An insertion in the methyltransferase domain of P. falciparum trimethylguanosine synthase harbors a classical nuclear localization signal

[Display omitted] •The methyltransferase domain of P. falciparum TGS1 (PfTGS1) has an insertion containing a predicted nuclear localization signal (NLS).•This NLS is functional in yeast and is recognized by P. falciparum importin-α in vitro.•Assays developed in this report can be used to study NLS from Plasmodium species that cannot be cultured in the laboratory. Many Plasmodium falciparum proteins do not share homology with, and are generally longer than their respective orthologs. This, to some extent, can be attributed to insertions. Here, we studied a P. falciparum RNA hypermethylase, trimethylguanosine synthase (PfTGS1) that harbors a 76 amino acid insertion in its methyltransferase domain. Bioinformatics analysis revealed that this insertion was present in TGS1 orthologs from other Plasmodium species as well. Interestingly, a classical nuclear localization signal (NLS) was predicted in the insertions of primate parasite TGS1 proteins. To check whether these predicted NLS are functional, we developed an in vivo heterologous system using S. cerevisiae. The predicted NLS when fused to dimeric GFP were able to localize the fusion protein to the nucleus in yeast indicating that it is indeed recognized by the yeast nuclear import machinery. We further showed that the PfTGS1 NLS binds to P. falciparum importin-α in vitro, confirming that the NLS is also recognized by the P. falciparum classical nuclear import machinery. Thus, in this study we report a novel function of the insertion in PfTGS1.