title: Mitochondrial targeting of cytochrome P450 proteins containing NH2-terminal chimeric signals involves an unusual TOM20/TOM22 bypass mechanism.
abstract: Previously we showed that xenobiotic inducible cytochrome P450 (CYP) proteins are bimodally targeted to the endoplasmic reticulum and mitochondria. In this study, we investigated the mechanism of delivery of chimeric signal containing CYP proteins to the peripheral and channel-forming mitochondrial outer membrane translocases (TOMs). CYP+33/1A1 and CYP2B1 did not require peripheral TOM70, TOM20, or TOM22 for translocation through the channel-forming TOM40 protein. In contrast, CYP+5/1A1 and CYP2E1 were able to bypass TOM20 and TOM22 but required TOM70. CYP27, which contains a canonical cleavable mitochondrial signal, required all of the peripheral TOMs for its mitochondrial translocation. We investigated the underlying mechanisms of bypass of peripheral TOMs by CYPs with chimeric signals. The results suggested that interaction of CYPs with Hsp70, a cytosolic chaperone involved in the mitochondrial import, alone was sufficient for the recognition of chimeric signals by peripheral TOMs. However, sequential interaction of chimeric signal containing CYPs with Hsp70 and Hsp90 resulted in the bypass of peripheral TOMs, whereas CYP27A1 interacted only with Hsp70 and was not able to bypass peripheral TOMs. Our results also show that delivery of a chimeric signal containing client protein by Hsp90 required the cytosol-exposed NH(2)-terminal 143 amino acids of TOM40. TOM40 devoid of this domain was unable to import CYP proteins. These results suggest that compared with the unimodal mitochondrial targeting signals, the chimeric mitochondrial targeting signals are highly evolved and dynamic in nature.
authors.affiliation: From the Department of Animal Biology and the Mari Lowe Center for Comparative Oncology, School of Veterinary Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104.
authors.forename: Hindupur K
authors.initials: HK
authors.lastname: Anandatheerthavarada
authors.affiliation: From the Department of Animal Biology and the Mari Lowe Center for Comparative Oncology, School of Veterinary Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104.
authors.forename: Naresh Babu V
authors.initials: NBV
authors.lastname: Sepuri
authors.affiliation: From the Department of Animal Biology and the Mari Lowe Center for Comparative Oncology, School of Veterinary Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104. Electronic address: narayan@vet.upenn.edu.
authors.forename: Narayan G
authors.initials: NG
authors.lastname: Avadhani
mesh_terms: D000818:Animals; D001693:Biological Transport, Active; D003577:Cytochrome P-450 Enzyme System / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D005800:Genes, Fungal; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism; D018841:HSP90 Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism; D066298:In Vitro Techniques; D026901:Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism*; D008930:Mitochondria, Liver / Q000378:metabolism*; D033681:Mitochondrial Membrane Transport Proteins; D000090622:Mitochondrial Precursor Protein Import Complex Proteins; D024101:Mitochondrial Proteins / Q000235:genetics / Q000378:metabolism*; D008954:Models, Biological; D009154:Mutation; D021382:Protein Sorting Signals / Q000235:genetics / Q000502:physiology; D011817:Rabbits; D051381:Rats; D011956:Receptors, Cell Surface; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism
chemical_list: D018840:HSP70 Heat-Shock Proteins; D018841:HSP90 Heat-Shock Proteins; D026901:Membrane Transport Proteins; D033681:Mitochondrial Membrane Transport Proteins; D000090622:Mitochondrial Precursor Protein Import Complex Proteins; D024101:Mitochondrial Proteins; D021382:Protein Sorting Signals; D011956:Receptors, Cell Surface; D018160:Receptors, Cytoplasmic and Nuclear; D011993:Recombinant Fusion Proteins; C530938:TOMM22 protein, rat; C489998:Tomm20 protein, rat; C000707766:Tomm40 protein, rat; D003577:Cytochrome P-450 Enzyme System