Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.1.30.2 (endonuclease)
 18,621 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

RNA editing in Trypanosoma brucei inserts and deletes uridines in mitochondrial mRNAs by a series of enzymatic steps that are catalyzed by a multiprotein complex, the editosome. KREPB1 and two related editosome proteins KREPB2 and KREPB3 contain motifs that suggest endonuclease and RNA/protein interaction functions. Repression of KREPB1 expression in procyclic forms by RNAi inhibited growth, in vivo editing, and in vitro endoribonucleolytic cleavage of deletion substrates. However, cleavage of insertion substrates and the exoUase, TUTase, and ligase catalytic activities of editing were retained by 20S editosomes. Repression of expression of an ectopic KREPB1 allele in bloodstream forms lacking both endogenous alleles or exclusive expression of KREPB1 with point mutations in the putative RNase III catalytic domain also blocked growth, in vivo editing, and abolished cleavage of deletion substrates, without affecting the other editing steps. These data indicate that KREPB1 is an endoribonuclease that is specific for RNA editing deletion sites.
 ...
PMID:A deletion site editing endonuclease in Trypanosoma brucei. 1628 22

RNA editing in the sleeping sickness parasite Trypanosoma brucei remodels mitochondrial transcripts by the addition and deletion of uridylates as specified by guide RNAs. Editing is catalyzed by at least three distinct approximately 20S multiprotein editosomes, all of which contain KREPB4, a protein with RNase III and Pumilio motifs. RNAi repression of KREPB4 expression in procyclic forms (PFs) strongly inhibited growth and in vivo RNA editing, greatly diminished the abundance of 20S editosomes, reduced cellular levels of editosome proteins, and generated approximately 5-10S editosome subcomplexes. Editing TUTase, exoUase, and RNA ligase activities were largely shifted from approximately 20S to approximately 5-10S fractions of cellular lysates. Insertion and deletion endonuclease activities in approximately 20S fractions were strongly reduced upon KREPB4 repression but were not detected in the 5-10S subcomplex fraction. Abundance of MRP1 and RBP16 proteins, which appear to be involved in RNA processing but are not components of the 20S editosome, was unaltered upon KREPB4 repression. These data suggest that KREPB4 is important for the structural integrity of approximately 20S editosomes, editing endonuclease activity, and the viability of PF T. brucei cells.
 ...
PMID:An essential role of KREPB4 in RNA editing and structural integrity of the editosome in Trypanosoma brucei. 1736 11

Trypanosome mitochondrial mRNAs achieve their coding sequences through RNA editing. This process, catalyzed by approximately 20S protein complexes, involves large numbers of uridylate (U) insertions and deletions within mRNA precursors. Here we analyze the role of the essential TbMP42 protein (band VI/KREPA2) by individually examining each step of the U-deletional and U-insertional editing cycles, using reactions in the approximately linear range. We examined control extracts and RNA interference (RNAi) extracts prepared soon after TbMP42 was depleted (when primary effects should be most evident) and three days later (when precedent shows secondary effects can become prominent). This analysis shows TbMP42 is critical for cleavage of editing substrates by both the U-deletional and U-insertional endonucleases. However, on simple substrates that assess cleavage independent of editing features, TbMP42 is similarly required only for the U-deletional endonuclease, indicating TbMP42 affects the two editing endonucleases differently. Supplementing RNAi extract with recombinant TbMP42 partly restores these cleavage activities. Notably, we find that all the other editing steps (the 3'-U-exonuclease [3'-U-exo] and ligation steps of U-deletion and the terminal-U-transferase [TUTase] and ligation steps of U-insertion) remain at control levels upon RNAi induction, and hence are not dependent on TbMP42. This contrasts with an earlier report that TbMP42 is a 3'-U-exo that may act in U-deletion and additionally is critical for the TUTase and/or ligation steps of U-insertion, observations our data suggest reflect indirect effects of TbMP42 depletion. Thus, trypanosomes require TbMP42 for both endonucleolytic cleavage steps of RNA editing, but not for any of the subsequent steps of the editing cycles.
 ...
PMID:Trypanosoma brucei RNA editing protein TbMP42 (band VI) is crucial for the endonucleolytic cleavages but not the subsequent steps of U-deletion and U-insertion. 1844 Oct 50

The transcriptome of kinetoplastid mitochondria undergoes extensive RNA editing that inserts and deletes uridine residues (U's) to produce mature mRNAs. The editosome is a multiprotein complex that provides endonuclease, TUTase, exonuclease, and ligase activities required for RNA editing. The editosome's KREPB4 and KREPB5 proteins are essential for editosome integrity and parasite viability and contain semi-conserved motifs corresponding to zinc finger, RNase III, and PUF domains, but to date no functional analysis of these domains has been reported. We show here that various point mutations to KREPB4 and KREPB5 identify essential domains, and suggest that these proteins do not themselves perform RNase III catalysis. The zinc finger of KREPB4 but not KREPB5 is essential for editosome integrity and parasite viability, and mutation of the RNase III signature motif in KREPB5 prevents integration into editosomes, which is lethal. Isolated TAP-tagged KREPB4 and KREPB5 complexes preferentially associate with components of the deletion subcomplex, providing additional insights into editosome architecture. A new alignment of editosome RNase III sequences from several kinetoplastid species implies that KREPB4 and KREPB5 lack catalytic activity and reveals that the PUF motif is present in the editing endonucleases KREN1, KREN2, and KREN3. The data presented here are consistent with the hypothesis that KREPB4 and KREPB5 form intermolecular heterodimers with the catalytically active editing endonucleases, which is unprecedented among known RNase III proteins.
 ...
PMID:Mutational analysis of Trypanosoma brucei editosome proteins KREPB4 and KREPB5 reveals domains critical for function. 2291 50