Unraveling the Ubiquitome of the Human Malaria Parasite

Abstract

Malaria is one of the deadliest infectious diseases worldwide. The most severe form is caused by the eukaryotic protozoan parasite Plasmodium falciparum. Recent studies have highlighted the importance of post-translational regulations for the parasite's progression throughout its life cycle, protein ubiquitylation being certainly one of the most abundant. The specificity of its components and the wide range of biological processes in which it is involved make the ubiquitylation pathway a promising source of suitable targets for anti-malarial drug development. Here, we combined immunofluorescent microscopy, biochemical assays, in silico prediction, and mass spectrometry analysis using the multidimensional protein identification technology, or MudPIT, to describe the P. falciparum ubiquitome. We found that ubiquitin conjugates are detected at every morphological stage of the parasite erythrocytic cycle. Furthermore, we detected that more than half of the parasite's proteome represents possible targets for ubiquitylation, especially proteins found to be present at the most replicative stage of the asexual cycle, the trophozoite stage. A large proportion of ubiquitin conjugates were also detected at the schizont stage, consistent with a cell activity slowdown to prepare for merozoite differentiation and invasion. Finally, for the first time in the human malaria parasite, our results strongly indicate the presence of heterologous mixed conjugations, SUMO/UB. This discovery suggests that sumoylated proteins may be regulated by ubiquitylation in P. falciparum. Altogether, our results present the first stepping stone toward a better understanding of ubiquitylation and its role(s) in the biology of the human malaria parasite.

The Ubiquitin Proteome of Toxoplasma gondii Reveals Roles for Protein Ubiquitination in Cell-Cycle Transitions

“…Ubiquitination sites were detected on S27A and three ubiquitin-like proteins on T. gondii -specific peptides. Unlike studies in P. falciparum (Ponts et al, 2011), ubiquitination of SUMO was not detected.…”

“…These yields are comparable to a ubiquitin survey of HEK293T cells (Xu et al, 2010). Seventy-six ubiquitinated proteins were identified in Plasmodium falciparum using a less sensitive, whole protein immunoprecipitation method (Ponts et al, 2011). We detected known substrates such as histones and ubiquitin, as well as atypical targets.…”

“…During intraerythrocytic development of P. falciparum ubiquitination is altered (Lasonder et al, 2015; Ponts et al, 2011), suggesting a role for ubiquitin in cell cycle regulation. Since a large proportion of the T. gondii transcriptome is cell cycle regulated (Behnke et al, 2010), we hypothesised that the ubiquitinome of T. gondii would be enriched for cell cycle-regulated proteins.…”

“…This is consistent with what is known about the IMC; during endodyogeny, the IMC is continuously rebuilt and dissembled (Ouologuem and Roos, 2014). The IMC is highly conserved in Apicomplexa (Kono et al, 2012), but ubiquitinated IMC proteins were not identified in P. falciparum (Ponts et al, 2011). Ubiquitination of Plasmodium IMC proteins could be substoichoimetric and under the detection limit of the approach used.…”

New Chemotherapeutic Strategies Against Malaria, Leishmaniasis and Trypanosomiases

“…Moreover, the temperature shift to which the parasite must rapidly adapt when it moves back and forth between humans and mosquitoes induces a stress response that is managed by the ubiquitin-proteasome system. In silico predictions indicate that over half of the parasite proteins represent targets for ubiquitination [83,84]. Therefore, proteasome inhibitors are considered promising antimalarial agents.…”