Transcription factor mobility

Division Of Basic Sciences - Nci

Investigators

Linked publications & trials

Abstract

Expression of many important genes is cyclical, and may incorporate multiple cycles at the different time scale, such as circadian or ultradian cycles on the scale of hours, and fast and short bursts of transcription on the scale of minutes. The mechanisms of this cycling are poorly understood, but very important for the correct application of the drugs. At the transcription level, gene expression is controlled by the accessibility of the regulatory elements within DNA - promoters and enhancers. Research in yeast and mammalian cells indicates that modulation of chromatin accessibility occurs through interaction of transcription factors (TF) and chromatin remodelers. Binding of certain TF to their target DNA sequences is highly dynamic, on the scale of seconds. Also, some TF undergo the "slow" cycling on the scale of minutes, which consist of alternating "ON" and "OFF" gene states. It is unclear how this TF cycling is related to the transcriptional cycling. We are interested in molecular mechanisms of the TF cycling on promoters and its correlation with transcription cycles. We use as a model yeast gene CUP1 involved in heavy metal stress, and HSP104, involved in heat shock stress. CUP1 is activated by copper-bound TF Ace1p. HSP104 is activated by HSF1. We are studying dynamics of these TF factors on specific promoters by Single Molecule Tracking (SMT). Previously, we have built a custom instrument for SMT and optimized SMT data interpretation. We have developed methods for SMT in yeast cell nuclei and methods of performing SMT on specific promoters.We constructed reporter genes with promoters of either CUP1 or HSP104. Transcription may be visualized by smFISH (Fluorescence In Situ Hybridization) or by live imaging of the fluorescently tagged mRNA. We correlate the dynamics of the TF at the promoter with transcriptional output. We demonstrated that the transient recruitment of TFs is regulated by fast cyclical chromatin remodeling events to ensure the best dynamic transcriptional response. In our recent publication we demonstrate that the slow cycle of transcription consists of the fast cycles (bursts) of expression on individual genes. This is the first genetic model where the superimposed slow and fast cycles of transcription correlate with superimposed slow and fast cycling of the transcriptional factor. Ultimately, these studies will lay the groundwork for the analysis of in vivo interactions of the components of the transcriptional machinery. The technique of correlation of the TF biophysics with transcription modeling that we are in process of developing may be applied to a number of other problems of cellular biology where the information for molecular regulation of transcription is desired.