratory have shown that TOC1 binds to RNA in vitro. We propose to validate this biochemical property in vivo and study its biological relevance. We will also characterize TOC1 association with the transcriptional machinery and with hub components of hormone signaling pathways, which will be instrumental to understand the role of TOC1 in the circadian system. We further seek to explore the GI molecular network and propose to combine molecular, genetic, and biochemical analyses to study novel partners including transcription factors associated with pathogen responses, hormone and growth signaling. Preliminary data from our laboratory also suggest a potential role of the circadian clock in nitrate uptake. We aim to characterize nitrate uptake dynamics and by utilizing genetic, biochemical, and physiological studies we seek to identify the molecular mechanisms underlying the potential regulation of root physiology by the endogenous clock. By integrating different approaches including bioinformatic, classic plant photobiology tools and mutant screens, we propose to elucidate three distinct layers of the clock: to uncover the signaling pathway of light input to the clockwork; to identify new core-clock elements; and to discover components that are necessary for the oscillator to achieve different developmental or physiological traits. Mechanistic details from our study can be extrapolated outside the circadian field to advance research of other complex regulatory systems and ultimately impact work in human health, disease, and food security. D RELEVANCE (See instructions): This proposal will improve our insights of plant physiology and will provide a comprehensive understanding of the circadian system and the cellular processes associated with it. The study will ultimately impact research in human biological rhythms and treatment of circadian-linked health disorders.