Cloning and nucleotide sequence of rat ornithine decarboxylase cDNA

The enzyme ornithine decarboxylase (ODC; EC 4.1.1.17) catalyses the first and rate-limiting step in polyamine biosynthesis. Its activity is markedly increased in rapidly growing or regenerating tissue and is subject to regulation by a variety of trophic and mitogenic stimuli. ODC is therefore believed to play an essential role in the onset of cellular proliferation. In a molecular-biological approach to investigate ODC regulation upon induction by tumor promoters in rat liver we isolated an almost full-length rat ODC cDNA clone of 2.4 kb (designated pODC.E10) from a cDNA library of testosterone-induced rat kidney poly(A)+ RNA. Characterization by restriction-endonuclease mapping and sequence analysis showed strong homology to mouse ODC cDNA sequences previously published [Gupta and Coffino, J. Biol. Chem. 260 (1985) 2941-2944; Kahana and Nathans, Proc. Natl. Acad. Sci. USA 82 (1985) 1673-1677; Hickok et al., Proc. Natl. Acad. Sci. USA 83 (1986) 594-598]. This homology is most pronounced in the 461-aa-spanning coding region, amounting to 94% and 97% at the DNA and protein levels, respectively. In the 423-nt 5' leader the rat-mouse homology (approx. 75%) is most pronounced in a region of about 175 nt directly upstream from the translational start site. The leader sequence also contains a perfect inverted repeat of 54 nt and ten additional upstream ATG triplets, which are all followed by nonsense codons before the initiating ATG. In the 633-nt 3' trailer region of pODC.E10 an additional polyadenylation signal is observed more than 300 nt upstream from the 3' end. Rat-mouse homology is about 80% up to this first polyadenylation signal and is considerably less thereafter. The presence of two alternate polyadenylation sites most likely accounts for the 3' size heterogeneity observed in the two ODC mRNAs of 2.1 and 2.6 kb, respectively. In rat liver both mRNAs are coordinately induced by different tumor promoters. Finally, Southern blot analysis of normal rat liver and rat hepatoma DNA revealed that rat ODC, as in other rodents, belongs to a multigene family.     

Nucleotide sequence of hamster S-adenosylmethionine decarboxylase cDNA.

The nucleotide sequence of a cDNA encoding the proenzyme of hamster S-adenosylmethionine decarboxylase including 169 nucleotides of the 5' untranslated region has been determined. The deduced amino acid sequence shows a remarkable similarity to the human proenzyme with only seven differences out of 334 amino acids. The nucleotide sequence of the 5' untranslated region showed 93% homology with the corresponding rat and human sequences suggesting that this region may play an important role in the regulation of S-adenosylmethionine decarboxylase expression.     

Post-translational arginylation of ornithine decarboxylase from rat hepatocytes.

Ornithine decarboxylase (ODC) was purified 6500-fold from NMRI mouse kidneys under conditions designed to inhibit degradation by proteinases. The enzyme was homogeneous by SDS/polyacrylamide-gel electrophoresis, and the specific activity was among the highest reported. The yield was 70%. A monoclonal antibody against this preparation was generated and used in studies to investigate the half-life of ODC in cultured rat hepatocytes labelled with [35S]methionine. This value was 39 +/- 4 min and was unchanged when either NH4Cl (as a lysosomotropic agent) or leupeptin (as a lysosomal proteinase inhibitor) was added to the culture medium. Thus the intracellular turnover of ODC in cultured hepatocytes occurs mainly in extra-lysosomal compartments. Arginylation of rat ODC was investigated in vitro by incubation with L-[3H]arginyl-tRNA, and the incorporation of the label was compared with that of total cytosolic proteins. Arginylated ODC had a specific radioactivity 8600 times that of the bulk of cytosolic protein. Edman degradation of this ODC showed that the post-translational arginylation occurred only at the alpha-amino end of the enzyme. The inhibitor of arginyl-tRNA:protein arginyltransferase (EC 2.3.2.8), L-glutamyl-L-valyl-L-phenylalanine, increased the half-life of ODC in cultured hepatocytes from 39 min to more than 90 min. The possible significance of the preferential post-translational arginylation of ornithine decarboxylase to its rapid turnover is discussed.     

Allylisopropylacetamide induces rat hepatic ornithine decarboxylase

In rat liver, allylisopropylacetamide (AIA) treatment strongly induced (25-fold) the activity of rat hepatic ornithine decarboxylase (ODC). By either the oral or the subcutaneous route, AIA produced a long-lasting induction (30 to 40 hours) of hepatic ODC activity. Three analogs of AIA, propylisopropylacetamide (PIA), allobarbital, and allylbenzene, were active ODC inducers while a fourth, allylacetate, was not. Although induction of hepatic aminolevulinic acid (ALA) synthetase activity and the accumulation of hepatic porphyrins depend on the allyl moiety of AIA, this is not the case with hepatic ODC induction. Allylisopropylacetamide did not elevate serum alanine aminotransferase (SGPT) nor did it cause DNA damage, as measured by the alkaline elution assay. Thus, hepatic cell death is not a likely explanation of AIA's long-lasting induction of ODC. As AIA does not belong to any of the common categories of ODC inducers, it may be the chemical prototype of a new class of hepatic ODC inducers.     

Molecular cloning and sequence analysis of a chicken ornithine decarboxylase cDNA

A chicken embryo cDNA library was screened with a mouse probe for ornithine decarboxylase (ODC) and 14 positively hybridizing clones isolated. The longest of these (1.7 kb) was sub-cloned and sequenced. It is estimated that the clone comprises approximately 98% of the coding region for chicken ODC. The DNA sequence shows 78% identity with the human ODC cDNA sequence and the deduced amino acid sequence is almost 90% homologous to mouse and human. Both the peptide and cDNA sequences show interesting potential regulatory features which are discussed here.