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. 1995 Dec;7(12):2163–2174. doi: 10.1105/tpc.7.12.2163

Targeting and topology in the membrane of plant 3-hydroxy-3-methylglutaryl coenzyme A reductase.

N Campos 1, A Boronat 1
PMCID: PMC161070  PMID: 8718626

Abstract

The enzyme 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGR) catalyzes the synthesis of mevalonate. This is the first committed step of isoprenoid biosynthesis. A common feature of all known plant HMGR isoforms is the presence of two highly conserved hydrophobic sequences in the N-terminal quarter of the protein. Using an in vitro system, we showed that the two hydrophobic sequences of Arabidopsis HMGR1S function as internal signal sequences. Specific recognition of these sequences by the signal recognition particle mediates the targeting of the protein to microsomes derived from the endoplasmic reticulum. Arabidopsis HMGR is inserted into the microsomal membrane, and the two hydrophobic sequences become membrane-spanning segments. The N-terminal end and the C-terminal catalytic domain of Arabidopsis HMGR are positioned on the cytosolic side of the membrane, whereas only a short hydrophilic sequence is exposed to the lumen. Our results suggest that the plant HMGR isoforms known to date are primarily targeted to the endoplasmic reticulum and have the same topology in the membrane. This reinforces the hypothesis that mevalonate is synthesized only in the cytosol. The possibility that plant HMGRs might be located in different regions of the endomembrane system is discussed.

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Selected References

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  1. Baker A., Schatz G. Sequences from a prokaryotic genome or the mouse dihydrofolate reductase gene can restore the import of a truncated precursor protein into yeast mitochondria. Proc Natl Acad Sci U S A. 1987 May;84(10):3117–3121. doi: 10.1073/pnas.84.10.3117. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Blobel G., Dobberstein B. Transfer of proteins across membranes. I. Presence of proteolytically processed and unprocessed nascent immunoglobulin light chains on membrane-bound ribosomes of murine myeloma. J Cell Biol. 1975 Dec;67(3):835–851. doi: 10.1083/jcb.67.3.835. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bujard H., Gentz R., Lanzer M., Stueber D., Mueller M., Ibrahimi I., Haeuptle M. T., Dobberstein B. A T5 promoter-based transcription-translation system for the analysis of proteins in vitro and in vivo. Methods Enzymol. 1987;155:416–433. doi: 10.1016/0076-6879(87)55028-5. [DOI] [PubMed] [Google Scholar]
  4. Burnett R. J., Maldonado-Mendoza I. E., McKnight T. D., Nessler C. L. Expression of a 3-hydroxy-3-methylglutaryl coenzyme A reductase gene from Camptotheca acuminata is differentially regulated by wounding and methyl jasmonate. Plant Physiol. 1993 Sep;103(1):41–48. doi: 10.1104/pp.103.1.41. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Caelles C., Ferrer A., Balcells L., Hegardt F. G., Boronat A. Isolation and structural characterization of a cDNA encoding Arabidopsis thaliana 3-hydroxy-3-methylglutaryl coenzyme A reductase. Plant Mol Biol. 1989 Dec;13(6):627–638. doi: 10.1007/BF00016018. [DOI] [PubMed] [Google Scholar]
  6. Campos N., Schell J., Palme K. In vitro uptake and processing of maize auxin-binding proteins by ER-derived microsomes. Plant Cell Physiol. 1994 Mar;35(2):153–161. [PubMed] [Google Scholar]
  7. Casey W. M., Keesler G. A., Parks L. W. Regulation of partitioned sterol biosynthesis in Saccharomyces cerevisiae. J Bacteriol. 1992 Nov;174(22):7283–7288. doi: 10.1128/jb.174.22.7283-7288.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Choi D., Ward B. L., Bostock R. M. Differential induction and suppression of potato 3-hydroxy-3-methylglutaryl coenzyme A reductase genes in response to Phytophthora infestans and to its elicitor arachidonic acid. Plant Cell. 1992 Oct;4(10):1333–1344. doi: 10.1105/tpc.4.10.1333. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Chye M. L., Kush A., Tan C. T., Chua N. H. Characterization of cDNA and genomic clones encoding 3-hydroxy-3-methylglutaryl-coenzyme A reductase from Hevea brasiliensis. Plant Mol Biol. 1991 Apr;16(4):567–577. doi: 10.1007/BF00023422. [DOI] [PubMed] [Google Scholar]
  10. Enjuto M., Lumbreras V., Marín C., Boronat A. Expression of the Arabidopsis HMG2 gene, encoding 3-hydroxy-3-methylglutaryl coenzyme A reductase, is restricted to meristematic and floral tissues. Plant Cell. 1995 May;7(5):517–527. doi: 10.1105/tpc.7.5.517. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Genschik P., Criqui M. C., Parmentier Y., Marbach J., Durr A., Fleck J., Jamet E. Isolation and characterization of a cDNA encoding a 3-hydroxy-3-methylglutaryl coenzyme A reductase from Nicotiana sylvestris. Plant Mol Biol. 1992 Oct;20(2):337–341. doi: 10.1007/BF00014504. [DOI] [PubMed] [Google Scholar]
  12. Gilmore R. Protein translocation across the endoplasmic reticulum: a tunnel with toll booths at entry and exit. Cell. 1993 Nov 19;75(4):589–592. doi: 10.1016/0092-8674(93)90476-7. [DOI] [PubMed] [Google Scholar]
  13. Heintze A., Görlach J., Leuschner C., Hoppe P., Hagelstein P., Schulze-Siebert D., Schultz G. Plastidic Isoprenoid Synthesis during Chloroplast Development : Change from Metabolic Autonomy to a Division-of-Labor Stage. Plant Physiol. 1990 Jul;93(3):1121–1127. doi: 10.1104/pp.93.3.1121. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Hesse T., Feldwisch J., Balshüsemann D., Bauw G., Puype M., Vandekerckhove J., Löbler M., Klämbt D., Schell J., Palme K. Molecular cloning and structural analysis of a gene from Zea mays (L.) coding for a putative receptor for the plant hormone auxin. EMBO J. 1989 Sep;8(9):2453–2461. doi: 10.1002/j.1460-2075.1989.tb08380.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Imai Y., Matsushima Y., Sugimura T., Terada M. A simple and rapid method for generating a deletion by PCR. Nucleic Acids Res. 1991 May 25;19(10):2785–2785. doi: 10.1093/nar/19.10.2785. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Jefferson R. A., Kavanagh T. A., Bevan M. W. GUS fusions: beta-glucuronidase as a sensitive and versatile gene fusion marker in higher plants. EMBO J. 1987 Dec 20;6(13):3901–3907. doi: 10.1002/j.1460-2075.1987.tb02730.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Kreuz K., Kleinig H. Synthesis of prenyl lipids in cells of spinach leaf. Compartmentation of enzymes for formation of isopentenyl diphosphate. Eur J Biochem. 1984 Jun 15;141(3):531–535. doi: 10.1111/j.1432-1033.1984.tb08225.x. [DOI] [PubMed] [Google Scholar]
  18. Kunkel T. A., Roberts J. D., Zakour R. A. Rapid and efficient site-specific mutagenesis without phenotypic selection. Methods Enzymol. 1987;154:367–382. doi: 10.1016/0076-6879(87)54085-x. [DOI] [PubMed] [Google Scholar]
  19. 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]
  20. Learned R. M., Fink G. R. 3-Hydroxy-3-methylglutaryl-coenzyme A reductase from Arabidopsis thaliana is structurally distinct from the yeast and animal enzymes. Proc Natl Acad Sci U S A. 1989 Apr;86(8):2779–2783. doi: 10.1073/pnas.86.8.2779. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Lipp J., Flint N., Haeuptle M. T., Dobberstein B. Structural requirements for membrane assembly of proteins spanning the membrane several times. J Cell Biol. 1989 Nov;109(5):2013–2022. doi: 10.1083/jcb.109.5.2013. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Lumbreras V., Campos N., Boronat A. The use of an alternative promoter in the Arabidopsis thaliana HMG1 gene generates an mRNA that encodes a novel 3-hydroxy-3-methylglutaryl coenzyme A reductase isoform with an extended N-terminal region. Plant J. 1995 Oct;8(4):541–549. doi: 10.1046/j.1365-313x.1995.8040541.x. [DOI] [PubMed] [Google Scholar]
  23. McGarvey D. J., Croteau R. Terpenoid metabolism. Plant Cell. 1995 Jul;7(7):1015–1026. doi: 10.1105/tpc.7.7.1015. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Nelson A. J., Doerner P. W., Zhu Q., Lamb C. J. Isolation of a monocot 3-hydroxy-3-methylglutaryl coenzyme A reductase gene that is elicitor-inducible. Plant Mol Biol. 1994 Jun;25(3):401–412. doi: 10.1007/BF00043869. [DOI] [PubMed] [Google Scholar]
  25. Ng D. T., Walter P. Protein translocation across the endoplasmic reticulum. Curr Opin Cell Biol. 1994 Aug;6(4):510–516. doi: 10.1016/0955-0674(94)90069-8. [DOI] [PubMed] [Google Scholar]
  26. Park H., Denbow C. J., Cramer C. L. Structure and nucleotide sequence of tomato HMG2 encoding 3-hydroxy-3-methyl-glutaryl coenzyme A reductase. Plant Mol Biol. 1992 Oct;20(2):327–331. doi: 10.1007/BF00014502. [DOI] [PubMed] [Google Scholar]
  27. Ponce M. R., Micol J. L. PCR amplification of long DNA fragments. Nucleic Acids Res. 1992 Feb 11;20(3):623–623. doi: 10.1093/nar/20.3.623. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Prehn S., Wiedmann M., Rapoport T. A., Zwieb C. Protein translocation across wheat germ microsomal membranes requires an SRP-like component. EMBO J. 1987 Jul;6(7):2093–2097. doi: 10.1002/j.1460-2075.1987.tb02475.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. 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]
  30. Rogers L. J., Shah S. P., Goodwin T. W. Intracellular localization of mevalonate-activating enzymes in plant cells. Biochem J. 1966 May;99(2):381–388. doi: 10.1042/bj0990381. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Schutze M. P., Peterson P. A., Jackson M. R. An N-terminal double-arginine motif maintains type II membrane proteins in the endoplasmic reticulum. EMBO J. 1994 Apr 1;13(7):1696–1705. doi: 10.1002/j.1460-2075.1994.tb06434.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Stermer B. A., Bianchini G. M., Korth K. L. Regulation of HMG-CoA reductase activity in plants. J Lipid Res. 1994 Jul;35(7):1133–1140. [PubMed] [Google Scholar]
  33. Walter P., Blobel G. Preparation of microsomal membranes for cotranslational protein translocation. Methods Enzymol. 1983;96:84–93. doi: 10.1016/s0076-6879(83)96010-x. [DOI] [PubMed] [Google Scholar]
  34. Walter P., Blobel G. Signal recognition particle: a ribonucleoprotein required for cotranslational translocation of proteins, isolation and properties. Methods Enzymol. 1983;96:682–691. doi: 10.1016/s0076-6879(83)96057-3. [DOI] [PubMed] [Google Scholar]
  35. Walter P., Johnson A. E. Signal sequence recognition and protein targeting to the endoplasmic reticulum membrane. Annu Rev Cell Biol. 1994;10:87–119. doi: 10.1146/annurev.cb.10.110194.000511. [DOI] [PubMed] [Google Scholar]
  36. Wessels H. P., Spiess M. Insertion of a multispanning membrane protein occurs sequentially and requires only one signal sequence. Cell. 1988 Oct 7;55(1):61–70. doi: 10.1016/0092-8674(88)90009-8. [DOI] [PubMed] [Google Scholar]
  37. Wickner W. T., Lodish H. F. Multiple mechanisms of protein insertion into and across membranes. Science. 1985 Oct 25;230(4724):400–407. doi: 10.1126/science.4048938. [DOI] [PubMed] [Google Scholar]
  38. Wolin S. L. From the elephant to E. coli: SRP-dependent protein targeting. Cell. 1994 Jun 17;77(6):787–790. doi: 10.1016/0092-8674(94)90124-4. [DOI] [PubMed] [Google Scholar]
  39. Wright R., Basson M., D'Ari L., Rine J. Increased amounts of HMG-CoA reductase induce "karmellae": a proliferation of stacked membrane pairs surrounding the yeast nucleus. J Cell Biol. 1988 Jul;107(1):101–114. doi: 10.1083/jcb.107.1.101. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Yang Z., Park H., Lacy G. H., Cramer C. L. Differential activation of potato 3-hydroxy-3-methylglutaryl coenzyme A reductase genes by wounding and pathogen challenge. Plant Cell. 1991 Apr;3(4):397–405. doi: 10.1105/tpc.3.4.397. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. von Heijne G. Mitochondrial targeting sequences may form amphiphilic helices. EMBO J. 1986 Jun;5(6):1335–1342. doi: 10.1002/j.1460-2075.1986.tb04364.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. von Heijne G., Steppuhn J., Herrmann R. G. Domain structure of mitochondrial and chloroplast targeting peptides. Eur J Biochem. 1989 Apr 1;180(3):535–545. doi: 10.1111/j.1432-1033.1989.tb14679.x. [DOI] [PubMed] [Google Scholar]

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