Biogenesis of extracellular vesicles in protozoan parasites: The ESCRT complex in the trafficking fast lane?

Extracellular vesicles (EVs) provide a central mechanism of cell-cell communication. While EVs are found in most organisms, their pathogenesis-promoting roles in parasites are of particular interest given the potential for medical insight and consequential therapeutic intervention. Yet, a key feature of EVs in human parasitic protozoa remains elusive: their mechanisms of biogenesis. Here, we survey the current knowledge on the biogenesis pathways of EVs secreted by the four main clades of human parasitic protozoa: apicomplexans, trypanosomatids, flagellates, and amoebae. In particular, we shine a light on findings pertaining to the Endosomal Sorting Complex Required for Transport (ESCRT) machinery, as in mammals it plays important roles in EV biogenesis. This review highlights the diversity in EV biogenesis in protozoa, as well as the related involvement of the ESCRT system in these unique organisms.

High-throughput analysis of the transcriptional patterns of sexual genes in malaria.

BACKGROUND: Plasmodium falciparum (Pf) is the leading protozoan causing malaria, the most devastating parasitic disease. To ensure transmission, a small subset of Pf parasites differentiate into the sexual forms (gametocytes). Since the abundance of these essential parasitic forms is extremely low within the human host, little is currently known about the molecular regulation of their sexual differentiation, highlighting the need to develop tools to investigate Pf gene expression during this fundamental mechanism. METHODS: We developed a high-throughput quantitative Reverse-Transcription PCR (RT-qPCR) platform to robustly monitor Pf transcriptional patterns, in particular, systematically profiling the transcriptional pattern of a large panel of gametocyte-related genes (GRG). Initially, we evaluated the technical performance of the systematic RT-qPCR platform to ensure it complies with the accepted quality standards for: (i) RNA extraction, (ii) cDNA synthesis and (iii) evaluation of gene expression through RT-qPCR. We then used this approach to monitor alterations in gene expression of a panel of GRG upon treatment with gametocytogenesis regulators. RESULTS: We thoroughly elucidated GRG expression profiles under treatment with the antimalarial drug dihydroartemisinin (DHA) or the metabolite choline over the course of a Pf blood cycle (48 h). We demonstrate that both significantly alter the expression pattern of PfAP2-G, the gametocytogenesis master regulator. However, they also markedly modify the developmental rate of the parasites and thus might bias the mRNA expression. Additionally, we screened the effect of the metabolites lactate and kynurenic acid, abundant in severe malaria, as potential regulators of gametocytogenesis. CONCLUSIONS: Our data demonstrate that the high-throughput RT-qPCR method enables studying the immediate transcriptional response initiating gametocytogenesis of the parasites from a very low volume of malaria-infected RBC samples. The obtained data expand the current knowledge of the initial alterations in mRNA profiles of GRG upon treatment with reported regulators. In addition, using this method emphasizes that asexual parasite stage composition is a crucial element that must be considered when interpreting changes in GRG expression by RT-qPCR, specifically when screening for novel compounds that could regulate Pf sexual differentiation.

Malaria parasites both repress host CXCL10 and use it as a cue for growth acceleration.

Pathogens are thought to use host molecular cues to control when to initiate life-cycle transitions, but these signals are mostly unknown, particularly for the parasitic disease malaria caused by Plasmodium falciparum. The chemokine CXCL10 is present at high levels in fatal cases of cerebral malaria patients, but is reduced in patients who survive and do not have complications. Here we show a Pf 'decision-sensing-system' controlled by CXCL10 concentration. High CXCL10 expression prompts P. falciparum to initiate a survival strategy via growth acceleration. Remarkably, P. falciparum inhibits CXCL10 synthesis in monocytes by disrupting the association of host ribosomes with CXCL10 transcripts. The underlying inhibition cascade involves RNA cargo delivery into monocytes that triggers RIG-I, which leads to HUR1 binding to an AU-rich domain of the CXCL10 3'UTR. These data indicate that when the parasite can no longer keep CXCL10 at low levels, it can exploit the chemokine as a cue to shift tactics and escape.