Engineering riboswitch in L. major: From prediction to conceptualization

Post-transcriptional gene regulation is a vital process to regulate expression of the key genes in the eukaryotic cell. Such processes are essential for pathogens which reside inside the host cell. One such pathogen is Leishmania major, which causes cutaneous leishmaniasis. The parasite lives inside the macrophages of mammalian host (mostly human). Inside the macrophage, Leishmania genes show complex host-pathogen interaction regulating a plethora of gene expression. Till date, most of the studies have shown this kind of regulation with respect to the host macrophages. Here, based on an extensive in silico analysis, we have hypothesized a novel Theophylline binding riboswitch mediated post-transcriptional regulation of a gene i.e. RNA Polymerase III subunit1 (Lmjf_09_1060), an essential gene for the parasite's survival both in its promastigote as well as in its amastigote form. Later, we have conceptualized the working of the identified putative Theophylline binding riboswitch cassette in in vitro using E. coli based reporter assay, wherein, a reporter gene (eGFP) is used instead of RNA Polymerase III subunit1 gene and apparently have shown the downregulation of the reporter gene (eGFP) expression under the influence of in silico identified Theophylline binding riboswitch.