Development and characterization of FRET based probe to assess Mitochondria-Lipid droplet contact sites proximity (FLiMP).

Lipid droplets (LD) are organelles playing role in storage of neutral fatty acids and provide free fatty acids to mitochondria for β-oxidation through membrane contact sites. Few tether proteins regulating these contact sites have been discovered, but their mechanism remains unclear. In this work, we try to develop FRET (Förster Resonance Energy Transfer) system with CFP and YFP targeted to LD and mitochondria, respectively, with seamless cloning, to evaluate these contact sites. The probe contains FRB and FKBP domains in each moiety that can be heterodimerized by rapamycin, leading to an increase in energy transfer. This feature is exploited for further characterization of the probe. With this probe, it would help to identify unique pathways and genes responsible in the functioning of the cell and its metabolism.