Analyzing G1–S Transcriptional Control

G1–S transcriptional control involves the coordination of the expression of a large set of co-regulated genes as a function of cell cycle progression (Bertoli et al., Nat Rev Mol Cell Biol 14:518–528, 2013). Confining transcription to the G1 phase of the cell cycle requires the regulation of specific transcription factor activity through either co-factors or regulation of promoter DNA binding. Therefore, the analysis of G1–S transcriptional control involves cell cycle synchronization and monitoring cell cycle synchrony, in order to establish DNA binding of G1–S transcription factors to G1–S promoters and to investigate changes in gene expression during the different phases of the cell cycle. Here, we describe a cell cycle synchrony method and ways to monitor synchrony. We also describe a chromatin immunoprecipitation (ChIP) method to locate G1–S transcription factor components to promoters and a quantitative PCR (qPCR) protocol to determine gene expression. Defining the binding dynamics of G1–S transcription factors and changes in gene expression during the cell cycle should provide new insights into the mechanism that control G1–S transcription and will allow for investigation of the biological relevance of confining gene expression to G1.

Source: Springer protocols feed by Cell Biology - June 10, 2014 Category: Cytology Source Type: news