Study of NAT10 acetyltransferase in chromatin structure and gene expression regulation in Leishmania

Abstract

Leishmania spp. causes leishmaniases, and during its life cycle, the parasite shifts between two hosts and needs to adapt to various environmental conditions to survive. This adaptation could involve changes in gene expression and translation, which are regulated by chromatin structure modifications and post-transcriptional mechanisms. Recently, it was observed that mRNAs can be modified by acetylation in mammalian cells, a modification called N4-acetylcytidine (ac4C), catalyzed by the N-acetyltransferase (NAT10). Depending on the position ac4C is added could greatly impact the stability and/or the translation efficiency of the mRNA. NAT10 can act also as a protein acetyltransferase, and contribute to cellular processes independent of its function as an RNA acetyltransferase, including regulation of nuclear architecture, and nucleosome structure. Considering the need for Leishmania to adapt to different host environments and the described functions for NAT10, we decided to study the possible role of this enzyme in the chromatin dynamics regulation in the parasite. Preliminary data from our group found a L. mexicana NAT10 orthologue with all the required domains and structural characteristics described in other organisms. Attempts to generate NAT10 null parasites revealed that this gene is essential for the parasite, and as an alternative to studying this enzyme, we obtained NAT10 overexpressing parasites, that were confirmed by Western blotting and will be used in the experiments proposed here. Finally, TEM analysis of NAT10 single-knockout parasites showed significant alterations in chromatin structure. Thus, in this BEPE proposal, we will join forces with Dr. Langlais' expertise in functional genomics to study NAT10 epigenetics role: i) investigate the changes in chromatin accessibility of parasites overexpressing NAT10 by ATAC-seq experiments; ii) identify the chromatin binding regions of NAT10 by ChIP-seq; iii) analyze the changes in gene expression in parasites overexpressing NAT10. The results generated by this project will contribute to our knowledge about Leishmania gene expression regulation, a still poorly understood process.