Isolation and characterization of plant insulators

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Insulators are DNA elements that are able to block inappropriate or unwanted enhancer-promoter communication (enhancer blocker activity) and/or prevent the spread of repressive chromatin into active chromatin (barrier activity). Their action is mediated by protein(s) that recognize them leading to the recruitment of many proteins and ultimately the assembly "insulator bodies" the proposed "functional unit". Thus insulators represent an additional level of transcriptional control and their study represents an opportunity to understand a basic, universal mechanism of gene regulation. Insulators from number of species have been identified including fungi, yeast, fruit fly, chicken and human but not plant. Our work indicated that in Arabidopsis two insulators, UASrpg and BEAD-1C can function as insulators-first published demonstration of insulator activity in a plant. Given the potential importance of insulators in plant gene expression and possible key roles in plant biotechnology for the control of tissue-specific transgene expression, we have embarked on a research programme to isolate and study plant insulators. Our approach is to avoid the use of candidate sequences and to tackle the problem by adapting our transgenic system for the direct selection of DNA elements with insulating activity. Sequence(s) thus isolated can be cloned into other vectors to eliminate insulators that are not functional in a variety of promoter/enhancer combinations, orientation and a range of different species. With such sequences in hand we can devise means to isolate the proteins that recognize them and test models of insulator activity proposed for animal systems. Our long term goal is to understand how these interactions regulate gene expression in plants and their evolution. In addition a function and universal plant insulator would have an impact on the design of a new generation of plant transformation vectors.