Cap-Independent Translational Enhancement of Turnip Crinkle Virus Genomic and Subgenomic RNAs

ABSTRACT The presence of translational control elements and cap structures has not been carefully investigated for members of theCarmovirus genus, a group of small icosahedral plant viruses with positive-sense RNA genomes. In this study, we examined both the 5′ and 3′ untranslated regions (UTRs) of the turnip crinkle carmovirus (TCV) genomic RNA (4 kb) as well as the 5′ UTR of the coat protein subgenomic RNA (1.45 kb) for their roles in translational regulation. All three UTRs enhanced translation of the firefly luciferase reporter gene to different extents. Optimal translational efficiency was achieved when mRNAs contained both 5′ and 3′ UTRs. The synergistic effect due to the 5′-3′ cooperation was at least fourfold greater than the sum of the contributions of the individual UTRs. The observed translational enhancement of TCV mRNAs occurred in a cap-independent manner, a result consistent with the demonstration, using a cap-specific antibody, that the 5′ end of the TCV genomic RNA was uncapped. Finally, the translational enhancement activity within the 5′ UTR of 1.45-kb subgenomic RNA was shown to be important for the translation of coat protein in protoplasts and for virulent infection in Arabidopsis plants.

[1]  R. Vale,et al.  Circularization of mRNA by eukaryotic translation initiation factors. , 1998, Molecular cell.

[2]  W. Miller,et al.  A potential mechanism for selective control of cap-independent translation by a viral RNA sequence in cis and in trans. , 1999, RNA.

[3]  A. Simon,et al.  Analysis of the two subgenomic RNA promoters for turnip crinkle virus in vivo and in vitro. , 1997, Virology.

[4]  J. van Emmelo,et al.  Subgenomic RNAs mediate expression of cistrons located internally on the genomic RNA of tobacco necrosis virus strain A , 1992, Journal of virology.

[5]  Z. Weng,et al.  Genome organization and gene expression of saguaro cactus carmovirus. , 1997, The Journal of general virology.

[6]  J. Carrington,et al.  Turnip crinkle virus infection from RNA synthesized in vitro. , 1989, Virology.

[7]  J. Carrington,et al.  Nucleotide sequence and genome organization of carnation mottle virus RNA. , 1985, Nucleic acids research.

[8]  S. Harrison,et al.  Structure and assembly of turnip crinkle virus. I. X-ray crystallographic structure analysis at 3.2 A resolution. , 1986, Journal of molecular biology.

[9]  J. Skuzeski,et al.  Immunodetection, expression strategy and complementation of turnip crinkle virus p28 and p88 replication components. , 1995, Virology.

[10]  D. Gallie The cap and poly(A) tail function synergistically to regulate mRNA translational efficiency. , 1991, Genes & development.

[11]  R. Lührmann,et al.  A monoclonal antibody against 2,2,7-trimethylguanosine that reacts with intact, class U, small nuclear ribonucleoproteins as well as with 7-methylguanosine-capped RNAs. , 1987, European journal of biochemistry.

[12]  D. Gallie A tale of two termini: a functional interaction between the termini of an mRNA is a prerequisite for efficient translation initiation. , 1998, Gene.

[13]  W. Miller,et al.  A Sequence Located 4.5 to 5 Kilobases from the 5′ End of the Barley Yellow Dwarf Virus (PAV) Genome Strongly Stimulates Translation of Uncapped mRNA (*) , 1995, The Journal of Biological Chemistry.

[14]  M. Lai,et al.  An internal polypyrimidine-tract-binding protein-binding site in the hepatitis C virus RNA attenuates translation, which is relieved by the 3'-untranslated sequence. , 1999, Virology.

[15]  P. Ahlquist,et al.  A Brome Mosaic Virus Intergenic RNA3 Replication Signal Functions with Viral Replication Protein 1a To Dramatically Stabilize RNA In Vivo , 1999, Journal of Virology.

[16]  The genomic sequence of cardamine chlorotic fleck carmovirus. , 1993, The Journal of general virology.

[17]  J. Sambrook,et al.  Molecular Cloning: A Laboratory Manual , 2001 .

[18]  W. Miller,et al.  Barley yellow dwarf virus RNA requires a cap-independent translation sequence because it lacks a 5' cap. , 1999, Virology.

[19]  Lee Gehrke,et al.  Enhanced translation of chimaeric messenger RNAs containing a plant viral untranslated leader sequence , 1987, Nature.

[20]  J. Carrington,et al.  Cap-independent enhancement of translation by a plant potyvirus 5' nontranslated region , 1990, Journal of virology.

[21]  W. Miller,et al.  A viral sequence in the 3′‐untranslated region mimics a 5′ cap in facilitating translation of uncapped mRNA , 1997, The EMBO journal.

[22]  K. Kim,et al.  The 5' nontranslated region of potato virus X RNA affects both genomic and subgenomic RNA synthesis , 1996, Journal of virology.

[23]  P. Turner,et al.  Mutational analysis of the tobacco mosaic virus 5'-leader for altered ability to enhance translation. , 1988, Nucleic acids research.

[24]  S. Schlesinger,et al.  Translation of Sindbis virus mRNA: analysis of sequences downstream of the initiating AUG codon that enhance translation , 1996, Journal of virology.

[25]  Robert L. Tanguay,et al.  Translational efficiency is regulated by the length of the 3' untranslated region , 1996, Molecular and cellular biology.

[26]  W. J. Lucas,et al.  Visualizing mRNA expression in plant protoplasts: factors influencing efficient mRNA uptake and translation. , 1989, The Plant cell.

[27]  N. Sonenberg,et al.  Interaction of polyadenylate-binding protein with the eIF4G homologue PAIP enhances translation , 1998, Nature.

[28]  C. Song,et al.  Requirement of a 3'-terminal stem-loop in in vitro transcription by an RNA-dependent RNA polymerase. , 1995, Journal of molecular biology.

[29]  K. White,et al.  Uncoupled Expression of p33 and p92 Permits Amplification of Tomato Bushy Stunt Virus RNAs , 1998, Journal of Virology.

[30]  The 3' untranslated region of satellite tobacco necrosis virus RNA stimulates translation in vitro. , 1993, Molecular and cellular biology.

[31]  J W Watts,et al.  The 5'-leader sequence of tobacco mosaic virus RNA enhances the expression of foreign gene transcripts in vitro and in vivo. , 1987, Nucleic acids research.

[32]  V. Walbot,et al.  Identification of the motifs within the tobacco mosaic virus 5'-leader responsible for enhancing translation. , 1992, Nucleic acids research.

[33]  S. Harrison,et al.  Structure and assembly of turnip crinkle virus. IV. Analysis of the coat protein gene and implications of the subunit primary structure. , 1987, Journal of molecular biology.

[34]  F. Qu,et al.  Cell-to-cell movement of turnip crinkle virus is controlled by two small open reading frames that function in trans. , 1998, Virology.

[35]  S. Schlesinger,et al.  Translation of Sindbis virus mRNA: effects of sequences downstream of the initiating codon , 1994, Journal of virology.

[36]  F. Qu,et al.  Encapsidation of turnip crinkle virus is defined by a specific packaging signal and RNA size , 1997, Journal of virology.

[37]  Robert L. Tanguay,et al.  The tobacco etch viral 5' leader and poly(A) tail are functionally synergistic regulators of translation. , 1995, Gene.

[38]  B. Kastner,et al.  Isolation of S. cerevisiae snRNPs: comparison of U1 and U4/U6.U5 to their human counterparts. , 1994, Science.

[39]  M. Van Montagu,et al.  5'- and 3'-sequences of satellite tobacco necrosis virus RNA promoting translation in tobacco. , 1998, The Plant journal : for cell and molecular biology.