rRNA-complementarity in the 5' untranslated region of mRNA specifying the Gtx homeodomain protein: evidence that base- pairing to 18S rRNA affects translational efficiency.

Numerous eukaryotic mRNAs contain sequences complementary to segments of the 18S and 28S rRNAs. Previous studies raised the possibility that these complementarities might allow mRNA-rRNA interactions and affect rates of translation. In the present study, we investigated the mRNA encoding the mouse Gtx homeodomain protein. This mRNA contains within its 5' untranslated region (UTR) a segment that is complementary to two regions of the 18S rRNA, located at nucleotides 701-741 and 1104-1136. A Gtx RNA probe containing this complementarity could be photochemically cross-linked to ribosomal subunits through a linkage to 18S rRNA but not to 28S rRNA. Oligonucleotide-directed RNase H digestion of the rRNA and a reverse transcription analysis localized the cross-linked probe to the complementary segment of 18S rRNA at nucleotides 1104-1136 but not at nucleotides 701-741. To determine whether complementarity in the Gtx mRNA affected translation, a mutational analysis was performed with a Gtx-luciferase fusion construct and four related constructs with altered complementarity to the 18S rRNA. These constructs were examined for their ability to be translated in cell-free lysates prepared from P19 embryonal carcinoma and C6 glioma cell lines and after cellular transfection into these same cell lines. In both cell-free translation and transfection studies, the rate of translation decreased more than 9-fold as the degree of complementarity to nucleotides 1104-1136 of the 18S rRNA increased. We hypothesize that segments complementary to rRNA, such as those contained within the Gtx mRNA, form a category of cis-acting regulatory elements in mRNAs that affect translation by base pairing to rRNA within ribosomes.

[1]  G. Edelman,et al.  rRNA complementarity within mRNAs: a possible basis for mRNA-ribosome interactions and translational control. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[2]  D A Day,et al.  Post-transcriptional gene regulatory mechanisms in eukaryotes: an overview. , 1998, The Journal of endocrinology.

[3]  S. Scherer,et al.  Evidence That the Homeodomain Protein Gtx Is Involved in the Regulation of Oligodendrocyte Myelination , 1997, The Journal of Neuroscience.

[4]  M. Clemens,et al.  PKR--a protein kinase regulated by double-stranded RNA. , 1997, The international journal of biochemistry & cell biology.

[5]  J. Bachellerie,et al.  Guiding ribose methylation of rRNA. , 1997, Trends in biochemical sciences.

[6]  A G Porter,et al.  Functional importance of RNA interactions in selection of translation initiation codons , 1997, Molecular microbiology.

[7]  G. Dunny,et al.  Pheromone‐inducible expression of an aggregation protein in Enterococcus faecalis requires interaction of a plasmid‐encoded RNA with components of the ribosome , 1997, Molecular microbiology.

[8]  M. Zenkova,et al.  Studying functional significance of the sequence 980-1061 in the central domain of human 18S rRNA using complementary DNA probes. , 1997, Biochimica et biophysica acta.

[9]  G. Edelman,et al.  rRNA-like sequences occur in diverse primary transcripts: implications for the control of gene expression. , 1997, Proceedings of the National Academy of Sciences of the United States of America.

[10]  M. Yao,et al.  Antisense ribosomes: rRNA as a vehicle for antisense RNAs. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[11]  M. L. Sprengart,et al.  The downstream box: an efficient and independent translation initiation signal in Escherichia coli. , 1996, The EMBO journal.

[12]  T. Yoshida,et al.  Cloning of the rat Gadd45 cDNA and its mRNA expression in the brain. , 1994, Gene.

[13]  G. Scheper,et al.  Basepairing with 18S ribosomal RNA in internal initiation of translation , 1994, FEBS letters.

[14]  P. Wollenzien,et al.  mRNA binding track in the human 80S ribosome for mRNA analogues randomly substituted with 4-thiouridine residues. , 1994, Biochemistry.

[15]  O. Nygård,et al.  Probing the structure of mouse Ehrlich ascites cell 5.8S, 18S and 28S ribosomal RNA in situ. , 1994, Nucleic acids research.

[16]  Helen M. Blau,et al.  Tumor suppression by RNA from the 3′ untranslated region of α-tropomyosin , 1993, Cell.

[17]  R. Bodmer,et al.  Gtx: a novel murine homeobox‐containing gene, expressed specifically in glial cells of the brain and germ cells of testis, has a transcriptional repressor activity in vitro for a serum‐inducible promoter. , 1993, The EMBO journal.

[18]  O. Matveeva,et al.  Intermolecular mRNA-rRNA hybridization and the distribution of potential interaction regions in murine 18S rRNA. , 1993, Nucleic acids research.

[19]  M. Mathews,et al.  Interactions between double-stranded RNA regulators and the protein kinase DAI , 1992, Molecular and cellular biology.

[20]  P. Wollenzien,et al.  The mRNA binding track in the Escherichia coli ribosome for mRNAs of different sequences. , 1992, Biochemistry.

[21]  S. Dooley,et al.  Cross-hybridization between the avian myeloblastosis oncogene and eukaryotic 28S ribosomal RNA. , 1992, Gene.

[22]  A. Favre,et al.  Conformation and structural fluctuations of a 218 nucleotides long rRNA fragment: 4-thiouridine as an intrinsic photolabelling probe , 1991, Nucleic Acids Res..

[23]  D. Lipman,et al.  Improved tools for biological sequence comparison. , 1988, Proceedings of the National Academy of Sciences of the United States of America.

[24]  W. Hill,et al.  Probing ribosome structure using short oligodeoxyribonucleotides: the question of resolution. , 1987, Biochimie.

[25]  H. D. de Boer,et al.  Specialized ribosome system: preferential translation of a single mRNA species by a subpopulation of mutated ribosomes in Escherichia coli. , 1987, Proceedings of the National Academy of Sciences of the United States of America.

[26]  M. Santer,et al.  A single base change in the Shine-Dalgarno region of 16S rRNA of Escherichia coli affects translation of many proteins. , 1987, Proceedings of the National Academy of Sciences of the United States of America.

[27]  A. Shatkin,et al.  Proximity of mRNA5′-region and 18S rRNA in eukaryotic initiation complexes , 1980, Nature.

[28]  J. Shine,et al.  The 3'-terminal sequence of Escherichia coli 16S ribosomal RNA: complementarity to nonsense triplets and ribosome binding sites. , 1974, Proceedings of the National Academy of Sciences of the United States of America.

[29]  I. Wool,et al.  Determination of the number of proteins in liver ribosomes and ribosomal subunits by two-dimensional polyacrylamide gel electrophoresis. , 1972, The Journal of biological chemistry.

[30]  I. Wool,et al.  Dissociation and reassociation of skeletal muscle ribosomes. , 1969, Journal of molecular biology.

[31]  M. Mathews,et al.  The regulation of the protein kinase PKR by RNA. , 1996, Biochimie.

[32]  S. Peltz,et al.  Interrelationships of the pathways of mRNA decay and translation in eukaryotic cells. , 1996, Annual review of biochemistry.

[33]  E. Davies,et al.  Methods for isolation and analysis of polyribosomes. , 1995, Methods in cell biology.

[34]  O. Uhlenbeck,et al.  Synthesis of small RNAs using T7 RNA polymerase. , 1989, Methods in enzymology.

[35]  P. Wollenzien.  [21] Isolation and identification of RNA cross-links , 1988 .

[36]  D. Camp,et al.  Probing ribosome structure and function using short oligodeoxyribonucleotides. , 1988, Methods in enzymology.

[37]  W. Musters,et al.  Evolutionary conservation of structure and function of high molecular weight ribosomal RNA. , 1988, Progress in biophysics and molecular biology.