Increased exchange rate of histone H1 on chromatin by exogenous myo-genin expression

(整期优先)网络出版时间:2002-05-15
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ToexplorethemolecularmechanismofchromatinremodelinginvolvedintheregulationoftranscriptionalactivationofspecificgenesbyamyogenicregulatoryfactorMyogenin,weusedNIH3T3fibroblastswithastablyintegratedH1.1-GFPfusionproteintomonitorhistoneH1movementdirectlybyfluorescencerecov-eryafterphotobleaching(FRAP)inlivingcells.TheobservationfromFRAPexperimentswithmyogenintransfectedfibroblastsshowedthattheexchangerateofhistoneH1inchromatinwasobviouslyincreased,indicatingthatforcedexpressionofexogenousMyogenincaninducechromatinremodeling.Thehyper-acetylationofhistonesH3andH4frommyogenintransfectedfibroblastswasdetectedbytriton-acid-urea(TAU)/SDS(2-D)electrophoresisandWesternblotwithspecificantibodiesagainstacetylatedN-terminiofhistonesH3andH4.RT-PCRanalysisindicatedthatthenAChRa-subunitgenewasexpressedinthetrans-fectedfibroblasts.TheseresultssuggestthattheexpressionofexogenousMyogenincaninducechromatinremodelingandactivatethetranscriotionofMvogenin-targetedgeneinnon-musclecells.