Mitochondria uptake and buffer huge amounts of calcium via a channel called the "uniporter."  Calcium uptake into mitochondria activates the TCA cycle and is believed to serve as a key signal for coordinating mitochondrial ATP production with its utilization in the cytosol.  The uniporter has been studied for decades but its molecular composition has remained a complete mystery.  Our laboratory used comparative genomics and proteomics to discover all of the molecular components of the uniporter.  This strategy led initially to the identification of MICU1, which we now understand to be one of the gate-keepers of the uniporter.  Using integrative genomics, we could then discover MCU as the pore-forming subunit.  Although MCU fulfilled all of the criteria for serving as the uniporter’s pore, it was not sufficient for reconstituting uniporter activity.  We then performed an affinity purification of the uniporter holocomplex to identify the remaining distinct component, EMRE, a tiny trans-membrane protein required for activity of MCU and of coupling to the calcium sensing activity of MICU1.  With these components in hand, we achieved the first in vivo reconstitution of this channel complex in yeast, an organism whose mitochondria lack uniporter activity and machinery.  Detailed biophysical studies indicate that the MICU1/MICU2 heterodimer sense calcium in the inter membrane space to serve as an "on-off" switch for trans-membrane protein complex MCU/EMRE, the minimal components required for pore-activity.  Ongoing studies are aimed at a full molecular understanding of this channel complex in health and in disease.