Human MARC1 p.165T variant causes mislocalization of MARC1 protein in hepatic cells.

HuH-7 cells were transfected with plasmids containing wild type or 2 common genetic variants (p.A165T and p.M187K) of human MARC1 (with C terminal Flag tag and gene expression driven under CVM promoter). 48 hours after transfection cells were incubated with Mitotracker Orange CMTMRos staining (Thermo Fisher) solution in serum free media for 20 minutes, cells were washed, fixed, permeabilized and blocked as described in Methods. After additional washing, cells were incubated with Flag antibody (Sigma) for 1 hour, washed and incubated with Alexa Fluor 647 conjugated secondary antibody. After final wash, cells were incubated in mounting medium with DAPI (Ibidi) and imaged with Leica confocal microscope. The experiment was repeated once with the equivalent results. (TIF)