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J. Biol. Chem., Vol. 275, Issue 49, 38384-38392, December 8, 2000
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From the Mammalian topoisomerase II
Cell Cycle-coupled Variation in Topoisomerase II
mRNA Is
Regulated by the 3'-Untranslated Region
POSSIBLE ROLE OF REDOX-SENSITIVE PROTEIN BINDING IN mRNA
ACCUMULATION*
§¶,,,,,,,,, and§
Radiation Oncology Center, MIR, Washington
University Medical School, St. Louis, Missouri 63108 and the
§ Free Radical and Radiation Biology Program, University of
Iowa, Iowa City, Iowa 52242
(Topo II) is a
highly regulated enzyme essential for many cellular processes including
the G2 cell cycle checkpoint. Because Topo II gene
expression is regulated posttranscriptionally during the cell cycle, we
investigated the possible role of the 3'-untranslated region (3'-UTR)
in controlling Topo II mRNA accumulation. Reporter assays in stably
transfected cells demonstrated that, similar to endogenous Topo II
mRNA levels, the mRNA levels of reporter genes containing the
Topo II 3'-UTR varied during the cell cycle and were maximal in S and
G2/M relative to G1. Topo II 3'-UTR sequence
analysis and RNA-protein binding assays identified a 177-nucleotide
(base pairs 4772-4949) region containing an AUUUUUA motif sufficient
for protein binding. Multiple proteins (84, 70, 44, and 37 kDa) bound
this region, and the binding of 84- and 37-kDa (tentatively identified
as the adenosine- or uridine-rich element-binding factor AUF1)
proteins was enhanced in G1, correlating with decreased
Topo II mRNA levels. The binding activity was enhanced in cellular
extracts or cells treated with thiol-reducing agents, and increased
binding correlated with decreased Topo II mRNA levels. These
results support the hypothesis that cell cycle-coupled Topo II gene
expression is regulated by interaction of the 3'-UTR with
redox-sensitive protein complexes.
*
This work was supported by National Institutes of Health
Grants R29CA69593 (to P. C. G.), RO1HL51469 (to D. R. S.), CA50950 (to C. R. H.), and K08CA72602 (to D. G.).The costs of publication of this
article were defrayed in part by the
payment of page charges. The article
must therefore be hereby marked
"advertisement" in accordance with 18 U.S.C. Section
1734 solely to indicate this fact.
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