Effects of d(CAG)(n).d(CTG)(n) repeats on expression of a reporter gene in
human cell culture were studied using transient transfection, RNase protect
ion and coupled transcription/translation assays. Cloning these repeats int
o the reporter 3'-UTR did not affect gene functioning. In contrast, placing
the repeats in the reporter 5'-UTR led to strong inhibition of expression.
This inhibition depended on the repeat orientation, being prominent only w
hen the (CTG)(n) tracts were in the sense strand for transcription. Further
, the strength of inhibition increased exponentially with an increase in re
peat length. Our data indicate that expanded (CTG)(n) repeats prevent effic
ient translation of the reporter mRNA both in vitro and in vivo. We suggest
that formation of stable hairpins by (CUG)(n) runs of increasing length in
the 5'-UTR of a mRNA progressively inhibits the scanning step of translati
on initiation. This points to a novel mechanism of regulating gene expressi
on by expandable d(CTG)(n) repeats.