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. 1998 Jan;4(1):65–73.

Functionality of mutations at conserved nucleotides in eukaryotic SECIS elements is determined by the identity of a single nonconserved nucleotide.

G W Martin 3rd 1, J W Harney 1, M J Berry 1
PMCID: PMC1369597  PMID: 9436909

Abstract

In eukaryotes, the specific cotranslational insertion of selenocysteine at UGA codons requires the presence of a secondary structural motif in the 3' untranslated region of the selenoprotein mRNA. This selenocysteine insertion sequence (SECIS) element is predicted to form a hairpin and contains three regions of sequence invariance that are thought to interact with a specific protein or proteins. Specificity of RNA-binding protein recognition of cognate RNAs is usually characterized by the ability of the protein to recognize and distinguish between a consensus binding site and sequences containing mutations to highly conserved positions in the consensus sequence. Using a functional assay for the ability of wild-type and mutant SECIS elements to direct cotranslational selenocysteine incorporation, we have investigated the relative contributions of individual invariant nucleotides to SECIS element function. We report the novel finding that, for this consensus RNA motif, mutations at the invariant nucleotides are tolerated to different degrees in different elements, depending on the identity of a single nonconserved nucleotide. Further, we demonstrate that the sequences adjacent to the minimal element, although not required for function, can affect function through their propensity to base pair. These findings shed light on the specific structure these conserved sequences may form within the element. This information is crucial to the design of strategies for the identification of SECIS-binding proteins, and hence the elucidation of the mechanism of selenocysteine incorporation in eukaryotes.

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Selected References

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

  1. Battiste J. L., Mao H., Rao N. S., Tan R., Muhandiram D. R., Kay L. E., Frankel A. D., Williamson J. R. Alpha helix-RNA major groove recognition in an HIV-1 rev peptide-RRE RNA complex. Science. 1996 Sep 13;273(5281):1547–1551. doi: 10.1126/science.273.5281.1547. [DOI] [PubMed] [Google Scholar]
  2. Berry M. J., Banu L., Chen Y. Y., Mandel S. J., Kieffer J. D., Harney J. W., Larsen P. R. Recognition of UGA as a selenocysteine codon in type I deiodinase requires sequences in the 3' untranslated region. Nature. 1991 Sep 19;353(6341):273–276. doi: 10.1038/353273a0. [DOI] [PubMed] [Google Scholar]
  3. Berry M. J., Banu L., Harney J. W., Larsen P. R. Functional characterization of the eukaryotic SECIS elements which direct selenocysteine insertion at UGA codons. EMBO J. 1993 Aug;12(8):3315–3322. doi: 10.1002/j.1460-2075.1993.tb06001.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Berry M. J., Harney J. W., Ohama T., Hatfield D. L. Selenocysteine insertion or termination: factors affecting UGA codon fate and complementary anticodon:codon mutations. Nucleic Acids Res. 1994 Sep 11;22(18):3753–3759. doi: 10.1093/nar/22.18.3753. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Berry M. J., Kieffer J. D., Harney J. W., Larsen P. R. Selenocysteine confers the biochemical properties characteristic of the type I iodothyronine deiodinase. J Biol Chem. 1991 Aug 5;266(22):14155–14158. [PubMed] [Google Scholar]
  6. Calnan B. J., Biancalana S., Hudson D., Frankel A. D. Analysis of arginine-rich peptides from the HIV Tat protein reveals unusual features of RNA-protein recognition. Genes Dev. 1991 Feb;5(2):201–210. doi: 10.1101/gad.5.2.201. [DOI] [PubMed] [Google Scholar]
  7. Forchhammer K., Leinfelder W., Böck A. Identification of a novel translation factor necessary for the incorporation of selenocysteine into protein. Nature. 1989 Nov 23;342(6248):453–456. doi: 10.1038/342453a0. [DOI] [PubMed] [Google Scholar]
  8. Forchhammer K., Rücknagel K. P., Böck A. Purification and biochemical characterization of SELB, a translation factor involved in selenoprotein synthesis. J Biol Chem. 1990 Jun 5;265(16):9346–9350. [PubMed] [Google Scholar]
  9. Gossen M., Bujard H. Tight control of gene expression in mammalian cells by tetracycline-responsive promoters. Proc Natl Acad Sci U S A. 1992 Jun 15;89(12):5547–5551. doi: 10.1073/pnas.89.12.5547. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Heider J., Baron C., Böck A. Coding from a distance: dissection of the mRNA determinants required for the incorporation of selenocysteine into protein. EMBO J. 1992 Oct;11(10):3759–3766. doi: 10.1002/j.1460-2075.1992.tb05461.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Hill K. E., Lloyd R. S., Burk R. F. Conserved nucleotide sequences in the open reading frame and 3' untranslated region of selenoprotein P mRNA. Proc Natl Acad Sci U S A. 1993 Jan 15;90(2):537–541. doi: 10.1073/pnas.90.2.537. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Kollmus H., Flohé L., McCarthy J. E. Analysis of eukaryotic mRNA structures directing cotranslational incorporation of selenocysteine. Nucleic Acids Res. 1996 Apr 1;24(7):1195–1201. doi: 10.1093/nar/24.7.1195. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Leinfelder W., Forchhammer K., Zinoni F., Sawers G., Mandrand-Berthelot M. A., Böck A. Escherichia coli genes whose products are involved in selenium metabolism. J Bacteriol. 1988 Feb;170(2):540–546. doi: 10.1128/jb.170.2.540-546.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. 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]
  15. Martin G. W., 3rd, Harney J. W., Berry M. J. Selenocysteine incorporation in eukaryotes: insights into mechanism and efficiency from sequence, structure, and spacing proximity studies of the type 1 deiodinase SECIS element. RNA. 1996 Feb;2(2):171–182. [PMC free article] [PubMed] [Google Scholar]
  16. Ringquist S., Schneider D., Gibson T., Baron C., Böck A., Gold L. Recognition of the mRNA selenocysteine insertion sequence by the specialized translational elongation factor SELB. Genes Dev. 1994 Feb 1;8(3):376–385. doi: 10.1101/gad.8.3.376. [DOI] [PubMed] [Google Scholar]
  17. Rould M. A., Perona J. J., Söll D., Steitz T. A. Structure of E. coli glutaminyl-tRNA synthetase complexed with tRNA(Gln) and ATP at 2.8 A resolution. Science. 1989 Dec 1;246(4934):1135–1142. doi: 10.1126/science.2479982. [DOI] [PubMed] [Google Scholar]
  18. Shen Q., Chu F. F., Newburger P. E. Sequences in the 3'-untranslated region of the human cellular glutathione peroxidase gene are necessary and sufficient for selenocysteine incorporation at the UGA codon. J Biol Chem. 1993 May 25;268(15):11463–11469. [PubMed] [Google Scholar]
  19. Shen Q., Leonard J. L., Newburger P. E. Structure and function of the selenium translation element in the 3'-untranslated region of human cellular glutathione peroxidase mRNA. RNA. 1995 Jul;1(5):519–525. [PMC free article] [PubMed] [Google Scholar]
  20. Shen Q., McQuilkin P. A., Newburger P. E. RNA-binding proteins that specifically recognize the selenocysteine insertion sequence of human cellular glutathione peroxidase mRNA. J Biol Chem. 1995 Dec 22;270(51):30448–30452. doi: 10.1074/jbc.270.51.30448. [DOI] [PubMed] [Google Scholar]
  21. Walczak R., Westhof E., Carbon P., Krol A. A novel RNA structural motif in the selenocysteine insertion element of eukaryotic selenoprotein mRNAs. RNA. 1996 Apr;2(4):367–379. [PMC free article] [PubMed] [Google Scholar]
  22. Weeks K. M., Crothers D. M. RNA recognition by Tat-derived peptides: interaction in the major groove? Cell. 1991 Aug 9;66(3):577–588. doi: 10.1016/0092-8674(81)90020-9. [DOI] [PubMed] [Google Scholar]
  23. Williams D. L., Pierce R. J., Cookson E., Capron A. Molecular cloning and sequencing of glutathione peroxidase from Schistosoma mansoni. Mol Biochem Parasitol. 1992 May;52(1):127–130. doi: 10.1016/0166-6851(92)90042-i. [DOI] [PubMed] [Google Scholar]
  24. Zinoni F., Heider J., Böck A. Features of the formate dehydrogenase mRNA necessary for decoding of the UGA codon as selenocysteine. Proc Natl Acad Sci U S A. 1990 Jun;87(12):4660–4664. doi: 10.1073/pnas.87.12.4660. [DOI] [PMC free article] [PubMed] [Google Scholar]

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