Abstract
It is thought that Shine–Dalgarno-like sequences, which exhibit complementarity to the nucleotide sequences at the 3′-end of 18S rRNA, are not present in eukaryotic mRNAs. However, complementary sequences consisting of more than 5 nucleotides to the 3′-end of 18S rRNA, i.e., a CR sequence, are present at −17 to −32 upstream from the initiation codon AUG in 18 mRNAs involved in protein synthesis except eEF1A mRNA. Thus, effects of the CR sequence in mRNAs and polyamines on protein synthesis were examined using control and polyamine-reduced FM3A and NIH3T3 cells. Polyamines did not stimulate protein synthesis encoded by 18 mRNAs possessing a normal CR sequence. When the CR sequence was deleted, protein synthetic activities decreased to less than 70 % of intact mRNAs. In eEF1A mRNA, the CR sequence was located at −33 to −39 upstream from the initiation codon AUG, and polyamines stimulated eEF1A synthesis about threefold. When the CR sequence was shifted to −22 to −28 upstream from the AUG, eEF1A synthesis increased in polyamine-reduced cells and the degree of polyamine stimulation decreased greatly. The results indicate that the CR sequence exists in many eukaryotic mRNAs, and the location of a CR sequence in mRNAs influences polyamine stimulation of protein synthesis.
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Abbreviations
- APCHA:
-
N 1-(3-aminopropyl)-cyclohexylamine
- CD:
-
Circular dichroism
- CR sequence:
-
Complementary sequence to 18S rRNA
- DFMO:
-
α-Difluoromethylornithine
- GC7 :
-
N 1-guanyl-1,7-diaminoheptane
- PBS:
-
Phosphate-buffered saline
- 5′-UTR:
-
5′-Untranslated region of mRNA
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Acknowledgments
We thank Drs. A. J. Michael and K. Williams for their help in preparing the manuscript. We also thank Dr. R. G. Mirmira for his kind gift of antibody against hypusinated eIF5A. This work was supported by a Grant-in-Aid for Scientific Research from the Ministry of Education, Culture, Sports, Science and Technology, Japan.
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726_2014_1867_MOESM1_ESM.pptx
Fig. S1CR sequences in various mRNAs. The nucleotide sequences in the 5′-UTR of mRNAs were quoted from http://www.ncbi.nil.nih.gov/nucleotide/. The CR sequences in the 5′-UTR of various mRNAs are shown in red together with the numbers of the complementary nucleotide sequence of the 3′-end of 18S rRNA(Holmberg et al. 1994). Non-complementary base in CR sequences is shown in black. Consensus Kozak sequences (Kozak 1991) were shown in blue. Number of nucleotides of the 5′-UTR of specified mRNAs is shown in the parentheses (PPTX 68 kb)
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Terui, Y., Sakamoto, A., Yoshida, T. et al. Polyamine stimulation of eEF1A synthesis based on the unusual position of a complementary sequence to 18S rRNA in eEF1A mRNA. Amino Acids 47, 345–356 (2015). https://doi.org/10.1007/s00726-014-1867-z
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DOI: https://doi.org/10.1007/s00726-014-1867-z