Comparative analysis of the kinomes of Plasmodium falciparum, Plasmodium vivax and their host Homo sapiens

Abstract

Background: Novel antimalarials should be effective across all species of malaria parasites that infect humans, especially the two species that bear the most impact, Plasmodium falciparum and Plasmodium vivax. Protein kinases encoded by pathogens, as well as host kinases required for survival of intracellular pathogens, carry considerable potential as targets for antimalarial intervention (Adderley et al. Trends Parasitol 37:508-524, 2021; Wei et al. Cell Rep Med 2:100423, 2021). To date, no comprehensive P. vivax kinome assembly has been conducted; and the P. falciparum kinome, first assembled in 2004, requires an update. The present study, aimed to fill these gaps, utilises a recently published structurally-validated multiple sequence alignment (MSA) of the human kinome (Modi et al. Sci Rep 9:19790, 2019). This MSA is used as a scaffold to assist the alignment of all protein kinase sequences from P. falciparum and P. vivax, and (where possible) their assignment to specific kinase groups/families.

Results: We were able to assign six P. falciparum previously classified as OPK or 'orphans' (i.e. with no clear phylogenetic relation to any of the established ePK groups) to one of the aforementioned ePK groups. Direct phylogenetic comparison established that despite an overall high level of similarity between the P. falciparum and P. vivax kinomes, which will help in selecting targets for intervention, there are differences that may underlie the biological specificities of these species. Furthermore, we highlight a number of Plasmodium kinases that have a surprisingly high level of similarity with their human counterparts and therefore not well suited as targets for drug discovery.

Conclusions: Direct comparison of the kinomes of Homo sapiens, P. falciparum and P. vivax sheds additional light on the previously documented divergence of many P. falciparum and P. vivax kinases from those of their human host. We provide the first direct kinome comparison between the phylogenetically distinct species of P. falciparum and P. vivax, illustrating the key similarities and differences which must be considered in the context of kinase-directed antimalarial drug discovery, and discuss the divergences and similarities between the human and Plasmodium kinomes to inform future searches for selective antimalarial intervention.

Keywords: Kinase; Kinome; Malaria; Plasmodium falciparum; Plasmodium vivax.

Fig. 1

Sequence logos of the conserved regions in the multisequence alignment for the kinase domains of P. falciparum and P. vivax. Aligned regions of the kinase domain defined by [38] and logo generated using the webserver WebLogo (https://weblogo.berkeley.edu/) [39] and edited in Adobe Illustrator

Fig. 2

Phylogenetic tree containing the protein kinome of Homo sapiens, Plasmodium falciparum, Plasmodium vivax. The tree is represented in a circular format and contains a total of 671 protein kinases sequences excluding the atypical kinases (H. sapiens − 497, P. falciparum − 98 and P. vivax – 78). P. falciparum sequences were accessed from [8], H. sapiens sequences were accessed from [16] and P. vivax sequences were identified from PlasmoDB [17] and initially aligned using ClustalOmega [23]. All of the kinase sequences were imported into Jalview [24]. Using the human kinases as a template. the Plasmodium kinases were aligned into the conserved regions as defined by [16]. The resultant 230 column alignment was assessed using RAxML to infer phylogenetic distances and determine bootstrapping [41]. RAxML Gui 2.0 [42] was used with the following parameters: maximum-likelihood rapid bootstrap with the PROTgamma substitution model LG4M, with AutoMRE. A gene tree was inferred through the RAxML analysis using the ‘best tree’ and rendered with the interactive tree of life webserver (iTOL) [44]. HMMER profiling was performed for P. falciparum and P. vivax kinases using the defined families of Kinomer [25] with the addition of the NEK family (see Supplementary data 3). Using the tree structure, HMMER results and the known human kinase family assignments the Plasmodium kinases were assigned to the 9 typical protein kinase groups. These family assignments were annotated using Adobe illustrator along with the Aurora kinase family (ARK) and the Apicomplexan-specific kinase family FIKK. Orphan, or ‘other’ kinases are largely unassigned to families (white background). Bootstraps values above 50 are represented as circles on the associated branches, larger circles indicate higher bootstrap values. Plasmodium kinases are indicated with a red star and, the associated branches are bold

Fig. 3

Visualisation of the protein kinase family membership across Homo sapiens, Plasmodium falciparum, Plasmodium vivax. The nine typical protein kinase families along with the Aurora kinase family (ARK) and the Apicomplexan-specific family of FIKK were included here. The remaining unassigned kinases are denoted as Orphans. Each group/family is represented as a percentage of the total protein kinome for each organism. Note atypical protein kinases are not included in this analysis. The number next to each bar indicates the number of kinases which belong to each of the respective families for each organism. Blue = Plasmodium vivax, Red = Plasmodium falciparum and Grey = Homo sapiens

Fig. 4

Truncated phylogenetic tree of the CMGC group, focused on branches with strong bootstrap support between Plasmodium and human sequences. A small number of Plasmodium kinases within the CMGC group exhibited homology to human kinases. These kinases were Plasmodium kinases PfCLK3, PfSRPK1, PfMAPK1, PfCKα and PfGSK3 (denoted with a red circle). The associated strong bootstrap support (> 80) has been coloured blue, along with the branches to the human homologs. Bootstrap values are listed on the associated branches (when > 30), P. falciparum kinases are highlighted in blue, P. vivax kinases are highlighted in red

Fig. 5

Comparative Phylogenetic tree of Plasmodium vivax and Plasmodium falciparum. Phylogenetic tree indicating the kinases shared and unique to each of the two species, P. falciparum (red branches), P. vivax (black branches). Red arrow indicates the kinases PVP01_0118800 and PF3D7_0610600 (CDPK2) which do not have an equivalent kinase in the other species. Blue arrow indicates PVP01_0114800, the only member of the FIKK family that P. vivax encodes. See Fig. 1 legend for details regarding the assembly and construction of the phylogenetic tree. Typical protein kinase families were annotated using Adobe illustrator along with the Aurora kinase family (ARK) and the Apicomplexan-specific kinase family FIKK. Orphan, or ‘other’ kinases are largely unassigned to families (white background). Bootstraps values above 50 are represented as circles on the associated branches, larger circles indicate higher bootstrap values, note these values relate to the full tree illustrated in Fig. 2

Fig. 6

Phylogenetic tree of Plasmodium spp. illustrating CDPK2 and PVP01_118800 orthologs. Tree assembled using the mitochondrial genomes of each species (see methods for details). A gene tree was inferred through the RAxML analysis using the ‘best tree’ and rendered with the tree of life webserver (iTOL) [44]. CDPK2 and PVP01_118800 orthologs were identified through blast searches using the kinase domains. Plasmodium species which have orthologs to CDPK2 are indicated by the blue circle, while species with an ortholog of PVP01_118800 are indicated by the red circle. Bootstrap support for the gene tree is indicated on each of the branches, only values above 40 are displayed