A spatiotemporally resolved single cell atlas of the Plasmodium liver stage

Abstract

Malaria infection involves an obligatory, yet clinically silent liver stage1,2. Hepatocytes operate in repeating units termed lobules, exhibiting heterogeneous gene expression patterns along the lobule axis3, but the effects of hepatocyte zonation on parasite development have not been molecularly explored. Here, we combine single-cell RNA sequencing4 and single-molecule transcript imaging5 to characterize the host’s and parasite’s temporal expression programs in a zonally-controlled manner for the rodent malaria parasite Plasmodium berghei ANKA. We identify differences in parasite gene expression in distinct zones, and a sub-population of periportally-biased hepatocytes that harbor abortive infections associated with parasitophorous vacuole breakdown. These ‘abortive hepatocytes’ up-regulate immune recruitment and key signaling programs. They exhibit reduced levels of Plasmodium transcripts, perturbed parasite mRNA localization, and may give rise to progressively lower abundance of periportal infections. Our study provides a resource for understanding the liver stage of Plasmodium infection at high spatial resolution and highlights heterogeneous behavior of both the parasite and the host hepatocyte.

Single-cell RNA profiling of Plasmodium vivax-infected hepatocytes reveals parasite- and host- specific transcriptomic signatures and therapeutic targets

“…Additionally, they have been shown to be susceptible to treatment with primaquine in combination with chloroquine and ionophores, implying they were alive to begin with (Maher et al, 2021). At the molecular level, we did not find consistencies between our data and the transcriptomic signatures of “abortive” P. berghei liver forms reported by Afriat and colleagues (2021). For example, orthologous expression of genes encoding for HSP90, HSP70, and HOP—three markers defining abortive parasites in their dataset—displayed greater expression in the schizont cluster relative to hypnozoites.…”

“…For each scRNA-seq library processed, we dedicated cultures from the same infection to provide a visual snapshot of the parasite viability at each end point. While Afriat and colleagues (2021) define abortive forms as displaying “vacuole breakdown,” upon inspection of our cultures, we did not find indications of abortive or dead parasites. Moreover, the parasites morphologically identified as hypnozoites were highly similar in morphology to those described in humanized mice (Mikolajczak et al, 2015) and in vitro (Roth, Maher et al, 2018).…”

Plasmodium berghei-Mediated NRF2 Activation in Infected Hepatocytes Enhances Parasite Survival

“…Target genes are involved in several cytoprotective pathways including detoxification response, heme metabolism, cell proliferation, and apoptotic resistance/survival regulation ( [8][9][10]; [11,12]). Importantly, several NRF2 target genes have recently been shown by single cell mRNA seq analysis to be upregulated upon P. berghei infection of hepatocytes [13]. Another transcriptome study in P. vivax-infected hepatocytes also identified NRF2 regulated expression as a pathway supporting parasite persistence suggesting an important and conserved function of this host cell transcription factor [14].…”

“…Among these are proteins involved in glutathione and iron metabolism such as glutathione S-transferases (Gstm1 and Gstm3) and Ftl1, Fth, and Slc40a1, respectively. In addition, p62 expression that is also controlled by Nrf2 was found to be upregulated in infected host cells [13]. Another recent transcriptome analysis on cells infected with Plasmodium vivax, a human infecting Plasmodium species, found enrichment of transcripts governed by NRF2 upon infection of primary human hepatocytes.…”

Systems biology of malaria explored with nonhuman primates

“…This type of research comes with many challenges, given the need to distinguish quiescent infected cells that have low metabolic activity from developing or newly activated forms [ 217 , 218 ]. Today, such culture systems, single-cell -omic technologies [ 219 ], and cellular imaging advances [ 220 , 221 ] are paving the way for using these model systems to understand P. vivax sporozoites [ 222 ] and host–parasite interactions in infected hepatocytes, including dormant and activated hypnozoites [ 223 ].…”