The SET1 protein of Leishmania donovani moderates the parasite's response to a hostile oxidative environment

Abstract

SET domain proteins mediate their effects through the methylation of specific lysine residues on target substrates, resulting in either the stimulation or repression of downstream processes. Initially identified as histone lysine methyltransferases, they are now known to target a wide-ranging conglomeration of non-histone subtrates as well. Twenty-nine SET domain proteins have been identified in Leishmania donovani through genome sequence annotations. This study initiates the first investigation into these proteins. We find LdSET1 is predominantly cytosolic and constitutively expressed. The set1 gene is not essential, although its deletion slows promastigote growth and hypersensitizes the parasite to hydroxyurea-induced G1/S arrest. Intriguingly, set1-nulls proliferate more proficiently than set1+/+ parasites within host macrophages, suggesting that LdSET1 moderates the parasites response to the inhospitable intracellular oxidative environment. set1-null parasites are highly tolerant to H2O2-induced oxidative stress in in vitro promastigote cultures as well, reflected in their growth pattern as well as almost complete absence of DNA damage at the H2O2 concentrations tested. This is linked to ROS levels remaining virtually unperturbed in response to H2O2 treatment in set1-nulls, contrasting to increased ROS in set1+/+ cells under similar conditions. In analyzing the cells ability to scavenge hydroperoxides we find that peroxidase activity is not upregulated in response to H2O2 exposure in set1-nulls. Rather, the constitutive basal levels of peroxidase activity are significantly higher in these cells, implicating this to be a factor contributing to the parasites apparent resistance to H2O2. The higher levels of peroxidase activity in set1-nulls are coupled to upregulation of tryparedoxin peroxidase transcripts in these cells. Thus, LdSET1 tunes the parasites behavior within host cells, enabling establishment and persistence of infection, and maintaining the balance with host without eradicating the host cell population it needs for survival. These findings uncover a new dimension to the mechanisms controlling the interplay of Leishmania-host interactions. Author SummaryLeishmaniases are a group of diseases afflicting people across 88 countries. Manifested in three forms: cutaneous, subcutaneous and visceral, different species cause the different forms of the disease. Leishmania donovani is one of the species causing Visceral Leishmaniasis, and its cellular processes are an area of intensive investigations. These protozoan parasites display an unusual pattern of transcription, and gene regulation occurs primarily through epigenetic mechanisms and post-transcriptional processes. SET proteins control gene expression by methylating histones as well as non-histone proteins. In this study we have examined the role of the LdSET1 protein. We find that while the set1 gene is not essential to cell survival, deletion of the set1 gene makes the parasite highly tolerant to the effects of an oxidizing environment, signifying that the LdSET1 protein plays a role in regulating the cells response to oxidative stress. LdSET1 appears to moderate the parasites survival in the oxidative intracellular milieu of host macrophages, controlling it such that infection is firmly established, and persistent, without wiping out the population of host cells it needs for survival.