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The mechanism by which cohesin controls gene expression (COHESIN CONTROL)
Start date: May 1, 2012, End date: Apr 30, 2017 PROJECT  FINISHED 

How cells retain, lose, and regain developmental plasticity is poorly understood due to ignorance of the molecular mechanisms regulating gene expression. Each gene is regulated by a unique set of factors and as a consequence the trans-acting factors and cis-acting chromatin modification states regulating a given gene are extremely rare. Transcription is affected by events taking place many thousands of base pairs away from the start, a property enabling developmental and evolutionary plasticity, presumably made possible by DNA looping or translocation of factors along chromatin. Most factors regulating a given gene function at many other genes, complicating interpretation of the consequences of altering the activity of such factors. It is difficult to exclude the possibility that phenotypes are knock-on effects. This could be surmounted if it were possible to observe individual genes in real time in three-dimensional space and to analyse the immediate consequences of altering the activity of regulatory factors. Of these, those capable of inter-connecting DNAs or of translocating large distances along chromatin are of interest. Cohesin is such a factor, composed of three core subunits, a pair of Smc proteins and a kleisin subunit, that interact with each other to form a huge tripartite ring, within which it is thought chromatin fibres are entrapped. In proliferating cells, cohesin’s primary function is to connect sister chromatids during DNA replication until the onset of anaphase, possibly by virtue of co-entrapment within a single ring. However, cohesin is present in most quiescent cells and it is becoming clear that it also regulates gene expression and recombination. This proposal has two goals: To image gene expression on polytene chromosomes and to investigate cohesin’s role during ecdysone-induced transcription. The advantage of this system is that we can use micro-injection of TEV protease to inactivate cohesin. A second goal is to develop the TEV system to

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