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Figure 1


Fig. 1. LETM1 is a mitochondrial inner-membrane protein. (A) Total cell lysates from HeLa cells transfected with vector (lane 1), LETM1 (lane 2) or LETM1-3HA expression plasmid (lane 3) were subjected to immunoblotting using anti-LETM1 antibody. Gray and black arrowheads indicate precursor and mature forms of LETM1, respectively. (B) HeLa cells were fixed, permeabilized and immunostained with antibody against LETM1 (a) and the mitochondrial matrix protein Hsp60 (b). Images were acquired using confocal microscopy. Scale bar, 10 µm. (C) Total lysate (T) from HeLa cells carrying the LETM2 expression plasmid was fractionated and separated into mitochondria-enriched (Mt), microsomal (Ms) and cytosolic (Cyto) fractions. These fractions were subjected to immunoblotting using antibodies against LETM1, LETM2, the mitochondrial outer-membrane protein Tom40, the mitochondrial inner-membrane protein Tim17, the ER protein Sec61β and the cytosolic protein H450. Gray and solid arrowheads indicate precursor and mature forms of LETM1, respectively. (D) The mitochondrial fraction prepared from HeLa cells was treated at 4°C for 30 minutes with trypsin at the indicated concentrations with (lanes 4, 8) or without 1% Triton X-100 (lanes 1-3, 5-7) under isotonic (Mitochondria) or hypotonic (Mitoplasts) conditions and analyzed by immunoblotting with antibodies against LETM1, Tim17, Hsp60 and the outer membrane protein Tom20.





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