The 3'-untranslated region of cytochrome oxidase II mRNA functions in RNA editing of African trypanosomes exclusively as a cis guide RNA

AbstractBoth adenine base editors (ABEs) and cytosine base editors (CBEs) have been recently revealed to induce transcriptome-wide RNA off-target editing in a guide RNA-independent manner. Here we construct a reporter system containing E.coli Hokb gene with a tRNA-like motif for robust detection of RNA editing activities as the optimized ABE, ABEmax, induces highly efficient A-to-I (inosine) editing within an E.coli tRNA-like structure. Then, we design mutations to disrupt the potential interaction between TadA and tRNAs in structure-guided principles and find that Arginine 153 (R153) within TadA is essential for deaminating RNAs with core tRNA-like structures. Two ABEmax or mini ABEmax variants (TadA* fused with Cas9n) with deletion of R153 within TadA and/or TadA* (named as del153/del153* and mini del153) are successfully engineered, showing minimized RNA off-targeting, but comparable DNA on-targeting activities. Moreover, R153 deletion in recently reported ABE8e or ABE8s can also largely reduce their RNA off-targeting activities. Taken together, we develop a strategy to generate engineered ABEs (eABEs) with minimized RNA off-targeting activities.

Download Full-text

Identification of Diatraea spp. (Lepidoptera: Crambidae) based on cytochrome oxidase II

PLoS ONE ◽

10.1371/journal.pone.0184053 ◽

2017 ◽

Vol 12 (9) ◽

pp. e0184053 ◽

Cited By ~ 5

Author(s):

Gloria Patricia Barrera ◽

Laura Fernanda Villamizar ◽

Carlos Espinel ◽

Edgar Mauricio Quintero ◽

Mariano Nicolás Belaich ◽

...

Keyword(s):

Cytochrome Oxidase ◽

Cytochrome Oxidase Ii

Download Full-text

Upgraded adenine base editor (uABE) with minimized RNA off-targeting activity

10.21203/rs.3.rs-73017/v1 ◽

2020 ◽

Author(s):

Jianan Li ◽

Wenxia Yu ◽

Shisheng Huang ◽

Susu Wu ◽

Liping Li ◽

...

Keyword(s):

Rna Editing ◽

Reporter System ◽

Independent Manner ◽

Robust Detection ◽

Guide Rna ◽

Highly Efficient ◽

Adenine Base

Abstract Both adenine base editors (ABEs) and cytosine base editors (CBEs) have been recently revealed to induce transcriptome-wide RNA off-target editing in a guide RNA-independent manner. As the optimized ABE, ABEmax, induces highly efficient A-to-I (inosine) editing within an E.coli tRNA-like structure, we construct a reporter system containing E.coli Hokb gene with a tRNA-like motif for robust detection of RNA editing activities. Then, we design mutations to disrupt the interaction between TadA and tRNAs in structure-guided principles, and find that Arginine 153 (R153) within TadA is essential for recognizing core tRNA-like structures. Two ABEmax or mini ABEmax variants (TadA* fused with Cas9 (D10A)) with deletion of R153 within TadA and/or TadA* (named as del153/del153* and mini del153) are successfully engineered, showing minimized RNA editing, but comparable DNA on-targeting activities. Moreover, del153 in recently reported ABE8e or ABE8s can also largely reduce their RNA off-targeting activities. Taken together, we develop a strategy to generate upgraded ABEs (uABEs) with minimized RNA off-target activities.

Download Full-text

Mitochondrial DNAs provide insight into trypanosome phylogeny and molecular evolution

10.21203/rs.3.rs-27858/v1 ◽

2020 ◽

Author(s):

Christopher Kay ◽

Tom A Williams ◽

Wendy Gibson

Keyword(s):

Molecular Evolution ◽

Trypanosoma Brucei ◽

Rna Editing ◽

Phylogenetic Trees ◽

Nucleotide Composition ◽

Sub Saharan Africa ◽

African Trypanosomes ◽

Animal Pathogens ◽

Recent Emergence ◽

The Impact

Abstract Background: Trypanosomes are single-celled eukaryotic parasites characterised by the unique biology of their mitochondrial DNA (mtDNA). African livestock trypanosomes impose a major burden on agriculture across sub-Saharan Africa, but are poorly understood compared to those that cause sleeping sickness and Chagas disease in humans. Here we explore the potential of trypanosome mtDNA to study the evolutionary history of trypanosomes and the molecular evolution of their mtDNAs.Results: We used long-read sequencing to completely assemble mtDNAs from four previously uncharacterized African trypanosomes, and leveraged these assemblies to scaffold and assemble a further 103 trypanosome mtDNAs from published short-read data. While synteny was largely conserved, there were repeated, independent losses of Complex I genes. Comparison of edited and non-edited genes revealed the impact of RNA editing on nucleotide composition, with non-edited genes approaching the limits of GC loss. African tsetse-transmitted trypanosomes showed high levels of RNA editing compared to other trypanosomes. Whole mtDNA coding regions were used to construct time-resolved phylogenetic trees, revealing deep divergence events among isolates of the pathogens Trypanosoma brucei and T. congolense .Conclusions: Our mtDNA data represents a new resource for experimental and evolutionary analyses of trypanosome phylogeny, molecular evolution and function. Molecular clock analyses yielded a timescale for trypanosome evolution congruent with major biogeographical events in Africa and revealed the recent emergence of Trypanosoma brucei gambiense and T. equiperdum , major human and animal pathogens.

Download Full-text