Nucleic Acids Research, 1994, Vol. 22, No. 2 222-231
© 1994
MOLECULAR BIOLOGY |
Upstream and downstream c/s-acting elements for cleavage at the L4 polyadenylation site of adenovirus-2
Laboratoire de Génèwtique Moléculaire des Eucaryotes du CNRS, Unité 184 de Biologie Moleculaire et de Génie Génétique de I'lNSERM, Institut de Chimie Biologique, Faculté de Médecine 11 rue Humann, 67085 Strasbourg Cédex, France
*To whom correspondence should be addressed
Received September 13, 1993. Accepted December 13, 1993.
A study of the cis-actlng elements Involved in the 3' end formation of the RNAs from the major late L4 family of adenovirus-2 was undertaken. Series of 5' or 3' end deletion mutants and mutants harboring either Internal deletions or substitutions were prepared and assayed for In vitro cleavage. This first allowed the demonstration of a sequence, located at –6 to –29, relative to AAUAAA, whose deletion or substitution reduces cleavage efficiency at the L4 polyadenylation site two to three fold. This upstream efficiency element 5' AUCUUUGUUGUC/AUCUCUGUGCUG 3' is constituted of a partially repeated 12 nucleotlde long, UCG rich sequence. The activities of the 2 sequence elements in cleavage are additive. We also searched for regulatory sequences downstream of the L4 polyadenylation site. We found that the deletion or substitution of a 30 nucleotlde long UCG rich sequence, between nucleotides +7 and +35 relative to the cleavage site and harboring a UCCUGU repeat reduces cleavage efficiency at least ten fold. A GUUUUU sequence, starting at +35 had no influence. Thus, the usage of the L4 polyadenylation site requires downstream sequences different from the canonical GU or U boxes and is regulated by upstream sequence elements.
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