Abstract
Rat microsomal aldehyde dehydrogenase (msALDH) has no amino-terminal signal sequence, but instead it has a characteristic hydrophobic domain at the carboxyl terminus (Miyauchi, K., R. Masaki, S. Taketani, A. Yamamoto, A. Akayama, and Y. Tashiro. 1991. J. Biol. Chem. 266:19536- 19542). This membrane-bound enzyme is a useful model protein for studying posttranslational localization to its final destination. When expressed from cDNA in COS-1 cells, wild-type msALDH is localized exclusively in the well-developed ER. The removal of the hydrophobic domain results in the cytosolic localization of truncated proteins, thus suggesting that the portion is responsible for membrane anchoring. The last 35 amino acids of msALDH, including the hydrophobic domain, are sufficient for targeting of E. coli beta-galactosidase to the ER membrane. Further studies using chloramphenicol acetyltransferase fusion proteins suggest that two hydrophilic sequences on either side of the hydrophobic domain play an important role in ER targeting.
Full Text
The Full Text of this article is available as a PDF (6.7 MB).
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Adam S. A., Marr R. S., Gerace L. Nuclear protein import in permeabilized mammalian cells requires soluble cytoplasmic factors. J Cell Biol. 1990 Sep;111(3):807–816. doi: 10.1083/jcb.111.3.807. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Akagi S., Yamamoto A., Yoshimori T., Masaki R., Ogawa R., Tashiro Y. Distribution of protein disulfide isomerase in rat hepatocytes. J Histochem Cytochem. 1988 Dec;36(12):1533–1542. doi: 10.1177/36.12.3192937. [DOI] [PubMed] [Google Scholar]
- Anderson D. J., Mostov K. E., Blobel G. Mechanisms of integration of de novo-synthesized polypeptides into membranes: signal-recognition particle is required for integration into microsomal membranes of calcium ATPase and of lens MP26 but not of cytochrome b5. Proc Natl Acad Sci U S A. 1983 Dec;80(23):7249–7253. doi: 10.1073/pnas.80.23.7249. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Andres D. A., Dickerson I. M., Dixon J. E. Variants of the carboxyl-terminal KDEL sequence direct intracellular retention. J Biol Chem. 1990 Apr 15;265(11):5952–5955. [PubMed] [Google Scholar]
- Blobel G. Intracellular protein topogenesis. Proc Natl Acad Sci U S A. 1980 Mar;77(3):1496–1500. doi: 10.1073/pnas.77.3.1496. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Bordier C. Phase separation of integral membrane proteins in Triton X-114 solution. J Biol Chem. 1981 Feb 25;256(4):1604–1607. [PubMed] [Google Scholar]
- Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1016/0003-2697(76)90527-3. [DOI] [PubMed] [Google Scholar]
- Burnette W. N. "Western blotting": electrophoretic transfer of proteins from sodium dodecyl sulfate--polyacrylamide gels to unmodified nitrocellulose and radiographic detection with antibody and radioiodinated protein A. Anal Biochem. 1981 Apr;112(2):195–203. doi: 10.1016/0003-2697(81)90281-5. [DOI] [PubMed] [Google Scholar]
- Chernoff J., Schievella A. R., Jost C. A., Erikson R. L., Neel B. G. Cloning of a cDNA for a major human protein-tyrosine-phosphatase. Proc Natl Acad Sci U S A. 1990 Apr;87(7):2735–2739. doi: 10.1073/pnas.87.7.2735. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Clark B. J., Waterman M. R. The hydrophobic amino-terminal sequence of bovine 17 alpha-hydroxylase is required for the expression of a functional hemoprotein in COS 1 cells. J Biol Chem. 1991 Mar 25;266(9):5898–5904. [PubMed] [Google Scholar]
- Frangioni J. V., Beahm P. H., Shifrin V., Jost C. A., Neel B. G. The nontransmembrane tyrosine phosphatase PTP-1B localizes to the endoplasmic reticulum via its 35 amino acid C-terminal sequence. Cell. 1992 Feb 7;68(3):545–560. doi: 10.1016/0092-8674(92)90190-n. [DOI] [PubMed] [Google Scholar]
- Fujiki Y., Hubbard A. L., Fowler S., Lazarow P. B. Isolation of intracellular membranes by means of sodium carbonate treatment: application to endoplasmic reticulum. J Cell Biol. 1982 Apr;93(1):97–102. doi: 10.1083/jcb.93.1.97. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Gould S. J., Keller G. A., Hosken N., Wilkinson J., Subramani S. A conserved tripeptide sorts proteins to peroxisomes. J Cell Biol. 1989 May;108(5):1657–1664. doi: 10.1083/jcb.108.5.1657. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Hachiya N., Alam R., Sakasegawa Y., Sakaguchi M., Mihara K., Omura T. A mitochondrial import factor purified from rat liver cytosol is an ATP-dependent conformational modulator for precursor proteins. EMBO J. 1993 Apr;12(4):1579–1586. doi: 10.1002/j.1460-2075.1993.tb05802.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kalderon D., Richardson W. D., Markham A. F., Smith A. E. Sequence requirements for nuclear location of simian virus 40 large-T antigen. Nature. 1984 Sep 6;311(5981):33–38. doi: 10.1038/311033a0. [DOI] [PubMed] [Google Scholar]
- Kornfeld S., Mellman I. The biogenesis of lysosomes. Annu Rev Cell Biol. 1989;5:483–525. doi: 10.1146/annurev.cb.05.110189.002411. [DOI] [PubMed] [Google Scholar]
- Kramer W., Fritz H. J. Oligonucleotide-directed construction of mutations via gapped duplex DNA. Methods Enzymol. 1987;154:350–367. doi: 10.1016/0076-6879(87)54084-8. [DOI] [PubMed] [Google Scholar]
- Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
- Masaki R., Yamamoto A., Tashiro Y. Cytochrome P-450 and NADPH-cytochrome P-450 reductase are degraded in the autolysosomes in rat liver. J Cell Biol. 1987 May;104(5):1207–1215. doi: 10.1083/jcb.104.5.1207. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Matsuura S., Fujii-Kuriyama Y., Tashiro Y. Immunoelectron microscope localization of cytochrome P-450 on microsomes and other membrane structures of rat hepatocytes. J Cell Biol. 1978 Aug;78(2):503–519. doi: 10.1083/jcb.78.2.503. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Mitoma J., Ito A. The carboxy-terminal 10 amino acid residues of cytochrome b5 are necessary for its targeting to the endoplasmic reticulum. EMBO J. 1992 Nov;11(11):4197–4203. doi: 10.1002/j.1460-2075.1992.tb05513.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Miyauchi K., Masaki R., Taketani S., Yamamoto A., Akayama M., Tashiro Y. Molecular cloning, sequencing, and expression of cDNA for rat liver microsomal aldehyde dehydrogenase. J Biol Chem. 1991 Oct 15;266(29):19536–19542. [PubMed] [Google Scholar]
- Munro S., Pelham H. R. A C-terminal signal prevents secretion of luminal ER proteins. Cell. 1987 Mar 13;48(5):899–907. doi: 10.1016/0092-8674(87)90086-9. [DOI] [PubMed] [Google Scholar]
- Munro S. Sequences within and adjacent to the transmembrane segment of alpha-2,6-sialyltransferase specify Golgi retention. EMBO J. 1991 Dec;10(12):3577–3588. doi: 10.1002/j.1460-2075.1991.tb04924.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Nilsson T., Jackson M., Peterson P. A. Short cytoplasmic sequences serve as retention signals for transmembrane proteins in the endoplasmic reticulum. Cell. 1989 Aug 25;58(4):707–718. doi: 10.1016/0092-8674(89)90105-0. [DOI] [PubMed] [Google Scholar]
- Ohsumi T., Ichimura T., Sugano H., Omata S., Isobe T., Kuwano R. Ribosome-binding protein p34 is a member of the leucine-rich-repeat-protein superfamily. Biochem J. 1993 Sep 1;294(Pt 2):465–472. doi: 10.1042/bj2940465. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Okada Y., Frey A. B., Guenthner T. M., Oesch F., Sabatini D. D., Kreibich G. Studies on the biosynthesis of microsomal membrane proteins. Site of synthesis and mode of insertion of cytochrome b5, cytochrome b5 reductase, cytochrome P-450 reductase and epoxide hydrolase. Eur J Biochem. 1982 Feb;122(2):393–402. doi: 10.1111/j.1432-1033.1982.tb05894.x. [DOI] [PubMed] [Google Scholar]
- Okayama H., Berg P. A cDNA cloning vector that permits expression of cDNA inserts in mammalian cells. Mol Cell Biol. 1983 Feb;3(2):280–289. doi: 10.1128/mcb.3.2.280. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Omura T., Takesue S. A new method for simultaneous purification of cytochrome b5 and NADPH-cytochrome c reductase from rat liver microsomes. J Biochem. 1970 Feb;67(2):249–257. doi: 10.1093/oxfordjournals.jbchem.a129248. [DOI] [PubMed] [Google Scholar]
- Ozols J., Heinemann F. S. Chemical structure of rat liver cytochrome b5. Isolation of peptides by high-pressure liquid chromatography. Biochim Biophys Acta. 1982 May 21;704(1):163–173. doi: 10.1016/0167-4838(82)90143-1. [DOI] [PubMed] [Google Scholar]
- Pfeffer S. R., Rothman J. E. Biosynthetic protein transport and sorting by the endoplasmic reticulum and Golgi. Annu Rev Biochem. 1987;56:829–852. doi: 10.1146/annurev.bi.56.070187.004145. [DOI] [PubMed] [Google Scholar]
- Pihlajaniemi T., Helaakoski T., Tasanen K., Myllylä R., Huhtala M. L., Koivu J., Kivirikko K. I. Molecular cloning of the beta-subunit of human prolyl 4-hydroxylase. This subunit and protein disulphide isomerase are products of the same gene. EMBO J. 1987 Mar;6(3):643–649. doi: 10.1002/j.1460-2075.1987.tb04803.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Rapiejko P. J., Gilmore R. Protein translocation across the ER requires a functional GTP binding site in the alpha subunit of the signal recognition particle receptor. J Cell Biol. 1992 May;117(3):493–503. doi: 10.1083/jcb.117.3.493. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Rapoport T. A. Transport of proteins across the endoplasmic reticulum membrane. Science. 1992 Nov 6;258(5084):931–936. doi: 10.1126/science.1332192. [DOI] [PubMed] [Google Scholar]
- Sabatini D. D., Kreibich G., Morimoto T., Adesnik M. Mechanisms for the incorporation of proteins in membranes and organelles. J Cell Biol. 1982 Jan;92(1):1–22. doi: 10.1083/jcb.92.1.1. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Schatz G. 17th Sir Hans Krebs lecture. Signals guiding proteins to their correct locations in mitochondria. Eur J Biochem. 1987 May 15;165(1):1–6. doi: 10.1111/j.1432-1033.1987.tb11186.x. [DOI] [PubMed] [Google Scholar]
- Shibahara S., Müller R., Taguchi H., Yoshida T. Cloning and expression of cDNA for rat heme oxygenase. Proc Natl Acad Sci U S A. 1985 Dec;82(23):7865–7869. doi: 10.1073/pnas.82.23.7865. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Shibahara S., Yoshida T., Kikuchi G. Mechanism of increase of heme oxygenase activity induced by hemin in cultured pig alveolar macrophages. Arch Biochem Biophys. 1979 Oct 15;197(2):607–617. doi: 10.1016/0003-9861(79)90285-6. [DOI] [PubMed] [Google Scholar]
- Strittmatter P., Rogers M. J., Spatz L. The binding of cytochrome b 5 to liver microsomes. J Biol Chem. 1972 Nov 25;247(22):7188–7194. [PubMed] [Google Scholar]
- Suemori H., Kadodawa Y., Goto K., Araki I., Kondoh H., Nakatsuji N. A mouse embryonic stem cell line showing pluripotency of differentiation in early embryos and ubiquitous beta-galactosidase expression. Cell Differ Dev. 1990 Mar;29(3):181–186. doi: 10.1016/0922-3371(90)90120-l. [DOI] [PubMed] [Google Scholar]
- Swift A. M., Machamer C. E. A Golgi retention signal in a membrane-spanning domain of coronavirus E1 protein. J Cell Biol. 1991 Oct;115(1):19–30. doi: 10.1083/jcb.115.1.19. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Takagi Y., Ito A., Omura T. Biogenesis of microsomal aldehyde dehydrogenase in rat liver. J Biochem. 1985 Dec;98(6):1647–1652. doi: 10.1093/oxfordjournals.jbchem.a135435. [DOI] [PubMed] [Google Scholar]
- Verner K., Schatz G. Protein translocation across membranes. Science. 1988 Sep 9;241(4871):1307–1313. doi: 10.1126/science.2842866. [DOI] [PubMed] [Google Scholar]
- Wigler M., Pellicer A., Silverstein S., Axel R. Biochemical transfer of single-copy eucaryotic genes using total cellular DNA as donor. Cell. 1978 Jul;14(3):725–731. doi: 10.1016/0092-8674(78)90254-4. [DOI] [PubMed] [Google Scholar]
