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. 2014 Mar 6;53(5):726–737. doi: 10.1016/j.molcel.2014.01.013

Figure 1.

Figure 1

MICU1 and MICU2 Interact in the Mitochondrial Intermembrane Space

(A) HeLa cells were transfected with MICU1-HA and/or MICU2-Flag. After 24 hr the cells were fixed and immunocytochemistry was performed with αHA, αFlag, or αTOM20 antibodies followed by incubation with Alexa 488 (pseudocolored green) and Alexa 555 (pseudocolored red) -conjugated secondary antibodies. The scale bars represent 10 μm.

(B) HeLa cells were transfected with GFP and mCherry, MCU-GFP and MCU-mCherry, MICU1-GFP and MICU1-mCherry, MICU2-GFP and MICU2-mCherry, MICU2-GFP and MICU1-mCherry, and MICU1-GFP and MICU2-mCherry and analyzed after 24 hr. Images of donors and acceptors were taken before and after photobleaching of the indicated region (white box). FRET was calculated as detailed in Experimental Procedures. The histogram bar diagram shows FRET efficiency of the indicated donor and acceptor pairs. Data are presented as mean ± SD. Descriptive statistics can be found in Table S1. ∗p < 0.05 compared to control. The scale bars represent 10 μm.

(C) The same amount of mitoplasts isolated from mouse liver was subjected to the indicated proteolytic treatment, processed for electrophoresis and western blot, and probed for MICU1 and MCU.