The translation in vitro of rat ornithine decarboxylase mRNA is blocked by its 5' untranslated region in a polyamine-independent way.
The enzyme ornithine decarboxylase (ODC, EC 4.1.1.17) is believed to play an essential role in the growth and differentiation of cells by regulating the biosynthesis of polyamines. The 5' untranslated region (5' UTR) of the ODC mRNA of different species is rather unusual in length and GC content, and may therefore be involved in translational control of ODC protein synthesis. We cloned the rat ODC cDNA downstream of the phage T7 promoter in order to perform transcription/translation studies in vitro. Our results show that the intact 5' UTR of rat ODC mRNA, which is 303 nt in length, is a potent inhibitor of translation. Efficient synthesis in vitro of ODC protein is obtained when either 172 nt from the 5'-end or 236 nt from the 3'-end of the 5' UTR are removed. A truncated 5' UTR with a calculated free energy of less than -272 kJ (-65 kcal/mol) is unable to support the synthesis in vitro of ODC protein. The short open reading frame (ORF) present in the 5' UTR of rat ODC mRNA does not contribute to the observed inhibitory effect on translation efficiency in vitro. At low polyamine concentration the efficiency of translation in vitro of intact ODC mRNA is not relatively increased compared with that of an ODC mRNA having a truncated 5' UTR or with that of control globin mRNA. From this we conclude that the well-documented negative feedback control of intracellular polyamines on ODC expression is not regulated by effects of polyamines on the secondary structure of the 5' UTR of ODC mRNA.
Van Steeg
H
H
National Institute of Public Health and Environmental Protection, Bilthoven, The Netherlands.
Van Oostrom
C T
CT
Hodemaekers
H M
HM
Peters
L
L
Thomas
A A
AA
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002151:Calorimetry; D003001:Cloning, Molecular; D005246:Feedback; D008969:Molecular Sequence Data; D015345:Oligonucleotide Probes; D016366:Open Reading Frames; D009955:Ornithine Decarboxylase / Q000235:genetics*; D010957:Plasmids; D011073:Polyamines / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000235:genetics*; D051381:Rats; D015183:Restriction Mapping; D013604:T-Phages / Q000235:genetics; D014158:Transcription, Genetic
D015345:Oligonucleotide Probes; D011073:Polyamines; D012333:RNA, Messenger; D009955:Ornithine Decarboxylase