Reconstitution of uridine-deletion precleaved RNA editing with two recombinant enzymes.
Uridine insertion/deletion RNA editing in trypanosomatid mitochondria is a posttranscriptional RNA modification phenomenon required for translation of mitochondrial mRNAs. This process involves guide RNA-mediated cleavage at a specific site, insertion or deletion of Us from the 3' end of the 5' mRNA fragment, and ligation of the two mRNA fragments. The Leishmania major RNA ligase-containing complex protein 2 expressed in insect cells has a 3'-5' exoribonuclease activity and was therefore renamed RNA editing exonuclease 1 (REX1). Recombinant REX1 specifically trims 3' overhanging Us and stops at a duplex region. Evidence is presented that REX1 is responsible for deletion of the 3' overhanging Us from the bridged mRNA 5' cleavage fragment and that RNA editing ligase 1 is responsible for the ligation of the two mRNA cleavage fragments in U-deletion editing. The evidence involves both in vivo down-regulation of REX1 expression in Trypanosoma brucei by RNA interference and the reconstitution of precleaved U-deletion in vitro editing with only two recombinant enzymes: recombinant REX1 and recombinant RNA editing ligase 1.
Kang
Xuedong
X
Howard Hughes Medical Institute and Department of Microbiology, Immunology, and Molecular Genetics, University of California, Los Angeles, CA 90095, USA.
Rogers
Kestrel
K
Gao
Guanghan
G
Falick
Arnold M
AM
Zhou
Sharleen
S
Simpson
Larry
L
D000818:Animals; D019729:Carbon-Oxygen Ligases / Q000502:physiology*; D024101:Mitochondrial Proteins / Q000502:physiology*; D017393:RNA Editing; D011994:Recombinant Proteins / Q000494:pharmacology; D014346:Trypanosoma brucei brucei / Q000235:genetics*; D014529:Uridine / Q000378:metabolism*
D024101:Mitochondrial Proteins; D011994:Recombinant Proteins; D019729:Carbon-Oxygen Ligases; C476580:REL1 RNA ligase, Trypanosoma brucei; C476581:REL2 RNA ligase, Trypanosoma brucei; D014529:Uridine