Two Homologous Cytochromes <i>b</i><sub>5</sub> in a Single Cell

Lederer, Florence; Ghrir, Rachid; Guiard, Bernard; Cortial, Sylvie; Ito, Akio

doi:10.1111/j.1432-1033.1983.tb07330.x

Cited by 93 publications

(38 citation statements)

References 41 publications

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“…Proteins of this class, which have been called tailanchored (TA) (Kutay et al, 1993), lack an N-terminal signal sequence and reach their destination within the cell by posttranslational mechanisms Kutay et al, 1993). TA proteins identified on the MOM include the protooncogene bcl-2 (Akao et al, 1994), the mitochondrial isoform of cytochrome b(5) [outer membrane b(5) or OM b(5); Lederer et al, 1983;D'Arrigo et al, 1993], TOM components TOM 5 and 6 (Neupert, 1997), a synaptojanin binding protein (Nemoto and De Camilli, 1999), an alternatively spliced isoform of vesicle associated membrane protein, VAMP-1B (Isenmann et al, 1998). An important problem is the mechanism through which these proteins discriminate between the MOM and the endoplasmic reticulum (ER) membrane, because the latter is the target for many polypeptides with similar topology.…”

Section: Introductionmentioning

confidence: 99%

“…Mammalian cyt b(5) exists in two homologous isoforms (Lederer et al, 1983), which are localized specifically to the MOM [OM b(5)] or to the ER [ER b(5)], with hardly any overlap between the two distributions . Work in our laboratory showed that the tail region of OM b(5) contains the information for localization to the MOM (De Silvestris et al, 1995).…”

Section: Introductionmentioning

confidence: 99%

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Targeting of a Tail-anchored Protein to Endoplasmic Reticulum and Mitochondrial Outer Membrane by Independent but Competing Pathways

Borgese

Gazzoni

Barberi

et al. 2001

MBoC

124

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Many mitochondrial outer membrane (MOM) proteins have a transmembrane domain near the C terminus and an N-terminal cytosolic moiety. It is not clear how these tail-anchored (TA) proteins posttranslationally select their target, but C-terminal charged residues play an important role. To investigate how discrimination between MOM and endoplasmic reticulum (ER) occurs, we used mammalian cytochrome b(5), a TA protein existing in two, MOM or ER localized, versions. Substitution of the seven C-terminal residues of the ER isoform or of green fluorescent protein reporter constructs with one or two arginines resulted in MOM-targeted proteins, whereas a single C-terminal threonine caused promiscuous localization. To investigate whether targeting to MOM occurs from the cytosol or after transit through the ER, we tagged a MOM-directed construct with a C-terminal N-glycosylation sequence. Although in vitro this construct was efficiently glycosylated by microsomes, the protein expressed in vivo localized almost exclusively to MOM, and was nearly completely unglycosylated. The small fraction of glycosylated protein was in the ER and was not a precursor to the unglycosylated form. Thus, targeting occurs directly from the cytosol. Moreover, ER and MOM compete for the same polypeptide, explaining the dual localization of some TA proteins.

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Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Targeting of a Tail-anchored Protein to Endoplasmic Reticulum and Mitochondrial Outer Membrane by Independent but Competing Pathways

Borgese

Gazzoni

Barberi

et al. 2001

MBoC

124

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“…Although these similarities in the targeting signals allow some TA proteins to exhibit dual localization within the cell, most TA proteins are targeted exclusively to one organelle or the other . Notably, only mammalian cells have been shown to contain two distinct isoforms of Cb5 that are localized specifically to the ER or to the mitochondrial outer membrane (Ito, 1980;Lederer et al, 1983;D'Arrigo et al, 1993). Despite intensive efforts over the past few years, details related to the nature of the cellular machinery involved in targeting and membrane integration of TA proteins to either ER or mitochondrial membranes in any organism have not been resolved (reviewed in Borgese et al, 2003; for an alternate point of view, see Abell et al, 2004).…”

Section: Introductionmentioning

confidence: 99%

Novel Targeting Signals Mediate the Sorting of Different Isoforms of the Tail-Anchored Membrane Protein Cytochrome b₅ to Either Endoplasmic Reticulum or Mitochondria

et al. 2004

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Tail-anchored membrane proteins are a class of proteins that are targeted posttranslationally to various organelles and integrated by a single segment of hydrophobic amino acids located near the C terminus. Although the localization of tail-anchored proteins in specific subcellular compartments in plant cells is essential for their biological function, the molecular targeting signals responsible for sorting these proteins are not well defined. Here, we describe the biogenesis of four closely related tung (Aleurites fordii) cytochrome b5 isoforms (Cb5-A, -B, -C, and -D), which are small tail-anchored proteins that play an essential role in many cellular processes, including lipid biosynthesis. Using a combination of in vivo and in vitro assays, we show that Cb5-A, -B, and -C are targeted exclusively to the endoplasmic reticulum (ER), whereas Cb5-D is targeted specifically to mitochondrial outer membranes. Comprehensive mutational analyses of ER and mitochondrial Cb5s revealed that their C termini, including transmembrane domains (TMD) and tail regions, contained several unique physicochemical and sequence-specific characteristics that defined organelle-specific targeting motifs. Mitochondrial targeting of Cb5 was mediated by a combination of hydrophilic amino acids along one face of the TMD, an enrichment of branched β-carbon–containing residues in the medial portion of the TMD, and a dibasic -R-R/K/H-x motif in the C-terminal tail. By contrast, ER targeting of Cb5 depended primarily upon the overall length and hydrophobicity of the TMD, although an -R/H-x-Y/F- motif in the tail was also a targeting determinant. Collectively, the results presented provide significant insight into the early biogenetic events required for entry of tail-anchored proteins into either the ER or mitochondrial targeting pathways

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“…In mammals, the reductase in its various locations is molecularly identical (Meldolesi et al, 1980). Two distinct forms of cytochrome b 5 have been shown to exist in rat liver (Lederer et al, 1983). Cytochrome b 5 localized in the outer mitochondrial membrane is distinguishable from the microsomal form in spectral and immunological properties as well as in primary structure.…”

Section: Tablementioning

confidence: 98%

HEPATIC, EXTRAHEPATIC, MICROSOMAL, AND MITOCHONDRIAL ACTIVATION OF THEN-HYDROXYLATED PRODRUGS BENZAMIDOXIME, GUANOXABENZ, AND RO 48-3656 ([[1-[(2S)-2-[[4-[(HYDROXYAMINO)IMINOMETHYL]BENZOYL]AMINO]-1-OXOPROPYL]-4-PIPERIDINYL]OXY]-ACETIC ACID)

Clement¹,

Mau²,

Deters³

et al. 2005

Drug Metab Dispos

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ABSTRACT:In previous studies, it was shown that liver microsomes from rabbit, rat, pig, and human are involved in the reduction of N-hydroxylated amidines, guanidines, and amidinohydrazones of various drugs and model compounds (Drug Metab Rev 34: 565-579). One responsible enzyme system, the microsomal benzamidoxime reductase, consisting of cytochrome b 5 , its reductase, and a cytochrome P450 isoenzyme, was isolated from pig liver microsomes (J Biol Chem 272:19615-19620). Further investigations followed to establish whether such enzyme systems are also present in microsomes of other organs such as brain, lung, and intestine. In addition, the mitochondrial reduction in human and porcine liver and kidney preparations was studied. . Interestingly, preparations of all tested organs were capable of reducing the three compounds. The highest specific rates were found in kidney followed by liver, brain, lung, and intestine, and usually the mitochondrial reduction rates were superior. From the determined characteristics, similarities between the enzyme systems in the different organs and organelles were detected. Furthermore, properties of the benzamidoxime reductase located in the outer membrane of pig liver mitochondria were studied. In summary, these results demonstrate that in addition to the microsomal reduction, mitochondria are involved to a great extent in the activation of amidoxime prodrugs. The importance of extrahepatic metabolism in the reduction of N-hydroxylated prodrugs is demonstrated.Numerous drugs and drug candidates contain strongly basic functional groups, such as guanidines, amidinohydrazones, and amidines. Because of their strong basicity, they are protonated under physiological conditions and usually not absorbed in the gastrointestinal tract. In our laboratory, a prodrug principle was developed in particular for amidines by N-hydroxylating this functional group to the corresponding amidoximes, substances with a less basic functional group. Consequently, they are not protonated under physiological conditions and can easily be absorbed as free bases. By creating these prodrugs of the active principle, the oral bioavailability of amidines can be increased. Thus amidoximes and similar functional groups can be used as prodrugs for amidines and related groups.The prodrug principle was developed for pentamidine (Clement, 1993) and was then applied to other amidines such as trypanocidal compounds (Zhou et al., 2004), glycoprotein IIIa/IIb receptor antagonists, such as sibrafiban (Weller et al., 1996), and the thrombin inhibitor ximelagatran, which was recently approved and is the first orally available direct thrombin inhibitor on the market (Gustafsson et al., 2001). Benzamidoxime is a model compound for such amidoxime prodrugs, which are known to be reduced by liver microsomes from different species as well as by the purified enzyme system from pig liver (Clement et al., 1997). Therefore, the reduction of this substrate and its HPLC analysis can be taken as a reliable activity assay (Fig. 1).Sibrafiban...

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Two Homologous Cytochromes b₅ in a Single Cell

Cited by 93 publications

References 41 publications

Targeting of a Tail-anchored Protein to Endoplasmic Reticulum and Mitochondrial Outer Membrane by Independent but Competing Pathways

Targeting of a Tail-anchored Protein to Endoplasmic Reticulum and Mitochondrial Outer Membrane by Independent but Competing Pathways

Novel Targeting Signals Mediate the Sorting of Different Isoforms of the Tail-Anchored Membrane Protein Cytochrome b₅ to Either Endoplasmic Reticulum or Mitochondria

HEPATIC, EXTRAHEPATIC, MICROSOMAL, AND MITOCHONDRIAL ACTIVATION OF THEN-HYDROXYLATED PRODRUGS BENZAMIDOXIME, GUANOXABENZ, AND RO 48-3656 ([[1-[(2S)-2-[[4-[(HYDROXYAMINO)IMINOMETHYL]BENZOYL]AMINO]-1-OXOPROPYL]-4-PIPERIDINYL]OXY]-ACETIC ACID)

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Two Homologous Cytochromes b5 in a Single Cell

Cited by 93 publications

References 41 publications

Targeting of a Tail-anchored Protein to Endoplasmic Reticulum and Mitochondrial Outer Membrane by Independent but Competing Pathways

Targeting of a Tail-anchored Protein to Endoplasmic Reticulum and Mitochondrial Outer Membrane by Independent but Competing Pathways

Novel Targeting Signals Mediate the Sorting of Different Isoforms of the Tail-Anchored Membrane Protein Cytochrome b5 to Either Endoplasmic Reticulum or Mitochondria

HEPATIC, EXTRAHEPATIC, MICROSOMAL, AND MITOCHONDRIAL ACTIVATION OF THEN-HYDROXYLATED PRODRUGS BENZAMIDOXIME, GUANOXABENZ, AND RO 48-3656 ([[1-[(2S)-2-[[4-[(HYDROXYAMINO)IMINOMETHYL]BENZOYL]AMINO]-1-OXOPROPYL]-4-PIPERIDINYL]OXY]-ACETIC ACID)

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Two Homologous Cytochromes b₅ in a Single Cell

Novel Targeting Signals Mediate the Sorting of Different Isoforms of the Tail-Anchored Membrane Protein Cytochrome b₅ to Either Endoplasmic Reticulum or Mitochondria