Ibarra, Oneida (2017-12). Understanding the Circadian Output Gene Network Using the Clock-Controlled Transcription Factor ADV-1 in Neurospora Crassa. Master's Thesis. Thesis uri icon

abstract

  • The circadian clock is an evolutionary conserved time-keeping mechanism that, through the regulation of rhythmic gene expression, coordinates the physiology of an organism with daily environmental cycles. Clock-controlled transcripts are expressed at all possible phases of the circadian cycle; however, we lack a basic understanding of what controls phase. Phase is defined as a reference point on a waveform cycle relative to an environmental cue. To begin to determine the molecular mechanisms controlling circadian output pathways, we identified the direct targets of the core clock component and transcription factor (TF) WHITE-COLLAR COMPLEX (WCC) in Neurospora crassa using ChIP-seq and found an overrepresentation of 24 TFs (called first tier TFS) in the roughly 200 direct targets. Among these TFs, ADV-1 was shown to be robustly rhythmic, and was the only TF that when deleted was defective in clock-controlled development. Identification of genome-wide binding sites for the 24 first tier TFs revealed that in addition to the WCC, 13 of the TFs bind to the adv-1 promoter, and that ADV-1 feeds back to bind to the promoters of these same TFs. The first tier TFs also bind and potentially co-regulate each other, as well as some of the downstream targets of ADV-1. These data suggested that the first tier TFs that bind to the adv-1 promoter form a network, rather acting independently in linear pathways from the clock to ADV-1. To test this hypothesis, my studies focused on how the TF network controls the period and phase of ADV-1 protein rhythms. I discovered that deletion of only 4 out of 13 TFs that bind to the adv-1 promoter in the network altered the period and/or phase of ADV-1 protein rhythms. These data support a network model for circadian phase regulation, and suggest that one role of the network is to compensate for changes in the levels of components of the network, including changes that might occur during stress.

publication date

  • December 2017