Selenocysteine insertion during selenoprotein biosynthesis
begins with the aminoacylation of selenocysteine tRNA[Ser]Sec
with serine, the conversion of the serine moiety to selenocysteine,
and the recognition of specific UGA codons within the mRNA.
Selenocysteine tRNA[Ser]Sec exists
as two major forms, differing by methylation of the ribose
portion of the nucleotide at the wobble position of the
anticodon. The levels and relative distribution of these
two forms of the tRNA are influenced by selenium in mammalian
cells and tissues. We have generated Chinese hamster ovary
cells that exhibit increased levels of tRNA[Ser]Sec
following transfection of the mouse tRNA[Ser]Sec
gene. The levels of selenocysteine tRNA[Ser]Sec
in transfectants increased proportionally to the number
of stably integrated copies of the tRNA[Ser]Sec
gene. Although we were able to generate transfectants overproducing
tRNA[Ser]Sec by as much as tenfold,
the additional tRNA was principally retained in the unmethylated
form. Selenium supplementation could not significantly
affect the relative distributions of the two major selenocysteine
tRNA[Ser]Sec isoacceptors. In addition,
increased levels of tRNA[Ser]Sec did
not result in measurable alterations in the levels of selenoproteins,
including glutathione peroxidase.