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Progression of a loop–loop complex to a four‐way junction is crucial for the activity of a regulatory antisense RNA
Author(s) -
Kolb Fabrice A.,
Engdahl Hilde M.,
SlagterJäger Jacoba G.,
Ehresmann Bernard,
Ehresmann Chantal,
Westhof Eric,
Wagner E.Gerhart H.,
Romby Pascale
Publication year - 2000
Publication title -
the embo journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 7.484
H-Index - 392
eISSN - 1460-2075
pISSN - 0261-4189
DOI - 10.1093/emboj/19.21.5905
Subject(s) - biology , loop (graph theory) , antisense rna , rna , stem loop , microbiology and biotechnology , genetics , computational biology , gene , mathematics , combinatorics
The antisense RNA, CopA, regulates the replication frequency of plasmid R1 through inhibition of RepA translation by rapid and specific binding to its target RNA (CopT). The stable CopA–CopT complex is characterized by a four‐way junction structure and a side‐by‐side alignment of two long intramolecular helices. The significance of this structure for binding in vitro and control in vivo was tested by mutations in both CopA and CopT. High rates of stable complex formation in vitro and efficient inhibition in vivo required initial loop–loop complexes to be rapidly converted to extended interactions. These interactions involve asymmetric helix progression and melting of the upper stems of both RNAs to promote the formation of two intermolecular helices. Data presented here delineate the boundaries of these helices and emphasize the need for unimpeded helix propagation. This process is directional, i.e. one of the two intermolecular helices (B) must form first to allow formation of the other (B′). A binding pathway, characterized by a hierarchy of intermediates leading to an irreversible and inhibitory RNA–RNA complex, is proposed.