Engineering riboswitch in L. major: From prediction to conceptualization

Area of Science:

• Molecular Biology
• Parasitology
• Bioinformatics

Background:

• Post-transcriptional gene regulation is crucial for eukaryotic gene expression, particularly for intracellular pathogens like Leishmania major.
• Leishmania major resides within host macrophages, exhibiting complex host-pathogen interactions that modulate gene expression.

Purpose of the Study:

• To hypothesize and investigate a novel theophylline-binding riboswitch-mediated post-transcriptional regulation mechanism for the essential RNA Polymerase III subunit 1 gene (Lmjf_09_1060) in Leishmania major.
• To validate the function of the identified riboswitch in vitro.

Main Methods:

• Extensive in silico analysis to identify the putative theophylline-binding riboswitch.
• In vitro reporter assay using E. coli, replacing the target gene with eGFP to assess riboswitch function.

Main Results:

• Hypothesized a novel theophylline-binding riboswitch regulating the essential RNA Polymerase III subunit 1 gene in Leishmania major.
• Demonstrated downregulation of the reporter gene (eGFP) expression in vitro, supporting the in silico identified riboswitch's regulatory role.

Conclusions:

• The study presents a novel mechanism of post-transcriptional gene regulation mediated by a theophylline-binding riboswitch in Leishmania major.
• This finding provides a new understanding of gene regulation in Leishmania parasites and suggests potential targets for therapeutic intervention against leishmaniasis.