molecular_biology-2010_autumn-08.ppt

Interferon ? enhancer Enhancers often have binding sites for several transcription factors Transcription factors can bind cooperatively at adjacent sites Architectural factors (with no regulatory domains, i.e. HMG1) can assist assembly Remarkably increase binding affinity for both DNA and machinery HMG1 肩并肩、手挽手，根基稳、魅力足 香肩并立、玉指紧扣，脚如磐石、面若桃花 Regulatory mechanism from a distance Compaction Sliding Looping Why do enhancers act independent of distance and orientation? An enhancer may function by bringing proteins into the vicinity of the promoter. An enhancer does not act on a promoter at the opposite end of a long linear DNA, but becomes effective when the DNA is joined into a circle by a protein bridge. An enhancer and promoter on separate circular DNAs do not interact, but can interact when the two molecules are catenated. Two experiments support the looping model --The essential role of the enhancer is to increase the concentration of activator in the vicinity of the promoter--- Receptors for many steroid and thyroid hormones have a similar organization, with an individual N-terminal region, conserved DNA-binding region, and a C-terminal hormone-binding region Steroid receptors are transcription factors Zinc finger TF Glucocorticoids regulate gene transcription by causing their receptor to transport into the nucleus and bind to an enhancer whose action is needed for promoter function. (or dexamethasone) Activation of Glucocorticoid Receptor (GR) Nuclear shuttling Glucocorticoid receptor (GR) fusion protein induction Cortisol Dexamethasone GR GR GR X X X Dex binds to GR, exposes nucleus localization signal (NLS), allows fusion protein to transport into the nucleus to activate transcription ----A good system to activate the function of nuclear-localized proteins (including transcription factors) Activation Tagging approach in plants Plant transformation phenotypic characterization of mutants locate T-DNA insertion site in Arabidopsis genome (how?) identify the