J. Baileyserres et Rk. Dawe, BOTH 5'-SEQUENCES AND 3'-SEQUENCES OF MAIZE ADH1 MESSENGER-RNA ARE REQUIRED FOR ENHANCED TRANSLATION UNDER LOW-OXYGEN CONDITIONS, Plant physiology, 112(2), 1996, pp. 685-695
Alcohol dehydrogenase-1 (ADH1) synthesis in O-2-deprived roots of maiz
e (Zea mays L.) results from induced transcription and selective trans
lation of ADH1 mRNA. The effect of ADH1 mRNA sequences on message stab
ility and translation was studied in protoplasts of the maize cell lin
e P3377. 5' capped and 3' polyadenylated mRNA constructs containing th
e firefly gene (luc) for luciferase (LUG) or the Escherichia coli gene
(uidA) for beta-glucuronidase (GUS) coding region were synthesized in
vitro and electroporated into protoplasts that were cultured at 40 or
5% O-2. A LUC mRNA with a 17-nucleotide polylinker 5' untranslated re
gion (UTR) was expressed 10-fold higher under aerobic conditions than
under hypoxic conditions. Expression of five chimeric ADH1-GUS mRNAs w
as measured relative to this LUC mRNA. An mRNA containing the 5'-UTR a
nd the first 18 codons of adh 1 in a translational fusion with the GUS
coding region and followed by the 3'-UTR of adh1 was expressed 57-fol
d higher at 5% O-2. Progressive deletion of adh1 5'-UTR and coding seq
uences reduced expression of the GUS-mRNA at 5% O-2, but had little im
pact on expression of 40% O-2. Enhancement of expression in hypoxic pr
otoplasts conferred by the adh1 5'-UTR and the first 26 codons decreas
ed more than 3-fold when the adh1 3'-UTR was removed. In addition, the
adh1 3'-UTR slightly inhibited expression in aerobic protoplasts. The
physical half-lives of the GUS and LUC mRNAs were similar under both
anaerobic and hypoxic conditions, indicating that expression levels we
re largely independent of mRNA stability. Thus, both adh1 5' and 3' mR
NA sequences are required for enhanced translation in protoplasts unde
r O-2 deprivation.