StitchR: Ribozyme-activated mRNA Trans-ligation and Translation
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA1164324 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR30775686_1.fastq.gz | Fastqsanger.gz | |||
| SRR30775686_2.fastq.gz | Fastqsanger.gz | |||
| SRR30775687_1.fastq.gz | Fastqsanger.gz | |||
| SRR30775687_2.fastq.gz | Fastqsanger.gz |
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Project description:Conventional reverse genetic approaches for study of Plasmodium malaria parasite gene function are limited, or not applicable. Hence, new inducible systems are needed. Here we describe a method to control P. falciparum gene expression in which target genes bearing a glmS ribozyme in the 3′ untranslated region (3′-UTR) are efficiently knocked down in transgenic P. falciparum parasites in response to exogenous glucosamine (GlcN) inducer. Using reporter genes, we show that the glmS ribozyme cleaves reporter mRNA in vivo leading to reduction in mRNA expression following GlcN treatment. GlcN-induced ribozyme activation also led to efficient reduction of reporter protein, which could be rapidly reversed by removing the inducer. The glmS ribozyme was validated as a reverse-genetic tool by integration into the essential gene and antifolate drug target dihydrofolate reductase-thymidylate synthase (PfDHFR-TS). GlcN treatment of transgenic parasites led to rapid and efficient knockdown of PfDHFR-TS mRNA and protein. PfDHFR-TS knockdown led to a growth/arrest mutant phenotype and hypersensitivity to pyrimethamine. The glmS ribozyme is thus an important tool for study of P. falciparum essential genes and anti-malarial drug discovery.
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