CycliX - Transcription regulatory networks of three interacting cycles
Cyclic regulatory programs are fundamental building blocks of living systems. In this project we aim to study three of these cycles, the circadian, cell-division, and nutrient-response cycles, and the interrelationships among them. The circadian cycle involves orchestrated genome regulation depending on a self-sustaining clock that adjusts to day and night conditions. The cell-division cycle corresponds to a series of molecular events that leads to genome duplication and segregation during cell proliferation, and is characterized by checkpoints that allow cycle progression only if certain conditions have been met. The nutrient-response cycle is initiated by exposure to nutrients, which leads a "fasting" cell to undergo a program of gene expression that returns to the fasting state once the nutrients have been exhausted.
The phases of each of these cycles are characterized by genomic states that reflect a response to phase-specific signals. These genomic states set up transcriptional programs that then motor the cycle forward to the next phase. Although each individual cycle has been studied extensively, we still know little about the global genomic responses to the cycles and their associated transcriptional regulatory programs. Moreover, the current knowledge is largely limited to RNA polymerase (pol) II mRNA-encoding genes, with few or no studies examining pol III transcription units or pol II non-coding RNA (ncRNA) genes. We know even less about how the three cycle transcription programs interconnect and influence each other.
The "CycliX: transcription regulatory networks of three interconnecting cycles" project aims at a quantitative and comprehensive understanding of the global genomic responses and pol II and III transcriptional programs characterizing each cycle. Importantly CycliX will focus on how and when these transcription programs intersect at a shared "core" regulatory network to assure proper integration and coordination among the three cycling systems.
A special aspect of this project is the analysis of pol III transcription alongside that of pol II. Pol III synthesizes ncRNAs (e.g., tRNAs, 5S RNA, snRNAs, some microRNAs) that are highly regulated during nutrient responses and in proliferating versus non-proliferating cells, as they must double in mass upon each cell division but owing to their great stability need little synthesis in resting cells. Nevertheless, pol III transcription analyses on a genomic scale have been lacking because most of these transcription units are repeated in the genome, making hybridization approaches essentially impossible.
As described in the proposal, we find that by ChIP-Seq analysis, we can determine the transcription state of individual repeated pol III transcribed genes. Thus, in this proposal, except for the ribosomal RNA genes transcribed by pol I and the few genes transcribed by the recently discovered spRNAP-IV, we interrogate all transcription activity.
The project is divided into three subprojects, which combine experimental and computational approaches. In subproject 1, titled "Identification of cyclic nodes", we will analyze the genomic states at different phases of the three cycles. We will quantify transcriptional activity by measuring levels of pol II and III on the entire genome and by mapping certain histone marks, using chromatin immunoprecipitation (ChIP) with antibodies directed against the factors of interest followed by deep sequencing (ChIP-Seq).
We will thus identify exhaustive sets of pol II and III genes whose transcription activity varies during a cycle as well as sets of genes whose activity varies during at least two of the three cycles, referred to as "cyclic-nodes" in this proposal. These efforts will be integrated with extensive mining and bioinformatics analyses of existing functional datasets, e.g. mRNA accumulation, during these cycles.
CyCliX Home Page (click on image)
