Characterization of the Novel Mitochondrial Protein Import Component, Tom 34, in Mammalian Cells

Abstract

Tom 34 is a newly-found component of the mitochondrial protein import machinery in mammalian cells with no apparent counterpart in fungi. RNA blot and immunoblot analyses showed that the expression of Tom 34 varies among tissues and differs from that of the core translocase component Tom 20. In contrast to a previous report [Nuttal, S. D. et al. (1997) DNA Cell Biol. 16, 1067-1074], the present study using a newly-prepared anti-Tom 34 antibody with a high titer showed that Tom 34 is present largely in the cytosolic fraction and partly in the mitochondrial and membrane fractions after fractionation of tissues and cells, and that the membrane-associated form is largely extractable with 0.1M sodium carbonate. The in vitro import of preproteins into isolated rat mitochondria was strongly inhibited by ΔTom 34 which lacks the NH²-terminal hydrophobic region of human Tom 34 (hTom 34). Import was also strongly inhibited by anti-hTom 34. In pulsechase experiments using COS-7 cells, pre-ornithine transcarbamylase (pOTC) was rapidly processed to the mature form. Coexpression of hTom 34 resulted in a stimulation of pOTC processing, whereas the coexpression of hTom 34 antisense RNA caused inhibition. The results confirm that Tom 34 plays a role in mitochondria) protein import in mammals, and suggest it to be an ancillary component of the translocation machinery in mammalian cells.