The Function of a Central RNA Helicase RNP in Editosomes of Trypanosomes

Abstract

Trypanosoma brucei requires extensive remodeling of its mitochondrial gene expression during different stages of its life cycle. RNA editing is one of the vital processes for these mitochondrial changes. This process involves insertions and deletions of uridylates in mitochondrial mRNAs that is governed by a multi-protein RNA editing core complex (RECC). Editing proceeds in small blocks from 3’ to 5’ direction leaded by small trans-acting guide RNAs (gRNAs). RECC appears to lack both, the mRNA substrates and gRNAs, indicating a role of accessory proteins in RNA editing. Many other non-RECC proteins have been discovered that directly impact the editing process, including mitochondrial RNA-binding complex 1(MRB1) that was shown to contain gRNAs. Despite of all the progress made, central long-standing question still remains unanswered including the mechanism for substrate delivery and regulation of RNA editing process.

This dissertation presents the discovery of two variants of native MRB1 (mitochondrial RNA-binding complex 1) that we termed REH2-MRB and 3010-MRB. These MRB1 variants contain both mRNAs and gRNAs and show specialized functions, 3010-MRB and REH2-MRB seem to serve as scaffolds for RNA editing. REH2-MRB is defined by the critical RNA helicase termed REH2 (RNA editing helicase 2) that acts in trans, affecting multiple steps of the editing function in 3010-MRB. In addition, we discovered two cofactors of REH2. This novel RNA editing helicase 2-associated subcomplex (REH2C) binds mRNA substrates and products and therefore represents a stable mRNA-bound protein subcomplex (mRNP). Our working model is that MRB1 variant complexes are formed by the coupling of this mRNP with gRNA-bound subcomplexes (gRNPs). The mRNP/gRNP complexes form a platform for the assembly of functional mRNA-gRNA hybrids and catalytic RECC enzyme. Thus, in our proposed model editosomes are assembled in a stepwise process that involves the docking of mRNP and gRNP modules through specific base-pairing of respective mRNA and gRNAs. These subunits of the REH2C may control specific checkpoints in the editing pathway.