Skip to main content
NCBI home page
RNA logoLink to RNA
. 2000 Sep;6(9):1306–1315. doi: 10.1017/s1355838200000388

Methylation of the ribosyl moiety at position 34 of selenocysteine tRNA[Ser]Sec is governed by both primary and tertiary structure.

L K Kim 1, T Matsufuji 1, S Matsufuji 1, B A Carlson 1, S S Kim 1, D L Hatfield 1, B J Lee 1
PMCID: PMC1370003  PMID: 10999607

Abstract

The selenocysteine (Sec) tRNA[Ser]Sec population in higher vertebrates consists of two major isoacceptors that differ from each other by a single nucleoside modification in the wobble position of the anticodon (position 34). One isoacceptor contains 5-methylcarboxymethyluridine (mcmU) in this position, whereas the other contains 5-methylcarboxymethyluridine-2'-O-methylribose (mcmUm). The other modifications in these tRNAs are N6-isopentenyladenosine (i6A), pseudouridine (psi), and 1-methyladenosine (m1A) at positions 37, 55, and 58, respectively. As methylation of the ribose at position 34 is influenced by the intracellular selenium status and the presence of this methyl group dramatically alters tertiary structure, we investigated the effect of the modifications at other positions as well as tertiary structure on its formation. Mutations were introduced within a synthetic gene encoded in an expression vector, transcripts generated and microinjected into Xenopus oocytes, and the resulting tRNA products analyzed for the presence of modified bases. The results suggest that efficient methylation of mcmU to yield mcmUm requires the prior formation of each modified base and an intact tertiary structure, whereas formation of modified bases at other positions, including mcmU, is not as stringently connected to precise primary and tertiary structure. These results, along with the observations that methylation of mcmU is enhanced in the presence of selenium and that this methyl group affects tertiary structure, further suggest that the mcmUm isoacceptor must have a role in selenoprotein synthesis different from that of the mcmU isoacceptor.

Full Text

The Full Text of this article is available as a PDF (808.0 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Amberg R., Mizutani T., Wu X. Q., Gross H. J. Selenocysteine synthesis in mammalia: an identity switch from tRNA(Ser) to tRNA(Sec). J Mol Biol. 1996 Oct 18;263(1):8–19. doi: 10.1006/jmbi.1996.0552. [DOI] [PubMed] [Google Scholar]
  2. Arnez J. G., Steitz T. A. Crystal structure of unmodified tRNA(Gln) complexed with glutaminyl-tRNA synthetase and ATP suggests a possible role for pseudo-uridines in stabilization of RNA structure. Biochemistry. 1994 Jun 21;33(24):7560–7567. doi: 10.1021/bi00190a008. [DOI] [PubMed] [Google Scholar]
  3. Berggren M., Gallegos A., Gasdaska J., Powis G. Cellular thioredoxin reductase activity is regulated by selenium. Anticancer Res. 1997 Sep-Oct;17(5A):3377–3380. [PubMed] [Google Scholar]
  4. Bermano G., Nicol F., Dyer J. A., Sunde R. A., Beckett G. J., Arthur J. R., Hesketh J. E. Tissue-specific regulation of selenoenzyme gene expression during selenium deficiency in rats. Biochem J. 1995 Oct 15;311(Pt 2):425–430. doi: 10.1042/bj3110425. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Chittum H. S., Hill K. E., Carlson B. A., Lee B. J., Burk R. F., Hatfield D. L. Replenishment of selenium deficient rats with selenium results in redistribution of the selenocysteine tRNA population in a tissue specific manner. Biochim Biophys Acta. 1997 Oct 30;1359(1):25–34. doi: 10.1016/s0167-4889(97)00092-x. [DOI] [PubMed] [Google Scholar]
  6. Choi I. S., Diamond A. M., Crain P. F., Kolker J. D., McCloskey J. A., Hatfield D. L. Reconstitution of the biosynthetic pathway of selenocysteine tRNAs in Xenopus oocytes. Biochemistry. 1994 Jan 18;33(2):601–605. doi: 10.1021/bi00168a027. [DOI] [PubMed] [Google Scholar]
  7. Diamond A. M., Choi I. S., Crain P. F., Hashizume T., Pomerantz S. C., Cruz R., Steer C. J., Hill K. E., Burk R. F., McCloskey J. A. Dietary selenium affects methylation of the wobble nucleoside in the anticodon of selenocysteine tRNA([Ser]Sec). J Biol Chem. 1993 Jul 5;268(19):14215–14223. [PubMed] [Google Scholar]
  8. Droogmans L., Haumont E., de Henau S., Grosjean H. Enzymatic 2'-O-methylation of the wobble nucleoside of eukaryotic tRNAPhe: specificity depends on structural elements outside the anticodon loop. EMBO J. 1986 May;5(5):1105–1109. doi: 10.1002/j.1460-2075.1986.tb04329.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Grosjean H., Edqvist J., Stråby K. B., Giegé R. Enzymatic formation of modified nucleosides in tRNA: dependence on tRNA architecture. J Mol Biol. 1996 Jan 12;255(1):67–85. doi: 10.1006/jmbi.1996.0007. [DOI] [PubMed] [Google Scholar]
  10. Grundner-Culemann E., Martin G. W., 3rd, Harney J. W., Berry M. J. Two distinct SECIS structures capable of directing selenocysteine incorporation in eukaryotes. RNA. 1999 May;5(5):625–635. doi: 10.1017/s1355838299981542. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Hatfield D., Lee B. J., Hampton L., Diamond A. M. Selenium induces changes in the selenocysteine tRNA[Ser]Sec population in mammalian cells. Nucleic Acids Res. 1991 Feb 25;19(4):939–943. doi: 10.1093/nar/19.4.939. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Higuchi R., Krummel B., Saiki R. K. A general method of in vitro preparation and specific mutagenesis of DNA fragments: study of protein and DNA interactions. Nucleic Acids Res. 1988 Aug 11;16(15):7351–7367. doi: 10.1093/nar/16.15.7351. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Hubert N., Sturchler C., Westhof E., Carbon P., Krol A. The 9/4 secondary structure of eukaryotic selenocysteine tRNA: more pieces of evidence. RNA. 1998 Sep;4(9):1029–1033. doi: 10.1017/s1355838298980888. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Ioudovitch A., Steinberg S. V. Structural compensation in an archaeal selenocysteine transfer RNA. J Mol Biol. 1999 Jul 9;290(2):365–371. doi: 10.1006/jmbi.1999.2901. [DOI] [PubMed] [Google Scholar]
  15. Jung J. E., Karoor V., Sandbaken M. G., Lee B. J., Ohama T., Gesteland R. F., Atkins J. F., Mullenbach G. T., Hill K. E., Wahba A. J. Utilization of selenocysteyl-tRNA[Ser]Sec and seryl-tRNA[Ser]Sec in protein synthesis. J Biol Chem. 1994 Nov 25;269(47):29739–29745. [PubMed] [Google Scholar]
  16. Kelmers A. D., Heatherly D. E. Columns for rapid chromatographic separation of small amounts of tracer-labeled transfer ribonucleic acids. Anal Biochem. 1971 Dec;44(2):486–495. doi: 10.1016/0003-2697(71)90236-3. [DOI] [PubMed] [Google Scholar]
  17. Kim S. H., Sussman J. L., Suddath F. L., Quigley G. J., McPherson A., Wang A. H., Seeman N. C., RICH A. The general structure of transfer RNA molecules. Proc Natl Acad Sci U S A. 1974 Dec;71(12):4970–4974. doi: 10.1073/pnas.71.12.4970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Lee B. J., de la Peña P., Tobian J. A., Zasloff M., Hatfield D. Unique pathway of expression of an opal suppressor phosphoserine tRNA. Proc Natl Acad Sci U S A. 1987 Sep;84(18):6384–6388. doi: 10.1073/pnas.84.18.6384. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Low S. C., Berry M. J. Knowing when not to stop: selenocysteine incorporation in eukaryotes. Trends Biochem Sci. 1996 Jun;21(6):203–208. [PubMed] [Google Scholar]
  20. Melton D. A., De Robertis E. M., Cortese R. Order and intracellular location of the events involved in the maturation of a spliced tRNA. Nature. 1980 Mar 13;284(5752):143–148. doi: 10.1038/284143a0. [DOI] [PubMed] [Google Scholar]
  21. Morin A., Auxilien S., Senger B., Tewari R., Grosjean H. Structural requirements for enzymatic formation of threonylcarbamoyladenosine (t6A) in tRNA: an in vivo study with Xenopus laevis oocytes. RNA. 1998 Jan;4(1):24–37. [PMC free article] [PubMed] [Google Scholar]
  22. Ohama T., Yang D. C., Hatfield D. L. Selenocysteine tRNA and serine tRNA are aminoacylated by the same synthetase, but may manifest different identities with respect to the long extra arm. Arch Biochem Biophys. 1994 Dec;315(2):293–301. doi: 10.1006/abbi.1994.1503. [DOI] [PubMed] [Google Scholar]
  23. Silberklang M., Gillum A. M., RajBhandary U. L. Use of in vitro 32P labeling in the sequence analysis of nonradioactive tRNAs. Methods Enzymol. 1979;59:58–109. doi: 10.1016/0076-6879(79)59072-7. [DOI] [PubMed] [Google Scholar]
  24. Sturchler C., Lescure A., Keith G., Carbon P., Krol A. Base modification pattern at the wobble position of Xenopus selenocysteine tRNA(Sec). Nucleic Acids Res. 1994 Apr 25;22(8):1354–1358. doi: 10.1093/nar/22.8.1354. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Tobian J. A., Drinkard L., Zasloff M. tRNA nuclear transport: defining the critical regions of human tRNAimet by point mutagenesis. Cell. 1985 Dec;43(2 Pt 1):415–422. doi: 10.1016/0092-8674(85)90171-0. [DOI] [PubMed] [Google Scholar]
  26. Wu X. Q., Gross H. J. The long extra arms of human tRNA((Ser)Sec) and tRNA(Ser) function as major identify elements for serylation in an orientation-dependent, but not sequence-specific manner. Nucleic Acids Res. 1993 Dec 11;21(24):5589–5594. doi: 10.1093/nar/21.24.5589. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Yeh J. Y., Vendeland S. C., Gu Q., Butler J. A., Ou B. R., Whanger P. D. Dietary selenium increases selenoprotein W levels in rat tissues. J Nutr. 1997 Nov;127(11):2165–2172. doi: 10.1093/jn/127.11.2165. [DOI] [PubMed] [Google Scholar]

Articles from RNA are provided here courtesy of The RNA Society

RESOURCES