Vital role forPlasmodium bergheiKinesin8B in axoneme assembly during male gamete formation and mosquito transmission

Abstract

Sexual development is an essential phase in the Plasmodium life cycle, where male gametogenesis is an unusual and extraordinarily rapid process. It produces 8 haploid motile microgametes, from a microgametocyte within 15 minutes. Its unique achievement lies in linking the assembly of 8 axonemes in the cytoplasm to the three rounds of intranuclear genome replication, forming motile microgametes, which are expelled in a process called exflagellation. Surprisingly little is known about the actors involved in these processes. We are interested in kinesins, molecular motors that could play potential roles in male gametogenesis. We have undertaken a functional characterization in Plasmodium berghei of kinesin-8B (PbKIN8B) expressed specifically in male gametocytes and gametes. By generating Pbkin8B-gfp parasites, we show that PbKIN8B is specifically expressed during male gametogenesis and is associated with the axoneme. We created a ΔPbkin8B knockout cell line and analysed the consequences of the absence of PbKIN8B on male gametogenesis. We show that the ability to produce sexually differentiated gametocytes is not affected in ΔPbkin8B parasites and that the 3 rounds of genome replication occur normally. Nevertheless, the development to free motile microgametes is halted and the life cycle is interrupted in vivo. Ultrastructural analysis revealed that intranuclear mitoses are unaffected whereas cytoplasmic microtubules, although assembled in doublets and elongated, fail to assemble in the normal axonemal '9+2' structure and become motile. Absence of a functional axoneme prevented microgamete assembly and release from the microgametocyte, severely reducing infection of the mosquito vector. This is the first functional study of a kinesin involved in male gametogenesis.These results reveal a previously unknown role for PbKIN8B in male gametogenesis, providing new insights into Plasmodium flagellar formation.

Genome-wide functional analysis reveals key roles for kinesins in the mammalian and mosquito stages of the malaria parasite life cycle

“…Δ kinesin-8B parasites produced no male gametes as shown previously (Depoix et al, 2020;Zeeshan et al, 2019a), while there was a significant decrease in male gamete formation in Δ kinesin-15 parasites (Fig. 4B).…”

A conserved malaria parasite antigen Pb22 plays a critical role in male gametogenesis inPlasmodium berghei

“…The mitogen‐activated protein kinase 2 (MAP‐2) affects the final stages of male gamete formation in both P. berghei and P. falciparum (Hitz, Balestra, Brochet, & Voss, 2020; Tewari, Dorin, Moon, Doerig, & Billker, 2005). Recent studies have revealed the participation of some structural and motor proteins in axoneme assembly (Depoix et al, 2020; Marques et al, 2015). Later in male gametogenesis, the egress process requires the release of lytic factors (e.g.…”

Mechanochemical tuning of a kinesin motor essential for malaria parasite transmission

“…Kinesin-8s are among the most widely distributed kinesin subfamilies across eukaryotes 8 , perhaps because of their functional adaptability to both move processively along MTs and to influence MT dynamics. To understand the molecular basis of P. berghei kinesin-8B function in parasite transmission 6,7 , we characterised its motor domain and compared it with kinesin-8B from P. falciparum . Our biochemical and structural data provide evidence of conserved and precisely tuned mechanochemistry in these motors, which is distinct compared to other kinesin-8s characterised to date, including those in the parasite’s mammalian hosts.…”

“…A conserved role for catalytic depolymerases in male gamete formation Plasmodium kinesin-8Bs are expressed exclusively in male gametocytes, the only flagellated stage of the parasite life cycle. While deletion of the P. berghei kinesin-8B has no effect on parasite blood stages 42 and also has no effect on genome duplication during male gametogenesis, kinesin-8B knockout parasites exhibit disrupted flagella assembly and parasite transmission is blocked 6,7 . The motor domains from P. berghei and P. falciparum kinesin-8Bs exhibit very similar properties in vitro which, together with their very similar expression profiles 43 , suggest a conserved function for kinesin-8Bs across Plasmodium species.…”

“…Kinesin-8B - a member of the kinesin superfamily of ATP-driven, MT-based molecular motors 5 - is required for flagella formation in P. berghei male gametes, and its knockout completely disrupts parasite transmission 6,7 . Kinesin-8s are one of the fifteen kinesin families identified across apicomplexa 8 that, together with kinesin-13s, are important regulators of MT dynamics 9 .…”