Single-cell RNA-seq reveals hidden transcriptional variation in malaria parasites

Abstract

Single-cell RNA-sequencing is revolutionising our understanding of seemingly homogeneous cell populations but has not yet been widely applied to single-celled organisms. Transcriptional variation in unicellular malaria parasites from the Plasmodium genus is associated with critical phenotypes including red blood cell invasion and immune evasion, yet transcriptional variation at an individual parasite level has not been examined in depth. Here, we describe the adaptation of a single-cell RNA-sequencing (scRNA-seq) protocol to deconvolute transcriptional variation for more than 500 individual parasites of both rodent and human malaria comprising asexual and sexual life-cycle stages. We uncover previously hidden discrete transcriptional signatures during the pathogenic part of the life cycle, suggesting that expression over development is not as continuous as commonly thought. In transmission stages, we find novel, sex-specific roles for differential expression of contingency gene families that are usually associated with immune evasion and pathogenesis.

eLife digest

Malaria is a life-threatening disease that affects hundreds of millions of people every year and causes around 500,000 deaths, mostly among young children. The disease is caused by Plasmodium parasites, which have a complex life cycle that involves different stages in different hosts. During mosquito bites, the parasites can be transmitted to people where they spend part of their life cycle inside red blood cells. Inside these cells, they can multiply rapidly and eventually burst the blood cells, which causes some of the symptoms of the disease. The parasite also produces sexual stages, which can be passed on to the next mosquito that feeds on the host.

Scientists have been studying these different stages to better understand how the parasites manage to evade the human immune system so successfully. Most of the research has looked at how genes differ between large pools of parasites, but this approach hides important differences between individual parasites. Understanding variation and how individual parasites behave could help to develop new and effective drugs and vaccines for malaria.

Now, Reid et al. used a technique called single-cell RNA sequencing, which allowed them to hone in on individual genes within a single parasite. This revealed hidden patterns in the way the parasites use their genes across the life cycle. When the parasite is developing inside a red blood cell, distinct groups of genes turn on simultaneously and are later switched off together. Reid et al. found clues about the genes that might be controlling these groups. The experiments also showed that a set of genes previously thought to be involved solely in evading the immune system is also important for the transition from human to mosquito.

A next step will be to see if single-cell RNA sequencing technology could be used to reveal more about the basic biology of the parasite and how it resists drugs or evades the immune system. In the future, this may help to develop drugs that interfere with the synchronisation of these groups of genes to disrupt the parasite’s development and stop it from causing the disease. The genes involved in transmission between hosts could be another promising drug target, and one day, may help to eliminate the disease.