The molecular events leading to globin gene transcriptional activation
01 / 2004 - 07 / 2008
Nederlandse Organisatie voor Wetenschappelijk Onderzoek (NWO)
We recently showed a stochastic basis in the activation of a and b globin genes in erythroid cells. Characterization of allelic globin expression patterns in single cells showed a significant proportion of cells (>25%) having failed to activate at least one globin allele, an observation that was reflected at both the nuclear primary transcript and cytoplasmic mRNA levels. Clonal assays showed these patterns to be clonally inherited, while analysis of a multicopy transgenic human b globin locus showed an all-or-none effect in the activation of all the genes in the multiple locus integrants. These observations indicated a critical 'window of opportunity' in erythroid differentiation during which decisions for globin activation (or not) are taken and subsequently fixed. In recent work using proerythroblastic cell lines that can be induced to undergo synchronous terminal differentiation, we determined the time interval during which globin genes become activated to be between 6 and 12 hours post induction. Thus, this interval most likely represents the window of opportunity for globin gene activation. It is our aim is to define in molecular terms this critical time interval for globin gene activation and to understand the parameters that influence decisions taken during this window for activation (or not). This translates into the key objectives of defining changes in chromatin structure and transcription factor binding in the a and b globin loci around the critical window of activation and of manipulating different parameters, such as transcription factor dosage or binding sites, in assessing their influence on decisions taken during the critical window.