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Roles for ligases in the RNA editing complex of Trypanosoma brucei : band IV is needed for U‐deletion and RNA repair
Author(s) -
Huang Catherine E.,
CruzReyes Jorge,
Zhelonkina Alevtina G.,
O'Hearn Sean,
Wirtz Elizabeth,
SollnerWebb Barbara
Publication year - 2001
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/20.17.4694
Subject(s) - biology , trypanosoma brucei , rna editing , rna , genetics , computational biology , guide rna , microbiology and biotechnology , gene , crispr , genome editing
Trypanosome RNA editing utilizes a seven polypeptide complex that includes two RNA ligases, band IV and band V. We now find that band IV protein contributes to the structural stability of the editing complex, so its lethal genetic knock‐out could reflect structural or catalytic requirements. To assess the catalytic role in editing, we generated cell lines which inducibly replaced band IV protein with an enzymatically inactive but structurally conserved version. This induction halts cell growth, showing that catalytic activity is essential. These induced cells have impaired in vivo editing, specifically of RNAs requiring uridylate (U) deletion; unligated RNAs cleaved at U‐deletion sites accumulated. Additionally, mitochondrial extracts of cells with reduced band IV activity were deficient in catalyzing U‐deletion, specifically at its ligation step, but were not deficient in U‐insertion. Thus band IV ligase is needed to seal RNAs in U‐deletion. U‐insertion does not appear to require band IV, so it might use the other ligase of the editing complex. Furthermore, band IV ligase was also found to serve an RNA repair function, both in vitro and in vivo .