Trypanosome REH1 is an RNA helicase involved with the 3′–5′ polarity of multiple gRNA-guided uridine insertion/deletion RNA editing

Li, Feng; Herrera, Jeremy; Zhou, Shi-Sheng; Maslov, Dmitri; Simpson, Larry

doi:10.1073/pnas.1014152108

Cited by 52 publications

(57 citation statements)

References 37 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Whether or not this activity affects editing substrate binding to other MRBs needs to be examined. REH2-dependent unwinding could also control global intra-strand mRNA structure or gRNA exchange during editing progression, as it was proposed for TbRGG2 and REH1, respectively (Ammerman et al 2010;Li et al 2011). It was proposed that RNAi-repression of REH2 reduces gRNA stability (Hashimi et al 2009).…”

Section: Discussionmentioning

confidence: 99%

Native mitochondrial RNA-binding complexes in kinetoplastid RNA editing differ in guide RNA composition

Madina

Kumar

Metz

et al. 2014

RNA

136

View full text Add to dashboard Cite

Mitochondrial mRNAs in kinetoplastids require extensive U-insertion/deletion editing that progresses 3 ′ -to-5 ′ in small blocks, each directed by a guide RNA (gRNA), and exhibits substrate and developmental stage-specificity by unsolved mechanisms. Here, we address compositionally related factors, collectively known as the mitochondrial RNA-binding complex 1 (MRB1) or gRNAbinding complex (GRBC), that contain gRNA, have a dynamic protein composition, and transiently associate with several mitochondrial factors including RNA editing core complexes (RECC) and ribosomes. MRB1 controls editing by still unknown mechanisms. We performed the first next-generation sequencing study of native subcomplexes of MRB1, immunoselected via either RNA helicase 2 (REH2), that binds RNA and associates with unwinding activity, or MRB3010, that affects an early editing step. The particles contain either REH2 or MRB3010 but share the core GAP1 and other proteins detected by RNA photo-crosslinking. Analyses of the first editing blocks indicate an enrichment of several initiating gRNAs in the MRB3010-purified complex. Our data also indicate fast evolution of mRNA 3 ′ ends and strain-specific alternative 3 ′ editing within 3 ′ UTR or C-terminal protein-coding sequence that could impact mitochondrial physiology. Moreover, we found robust specific copurification of edited and pre-edited mRNAs, suggesting that these particles may bind both mRNA and gRNA editing substrates. We propose that multiple subcomplexes of MRB1 with different RNA/protein composition serve as a scaffold for specific assembly of editing substrates and RECC, thereby forming the editing holoenzyme. The MRB3010-subcomplex may promote early editing through its preferential recruitment of initiating gRNAs.

show abstract

Section: Discussionmentioning

confidence: 99%

Native mitochondrial RNA-binding complexes in kinetoplastid RNA editing differ in guide RNA composition

Madina

Kumar

Metz

et al. 2014

RNA

136

View full text Add to dashboard Cite

show abstract

“…Resolution of both inter-and intramolecular RNA structure has long been recognized as essential for proper 3 ′ to 5 ′ progression of editing, and several protein factors that may contribute to this process have been described. These include the helicases REH1 (Li et al 2011) and REH2 (Hernandez et al 2010;Madina et al 2014Madina et al , 2015, the 6A). The bottom panel shows the abundance of pausing at each editing site in RPS12 (replicate 1), and intrinsic pause sites are denoted by an asterisk (from Fig.…”

Section: Discussionmentioning

confidence: 99%

“…That gRNA then needs to be at least partially removed because the subsequent gRNA forms an anchor duplex with a portion of the edited region guided by the first gRNA (Maslov and Simpson 1992). The mechanism of gRNA exchange is poorly understood, but may involve mitochondrial helicases (Hernandez et al 2010;Li et al 2011;Madina et al 2014Madina et al , 2015. This gRNA utilization process proceeds throughout the length of a given mRNA, resulting in the general, although not precise, progression of editing from the 3 ′ to the 5 ′ end of the transcript (Sturm and Simpson 1990;Koslowsky et al 1991;Souza et al 1992).…”

Section: Introductionmentioning

confidence: 99%

High-throughput sequencing of partially edited trypanosome mRNAs reveals barriers to editing progression and evidence for alternative editing

Simpson

Bruno

Bard

et al. 2016

RNA

145

View full text Add to dashboard Cite

Uridine insertion/deletion RNA editing in kinetoplastids entails the addition and deletion of uridine residues throughout the length of mitochondrial transcripts to generate translatable mRNAs. This complex process requires the coordinated use of several multiprotein complexes as well as the sequential use of noncoding template RNAs called guide RNAs. The majority of steadystate mitochondrial mRNAs are partially edited and often contain regions of mis-editing, termed junctions, whose role is unclear. Here, we report a novel method for sequencing entire populations of pre-edited partially edited, and fully edited RNAs and analyzing editing characteristics across populations using a new bioinformatics tool, the Trypanosome RNA Editing Alignment Tool (TREAT). Using TREAT, we examined populations of two transcripts, RPS12 and ND7-5 ′ , in wild-type Trypanosoma brucei. We provide evidence that the majority of partially edited sequences contain junctions, that intrinsic pause sites arise during the progression of editing, and that the mechanisms that mediate pausing in the generation of canonical fully edited sequences are distinct from those that mediate the ends of junction regions. Furthermore, we identify alternatively edited sequences that constitute plausible alternative open reading frames and identify substantial variability in the 5 ′ UTRs of both canonical and alternatively edited sequences. This work is the first to use high-throughput sequencing to examine full-length sequences of whole populations of partially edited transcripts. Our method is highly applicable to current questions in the RNA editing field, including defining mechanisms of action for editing factors and identifying potential alternatively edited sequences.

show abstract

“…The anchor helix and U-helix may keep the gRNA's template sequence bound to the premRNA after cleavage of the pre-mRNA during editing, and thereby keep the two internal ends of the cleaved mRNA in close proximity for religation (Koslowsky et al 2004). After the editosome has finished using the template domain of a particular gRNA, a RNA editing-specific helicase (Li et al 2011) displaces the gRNA strand from the mRNA, a different gRNA binds to the next ABS 59 to the last editing site on the mRNA, and the editing reaction cycles continue.…”

Section: Introductionmentioning

confidence: 99%

The crystal structure of an oligo(U):pre-mRNA duplex from a trypanosome RNA editing substrate

Mooers¹,

Singh²

2011

RNA

View full text Add to dashboard Cite

Guide RNAs bind antiparallel to their target pre-mRNAs to form editing substrates in reaction cycles that insert or delete uridylates (Us) in most mitochondrial transcripts of trypanosomes. The 59 end of each guide RNA has an anchor sequence that binds to the pre-mRNA by base-pair complementarity. The template sequence in the middle of the guide RNA directs the editing reactions. The 39 ends of most guide RNAs have~15 contiguous Us that bind to the purine-rich unedited pre-mRNA upstream of the editing site. The resulting U-helix is rich in G d U wobble base pairs. To gain insights into the structure of the U-helix, we crystallized 8 bp of the U-helix in one editing substrate for the A6 mRNA of Trypanosoma brucei. The fragment provides three samples of the 59-AGA-39/59-UUU-39 base-pair triple. The fusion of two identical U-helices head-to-head promoted crystallization. We obtained X-ray diffraction data with a resolution limit of 1.37 Å . The U-helix had low and high twist angles before and after each G d U wobble base pair; this variation was partly due to shearing of the wobble base pairs as revealed in comparisons with a crystal structure of a 16-nt RNA with all Watson-Crick base pairs. Both crystal structures had wider major grooves at the junction between the poly(U) and polypurine tracts. This junction mimics the junction between the template helix and the U-helix in RNA-editing substrates and may be a site of major groove invasion by RNA editing proteins.

show abstract

Trypanosome REH1 is an RNA helicase involved with the 3′–5′ polarity of multiple gRNA-guided uridine insertion/deletion RNA editing

Cited by 52 publications

References 37 publications

Native mitochondrial RNA-binding complexes in kinetoplastid RNA editing differ in guide RNA composition

Native mitochondrial RNA-binding complexes in kinetoplastid RNA editing differ in guide RNA composition

High-throughput sequencing of partially edited trypanosome mRNAs reveals barriers to editing progression and evidence for alternative editing

The crystal structure of an oligo(U):pre-mRNA duplex from a trypanosome RNA editing substrate

Contact Info

Product

Resources

About