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KRBP72 facilitates ATPase-dependent editing progression through a structural roadblock in mitochondrial A6 mRNA

ABSTRACT: KRBP72 facilitates ATPase-dependent editing progression through a structural roadblock in mitochondrial A6 mRNA

PROVIDER: PRJNA1136147 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR29837649.fastq.gz	Fastqsanger.gz
	SRR29837650.fastq.gz	Fastqsanger.gz
	SRR29837651.fastq.gz	Fastqsanger.gz

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KRBP72 facilitates ATPase-dependent editing progression through a structural roadblock in mitochondrial A6 mRNA

Project description:Uridine insertion/deletion editing of mitochondrial mRNAs in kinetoplastids entails the coordinated action of three complexes. RNA Editing Catalytic Complexes (RECCs) catalyze the enzymatic reactions, while the RNA Editing Substrate Binding Complex (RESC) and RNA Editing Helicase 1 Complex coordinate interactions between RECCs, mRNAs, and hundreds of gRNAs that direct edited sequences. Additionally, numerous auxiliary factors are required for productive editing of specific mRNAs. Here, we elucidate the role of KRBP72, an editing auxiliary factor of the ABC ATPase family that exhibits RNA binding activity. In procyclic form T. brucei, KRBP72 knockdown leads to a pause in editing at the base of a predicted stem loop structure in ATP synthase subunit 6 (A6) mRNA. Enhanced cross-linking and affinity purification revealed KRBP72 binding sites both within and upstream of this stem loop. KRBP72 ATPase activity is essential for its A6 mRNA editing function; however, its RNA binding activity is dispensable. KRBP72 interacts with most RESC proteins in an RNA-dependent manner. By contrast, RESC12A associates with KRBP72 in an RNA-independent fashion, and RESC12A strongly promotes KRBP72’s interaction with RNA. Hence, KRBP72 ATPase activity facilitates the progression of editing through a challenging RNA secondary structure, highlighting this protein’s crucial role in A6 mRNA editing.

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Project description:The bacterial transcription factor RpoN regulates an extensive network of genes whose products are involved in diverse biological functions. We constructed a small peptide termed the RpoN molecular roadblock, which binds to and blocks transcription from RpoN promoters. This RpoN molecular roadblock can be used in any bacterium to obtain information on the RpoN regulon. We expressed the RpoN molecular roadblock in P. aeruginosa PAO1 and used microarrays to identify genes that were differentially transcribed due to the RpoN molecular roadblock. The RpoN molecular roadblock was expressed in P. aeruginosa PAO1 in mid-exponential phase in rich media. Total RNA was isolated and prepped for Affymetrix GeneChips.

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