New Alternately Colored FRET Sensors for Simultaneous Monitoring of Zn2+ in Multiple Cellular Locations

摘要: Genetically encoded sensors based on fluorescence resonance energy transfer (FRET) are powerful tools for reporting ions, molecules and biochemical reactions in living cells. Here we describe the development of new Zn2+based alternate FRET-pairs that do not involve traditional CFP YFP. Zn2+ is an essential micronutrient plays fundamental roles cell biology. Consequently there a pressing need robust to monitor levels dynamics cells with high spatial temporal resolution. develop suite using FRET pairs, including tSapphire/TagRFP, tSapphire/mKO, Clover/mRuby2, mOrange2/mCherry, mOrange2/mKATE. These were targeted both nucleus cytosol characterized validated Sensors pair Clover/mRuby2 displayed higher dynamic range better signal-to-noise ratio than remaining tested optimal monitoring changes cytosolic nuclear Zn2+. Using green-red sensor cyan-yellow either ER, Golgi, or mitochondria, able uptake simultaneously two compartments, revealing rises quickly, whereas mitochondria all sequester more slowly delay 600–700 sec. Lastly, these studies provide first glimpse reveal buffered at level