Volume 272, Number 2,
Issue of January 10, 1997
pp. 984-991
©1997 by The American Society for Biochemistry and Molecular Biology, Inc.
Identification and Characterization of a Nuclease Specific
for the 3
End of the U6 Small Nuclear RNA
(Received for publication, July 22, 1996, and in revised form, October 4, 1996)
Bruce L.
Booth
Jr.
and
B. Franklin
Pugh
From the Center for Gene Regulation, Department of Biochemistry and
Molecular Biology, The Pennsylvania State University, University
Park, Pennsylvania 16802
The U6 small nuclear RNA is
post-transcriptionally processed by the addition and removal of
nontemplate uridylates (Us) at its 3
end prior to incorporation into
the U4·U6 small nuclear ribonucleoprotein complex. An enzyme
responsible for removing Us from the U6 3 end has not been previously
identified. Here we biochemically isolate and characterize an
exonuclease activity from HeLa cells that removes template and
nontemplate 3-nucleotides specifically from U6 RNA. We also report the
isolation of an inhibitor of this U6 nuclease. U6 nuclease rapidly
removes the four terminal 3 Us found in the human U6 coding sequence
in a magnesium-dependent manner. Mutagenesis studies on the
U6 RNA define regions essential for processing. U6 nuclease recognizes
specific sequences on both strands near the base of the major
intramolecular stem loop of the U6 RNA. The preprocessed 3 Us form
part of the base of the stem loop, but neither specific sequences nor
secondary structure at the four terminal nucleotides are required to
achieve processing.
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