• We are sorry, but NCBI web applications do not support your browser and may not function properly. More information
Logo of molcellbPermissionsJournals.ASM.orgJournalMCB ArticleJournal InfoAuthorsReviewers
Mol Cell Biol. Nov 1994; 14(11): 7322–7330.
PMCID: PMC359267

Cap-dependent and cap-independent translation by internal initiation of mRNAs in cell extracts prepared from Saccharomyces cerevisiae.


Translation extracts were prepared from various strains of Saccharomyces cerevisiae. The translation of mRNA molecules in these extracts were cooperatively enhanced by the presence of 5'-terminal cap structures and 3'-terminal poly(A) sequences. These cooperative effects could not be observed in other translation systems such as those prepared from rabbit reticulocytes, wheat germ, and human HeLa cells. Because the yeast translation system mimicked the effects of the cap structure and poly(A) tail on translational efficiency seen in vivo, this system was used to study cap-dependent and cap-independent translation of viral and cellular mRNA molecules. Both the 5' noncoding regions of hepatitis C virus and those of coxsackievirus B1 conferred cap-independent translation to a reporter coding region during translation in the yeast extracts; thus, the yeast translational apparatus is capable of initiating cap-independent translation. Although the translation of most yeast mRNAs was cap dependent, the unusually long 5' noncoding regions of mRNAs encoding cellular transcription factors TFIID and HAP4 were shown to mediate cap-independent translation in these extracts. Furthermore, both TFIID and HAP4 5' noncoding regions mediated translation of a second cistron when placed into the intercistronic spacer region of a dicistronic mRNA, indicating that these leader sequences can initiate translation by an internal ribosome binding mechanism in this in vitro translation system. This finding raises the possibility that an internal translation initiation mechanism exists in yeast cells for regulated translation of endogenous mRNAs.

Full text

Full text is available as a scanned copy of the original print version. Get a printable copy (PDF file) of the complete article (2.1M), or click on a page image below to browse page by page. Links to PubMed are also available for Selected References.

Images in this article

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Altmann M, Blum S, Pelletier J, Sonenberg N, Wilson TM, Trachsel H. Translation initiation factor-dependent extracts from Saccharomyces cerevisiae. Biochim Biophys Acta. 1990 Aug 27;1050(1-3):155–159. [PubMed]
  • Altmann M, Sonenberg N, Trachsel H. Translation in Saccharomyces cerevisiae: initiation factor 4E-dependent cell-free system. Mol Cell Biol. 1989 Oct;9(10):4467–4472. [PMC free article] [PubMed]
  • Anderson JT, Paddy MR, Swanson MS. PUB1 is a major nuclear and cytoplasmic polyadenylated RNA-binding protein in Saccharomyces cerevisiae. Mol Cell Biol. 1993 Oct;13(10):6102–6113. [PMC free article] [PubMed]
  • Bandyopadhyay PK, Wang C, Lipton HL. Cap-independent translation by the 5' untranslated region of Theiler's murine encephalomyelitis virus. J Virol. 1992 Nov;66(11):6249–6256. [PMC free article] [PubMed]
  • Banerjee AK. 5'-terminal cap structure in eucaryotic messenger ribonucleic acids. Microbiol Rev. 1980 Jun;44(2):175–205. [PMC free article] [PubMed]
  • Blanc A, Goyer C, Sonenberg N. The coat protein of the yeast double-stranded RNA virus L-A attaches covalently to the cap structure of eukaryotic mRNA. Mol Cell Biol. 1992 Aug;12(8):3390–3398. [PMC free article] [PubMed]
  • Borman A, Jackson RJ. Initiation of translation of human rhinovirus RNA: mapping the internal ribosome entry site. Virology. 1992 Jun;188(2):685–696. [PubMed]
  • Brawerman G. The Role of the poly(A) sequence in mammalian messenger RNA. CRC Crit Rev Biochem. 1981;10(1):1–38. [PubMed]
  • Brown EA, Day SP, Jansen RW, Lemon SM. The 5' nontranslated region of hepatitis A virus RNA: secondary structure and elements required for translation in vitro. J Virol. 1991 Nov;65(11):5828–5838. [PMC free article] [PubMed]
  • Cigan AM, Donahue TF. Sequence and structural features associated with translational initiator regions in yeast--a review. Gene. 1987;59(1):1–18. [PubMed]
  • Coward P, Dasgupta A. Yeast cells are incapable of translating RNAs containing the poliovirus 5' untranslated region: evidence for a translational inhibitor. J Virol. 1992 Jan;66(1):286–295. [PMC free article] [PubMed]
  • Dever TE, Feng L, Wek RC, Cigan AM, Donahue TF, Hinnebusch AG. Phosphorylation of initiation factor 2 alpha by protein kinase GCN2 mediates gene-specific translational control of GCN4 in yeast. Cell. 1992 Feb 7;68(3):585–596. [PubMed]
  • Everett JG, Gallie DR. RNA delivery in Saccharomyces cerevisiae using electroporation. Yeast. 1992 Dec;8(12):1007–1014. [PubMed]
  • Forsburg SL, Guarente L. Identification and characterization of HAP4: a third component of the CCAAT-bound HAP2/HAP3 heteromer. Genes Dev. 1989 Aug;3(8):1166–1178. [PubMed]
  • Fujimura T, Wickner RB. Replicase of L-A virus-like particles of Saccharomyces cerevisiae. In vitro conversion of exogenous L-A and M1 single-stranded RNAs to double-stranded form. J Biol Chem. 1988 Jan 5;263(1):454–460. [PubMed]
  • Gallie DR. The cap and poly(A) tail function synergistically to regulate mRNA translational efficiency. Genes Dev. 1991 Nov;5(11):2108–2116. [PubMed]
  • Gasior E, Herrera F, Sadnik I, McLaughlin CS, Moldave K. The preparation and characterization of a cell-free system from Saccharomyces cerevisiae that translates natural messenger ribonucleic acid. J Biol Chem. 1979 May 25;254(10):3965–3969. [PubMed]
  • Gerstel B, Tuite MF, McCarthy JE. The effects of 5'-capping, 3'-polyadenylation and leader composition upon the translation and stability of mRNA in a cell-free extract derived from the yeast Saccharomyces cerevisiae. Mol Microbiol. 1992 Aug;6(16):2339–2348. [PubMed]
  • Hellen CU, Witherell GW, Schmid M, Shin SH, Pestova TV, Gil A, Wimmer E. A cytoplasmic 57-kDa protein that is required for translation of picornavirus RNA by internal ribosomal entry is identical to the nuclear pyrimidine tract-binding protein. Proc Natl Acad Sci U S A. 1993 Aug 15;90(16):7642–7646. [PMC free article] [PubMed]
  • Hinnebusch AG. Transcriptional and translational regulation of gene expression in the general control of amino-acid biosynthesis in Saccharomyces cerevisiae. Prog Nucleic Acid Res Mol Biol. 1990;38:195–240. [PubMed]
  • Hofbauer R, Fessl F, Hamilton B, Ruis H. Preparation of a mRNA-dependent cell-free translation system from whole cells of Saccharomyces cerevisiae. Eur J Biochem. 1982 Feb;122(1):199–203. [PubMed]
  • Hsu CL, Stevens A. Yeast cells lacking 5'-->3' exoribonuclease 1 contain mRNA species that are poly(A) deficient and partially lack the 5' cap structure. Mol Cell Biol. 1993 Aug;13(8):4826–4835. [PMC free article] [PubMed]
  • Hussain I, Leibowitz MJ. Translation of homologous and heterologous messenger RNAs in a yeast cell-free system. Gene. 1986;46(1):13–23. [PubMed]
  • Iizuka N, Yonekawa H, Nomoto A. Nucleotide sequences important for translation initiation of enterovirus RNA. J Virol. 1991 Sep;65(9):4867–4873. [PMC free article] [PubMed]
  • Jackson RJ, Howell MT, Kaminski A. The novel mechanism of initiation of picornavirus RNA translation. Trends Biochem Sci. 1990 Dec;15(12):477–483. [PubMed]
  • Jang SK, Kräusslich HG, Nicklin MJ, Duke GM, Palmenberg AC, Wimmer E. A segment of the 5' nontranslated region of encephalomyocarditis virus RNA directs internal entry of ribosomes during in vitro translation. J Virol. 1988 Aug;62(8):2636–2643. [PMC free article] [PubMed]
  • Johnston M, Davis RW. Sequences that regulate the divergent GAL1-GAL10 promoter in Saccharomyces cerevisiae. Mol Cell Biol. 1984 Aug;4(8):1440–1448. [PMC free article] [PubMed]
  • Kozak M. The scanning model for translation: an update. J Cell Biol. 1989 Feb;108(2):229–241. [PMC free article] [PubMed]
  • Kühn R, Luz N, Beck E. Functional analysis of the internal translation initiation site of foot-and-mouth disease virus. J Virol. 1990 Oct;64(10):4625–4631. [PMC free article] [PubMed]
  • Leibowitz MJ, Barbone FP, Georgopoulos DE. In vitro protein synthesis. Methods Enzymol. 1991;194:536–545. [PubMed]
  • Macejak DG, Sarnow P. Internal initiation of translation mediated by the 5' leader of a cellular mRNA. Nature. 1991 Sep 5;353(6339):90–94. [PubMed]
  • Matunis MJ, Matunis EL, Dreyfuss G. PUB1: a major yeast poly(A)+ RNA-binding protein. Mol Cell Biol. 1993 Oct;13(10):6114–6123. [PMC free article] [PubMed]
  • McBratney S, Chen CY, Sarnow P. Internal initiation of translation. Curr Opin Cell Biol. 1993 Dec;5(6):961–965. [PubMed]
  • Meerovitch K, Svitkin YV, Lee HS, Lejbkowicz F, Kenan DJ, Chan EK, Agol VI, Keene JD, Sonenberg N. La autoantigen enhances and corrects aberrant translation of poliovirus RNA in reticulocyte lysate. J Virol. 1993 Jul;67(7):3798–3807. [PMC free article] [PubMed]
  • Merrick WC. Mechanism and regulation of eukaryotic protein synthesis. Microbiol Rev. 1992 Jun;56(2):291–315. [PMC free article] [PubMed]
  • Moye-Rowley WS, Harshman KD, Parker CS. Yeast YAP1 encodes a novel form of the jun family of transcriptional activator proteins. Genes Dev. 1989 Mar;3(3):283–292. [PubMed]
  • Oh SK, Scott MP, Sarnow P. Homeotic gene Antennapedia mRNA contains 5'-noncoding sequences that confer translational initiation by internal ribosome binding. Genes Dev. 1992 Sep;6(9):1643–1653. [PubMed]
  • Ow DW, DE Wet JR, Helinski DR, Howell SH, Wood KV, Deluca M. Transient and stable expression of the firefly luciferase gene in plant cells and transgenic plants. Science. 1986 Nov 14;234(4778):856–859. [PubMed]
  • Pelletier J, Sonenberg N. Insertion mutagenesis to increase secondary structure within the 5' noncoding region of a eukaryotic mRNA reduces translational efficiency. Cell. 1985 Mar;40(3):515–526. [PubMed]
  • Russell PJ, Hambidge SJ, Kirkegaard K. Direct introduction and transient expression of capped and non-capped RNA in Saccharomyces cerevisiae. Nucleic Acids Res. 1991 Sep 25;19(18):4949–4953. [PMC free article] [PubMed]
  • Sachs AB, Davis RW. The poly(A) binding protein is required for poly(A) shortening and 60S ribosomal subunit-dependent translation initiation. Cell. 1989 Sep 8;58(5):857–867. [PubMed]
  • Sachs AB, Deardorff JA. Translation initiation requires the PAB-dependent poly(A) ribonuclease in yeast. Cell. 1992 Sep 18;70(6):961–973. [PubMed]
  • Schmidt MC, Kao CC, Pei R, Berk AJ. Yeast TATA-box transcription factor gene. Proc Natl Acad Sci U S A. 1989 Oct;86(20):7785–7789. [PMC free article] [PubMed]
  • Sikorski RS, Hieter P. A system of shuttle vectors and yeast host strains designed for efficient manipulation of DNA in Saccharomyces cerevisiae. Genetics. 1989 May;122(1):19–27. [PMC free article] [PubMed]
  • Sonenberg N. Cap-binding proteins of eukaryotic messenger RNA: functions in initiation and control of translation. Prog Nucleic Acid Res Mol Biol. 1988;35:173–207. [PubMed]
  • Tsukiyama-Kohara K, Iizuka N, Kohara M, Nomoto A. Internal ribosome entry site within hepatitis C virus RNA. J Virol. 1992 Mar;66(3):1476–1483. [PMC free article] [PubMed]
  • Wang C, Sarnow P, Siddiqui A. Translation of human hepatitis C virus RNA in cultured cells is mediated by an internal ribosome-binding mechanism. J Virol. 1993 Jun;67(6):3338–3344. [PMC free article] [PubMed]
  • Wickner RB. Double-stranded and single-stranded RNA viruses of Saccharomyces cerevisiae. Annu Rev Microbiol. 1992;46:347–375. [PubMed]

Articles from Molecular and Cellular Biology are provided here courtesy of American Society for Microbiology (ASM)


Related citations in PubMed

See reviews...See all...

Cited by other articles in PMC

See all...


Recent Activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...