Functional characterization of both MAP kinases of the human malaria parasite Plasmodium falciparum by reverse genetics

Abstract

SummaryThe kinome of the human malaria parasite Plasmodium falciparum includes two genes encoding mitogen-activated protein kinase (MAPK) homologues, pfmap-1 and pfmap-2, but no clear orthologue of the MAPK kinase (MAPKK) family, raising the question of the mode of activation and function of the plasmodial MAPKs. Functional studies in the rodent malaria model Plasmodium berghei recently showed the map-2 gene to be dispensable for asexual growth and gametocytogenesis, but essential for male gametogenesis in the mosquito vector. Here, we demonstrate by using a reverse genetics approach that the map-2 gene is essential for completion of the asexual cycle of P. falciparum, an unexpected result in view of the non-essentiality of the orthologous gene for P. berghei erythrocytic schizogony. This validates Pfmap-2 as a potential target for chemotherapeutic intervention. In contrast, the other P. falciparum MAPK, Pfmap-1, is required neither for in vitro schizogony and gametocytogenesis in erythrocytes, nor for gametogenesis and sporogony in the mosquito vector. However, Pfmap-2 protein levels are elevated in pfmap-1 -parasites, suggesting that Pfmap-1 fulfils an important function in asexual parasites that necessitates compensatory adaptation in parasites lacking this enzyme.

Preerythrocytic, live-attenuated Plasmodium falciparum vaccine candidates by design

“…The P. falciparum p52 Ϫ and p36 Ϫ phenotypes are therefore similar to the phenotypes observed for the corresponding rodent malaria knockouts (11,12). Such experimental concurrence of phenotypes between rodent malaria parasite mutants and P. falciparum mutants is important because it has been observed that knockout of orthologous genes in rodent and human malaria parasites can result in distinct phenotypes (33,34).…”

Global kinomic and phospho-proteomic analyses of the human malaria parasite Plasmodium falciparum

“…Among these 22 ePKs are 8 enzymes that we previously reported as playing a crucial role in asexual proliferation: Pfmap-2 (PF11_0147) 9 , Pfnek-1 (PFL1370w) 10 , PfCK2 (PF11_ 0096) 11 , PfTKL3 (PF13_0258) 12 , Pfcrk-3 (PFD0740w) 13 , PfCLK1 (PF14_0431) 14 , PfCLK2 (PF14_0408) 14 and PfARK1 (PF13_0258) 15 . Others have provided reverse genetics evidence for essentiality of three additional ePKs, PfCDPK1 (PFB0815w) 16 , PfPKG (PF14_ 0346) 17 and PfPCDPK5 (PF13_0211) 18 .…”

An evolutionary perspective on the kinome of malaria parasites

“…Typically, this signal transduction pathway is formed by three or four protein kinases associated with distinct ePK families: MAP4K or MEKKK (a tyrosine-like kinase such as Raf, or a member of the STE20 family), MAP3K or MEKK (a member of the STE11 family), MAPKK or MEK (a STE7 member) and MAPK family (best characterized in the ERK1/ERK2, p38 and JNK subfamilies) [84]. However, in Plasmodium and most other apicomplexans, the STE group of kinases is reduced to one or two distantly related members, or even missing entirely [85], although two clear (but atypical) members of the MAPK family are present [86]. One of these, Pfmap-1, has been demonstrated to be phosphorylated on the conserved TXY motif in its activation loop [20], while this otherwise highly conserved activation motif is substituted in the other MAPK (Pfmap-2) by TSH [44]; no evidence for in vivo phosphorylation of this TSH motif is available, but mutating either the Thr or the His (but not the Ser) dramatically affects kinase activity of the recombinant enzyme [44].…”