Testing the impact of a single nucleotide polymorphism in a Plasmodium berghei ApiAP2 transcription factor on experimental cerebral malaria in mice

Abstract

Cerebral malaria (CM) is the deadliest form of severe Plasmodium infections. Currently, we have limited understanding of the mechanisms by which Plasmodium parasites induce CM. The mouse model of CM, experimental CM (ECM), induced by infection with the rodent parasite, Plasmodium berghei AnKA (PbANKA) has been extensively used to study the pathophysiology of CM. Recent genomic analyses revealed that the coding regions of PbAnKA and the closely related Plasmodium berghei NK65 (PbNK65), that does not cause ECM, differ in only 21 single nucleotide polymorphysims (SNPs). Thus, the SNP-containing genes might contribute to the pathogenesis of ECM. Although the majority of these SNPs are located in genes of unknown function, one SNP is located in the DNA binding site of a member of the Plasmodium ApiAP2 transcription factor family, that we recently showed functions as a virulence factor alternating the host's immune response to the parasite. Here, we investigated the impact of this SNP on the development of ECM. Our results using CRISPR-Cas9 engineered parasites indicate that despite its immune modulatory function, the SNP is neither necessary nor sufficient to induce ECM and thus cannot account for parasite strain-specific differences in ECM phenotypes. Malaria is a global health problem accounting for over 200 million cases each year worldwide and nearly 450,000 deaths in Africa alone, the majority of which are young children 1,2. Malaria is caused by mosquito-borne parasites belonging to genus Plasmodium that have complex life cycles involving two hosts 3. The outcome of the Plasmodium infection can vary from asymptomatic infections, to mild febrile disease, to severe malaria, the most deadly form of which is cerebral malaria (CM) 3-7. Approximately 1-2% of children with malaria develop CM that is

CCR6 expression reduces mouse survival upon malarial challenge with Plasmodium berghei NK65 strain

“…(13,14) Of note, the PbNK65 virulence was recently associated with a single polymorphism identified in the DNA binding domain of the parasitic APiAP2 transcription factor. The presence of a nucleotide (cytosine) at the position 5468 of the APiAP2 sequence would trigger a lethal iRBC infection with high parasitemia, (21) as observed in our data (Fig. 4A-B).…”

“…Parasitic infection -Mice were infected with either sporozoites or blood-stage PbNK65 strain parasites derived from the Department of Parasitology at the New York University (USA). (19,20,21) For the liver-stage infections, sporozoite challenges were performed through two distinct methods: (a) intravenous (iv) injection of parasites obtained from salivary glands of infected Anopheles stephensi mosquitoes; (b) natural infection by mosquito bites. Eighteen days before the mouse infection, reared mosquitoes were infected with Plasmodium berghei NK65 upon a C57BL/6 mouse blood meal in the insectary from the Department of Parasitology at the New York University, USA) as previously described.…”

Comparative transcriptomic analysis reveals translationally relevant processes in mouse models of malaria

“…In contrast to the P. yoelii strains, there are many SNPs that distinguish P. berghei ANKA from P. berghei NK65 ( Akkaya et al, 2020 ). Four SNPs (in genes PbANKA_1331700, PbANKA_0515200.1, PbANKA_1414600, and PbANKA_1222100.1) were examined in IGV using the P. berghei ANKA genome as a reference, confirming the absence of mismatch in the P. berghei ANKA infection samples and the presence of the expected mismatches in the P. berghei NK65 infection samples ( Supplementary file 1H ).…”

Disease modeling and stem cell immunoengineering in regenerative medicine using CRISPR/Cas9 systems

“…CRISPR/Cas9 systems have been routinely used to generate single-nucleotide polymorphism (SNP) animal models of human disease in rodents [169] , [175] , [176] . Such models provide a functional insight into human genetics that allow development of potential new therapies.…”