Motility and more: the flagellum of Trypanosoma brucei

Abstract

Trypanosoma brucei is a pathogenic unicellular eukaryote that infects humans and other mammals in sub-Saharan Africa. A central feature of trypanosome biology is the single flagellum of the parasite, which is an essential and multifunctional organelle that facilitates cell propulsion, controls cell morphogenesis and directs cytokinesis. Moreover, the flagellar membrane is a specialized subdomain of the cell surface that mediates attachment to host tissues and harbours multiple virulence factors. In this Review, we discuss the structure, assembly and function of the trypanosome flagellum, including canonical roles in cell motility as well as novel and emerging roles in cell morphogenesis and host-parasite interactions.

Figure 1. T. brucei flagellum overview

Bottom shows cartoon diagram of a T. brucei cell with flagellum in black. Cell movement (arrow) is with the flagellum tip leading. Anterior (A) and posterior (P) of the cell are labeled. Top shows a cartoon diagram of flagellum emergence from the flagellar pocket (FP) at the posterior end of the cell (boxed region in bottom panel). The flagellum is built around a core of microtubules that are arranged in characteristic patterns, as shown above the corresponding cross sections. The flagellar axoneme (AX) emanates from the basal body (BB) via the transition zone (TZ) and is laterally connected to the cell membrane (CM) via the flagellum attachment zone (FAZ). Extra-axonemal structures inside the flagellar membrane (FM) include the paraflagellar rod (PFR) and the ciliary necklace (CN). A tripartite attachment complex (TAC) links the basal body to the kinetoplast (KP) and transition fibers (TF) connect the basal body to the flagellar pocket. The distinctive architecture of the trypanosome flagellum is dictated by specialized membrane/cytoskeletal features, such as the flagellar pocket collar (FPC) and the FAZ filament (FAZF). Adjacent to the FAZ filament are the subpellicular microtubules (SPM), microtubule quartet (MTQ) and FAZ ER (FER). Numbered magenta lines indicate specific subdomains of the parasite cell surface, corresponding to the cell body membrane (1), flagellum (2) and cytoplasmic (3) sides of the flagellar pocket membrane (FPM), FAZ-associated flagellum (4) and cell body (5) membranes, the flagellum tip (6), and the remainder of the flagellum membrane (7)

Figure 2. Axoneme and PFR structure

(Bottom) Cartoon showing transverse section detail of axoneme and PFR, as viewed looking posterior to anterior. (Top) Cartoon showing longitudinal view of the axonemal repeating unit on one outer doublet microtubule, oriented with the basal body end toward the left. Prominent axonemal structures are labeled and include outer doublet (OD) and central pair microtubules, outer arm dyneins (OAD) and inner arm dyneins (IAD), radial spokes (RS) and the nexin-dynein regulatory complex (NDRC). Outer doublet microtubules are numbered 1 – 9, as indicated. Proximal (P), intermediate (I) and distal (D) domains of the PFR are shown.

Figure 3. The flagellum in cell division and cell morphogenesis

(A) Cartoon diagram showing the relationship of the flagellum and flagellum-associated structures to key steps of the cell division cycle in procyclic form trypomastigotes. (i) In G1 the cell has a single flagellum, FAZ , basal body and nucleus. (ii) The first recognizable event in cell division is formation of a new basal body, which moves posterior to the old basal body and elongates to form the new flagellum. The tip of the new flagellum is attached to the old flagellum by the flagella connector. (iii) The new flagellum elongates in parallel with a new FAZ and is guided by attachment to the old flagellum via the flagella connector. (iv) Elongation of the flagellum and FAZ continue until the flagella connector reaches a specific stop point, approximately 0.6 lengths of the old flagellum, at which point the two basal bodies separate fully. (v) Mitosis ensues and the new nucleus moves to lie between the new and old basal body. (v - vi) Cytokinesis initiates with cleavage furrow ingression beginning at the tip of the new flagellum and FAZ (red arrow) and progressing anterior to posterior along a line between the two flagella. Strikingly, cleavage furrow ingression does not depend on an actomyosin contractile ring, which governs membrane abscission in other eukaryotic cells. (vii) Ultimately, daughter cells are connected at their posterior ends and oriented in opposite directions, with their flagella exerting rotational and pulling forces that facilitate final cell separation to yield two G1 cells^, . Events for division of bloodstream form cells are essentially the same, except that there is no flagella connector and both nuclei remain anterior to both kinetoplasts. (B) Major morphological variants that occur during the developmental cycle within the tsetse are shown. Restructuring and repositioning of the flagellum is a prominent feature of cellular differentiation events that give rise to these developmental forms. (i) A mesocyclic trypomastigote is generated from a procyclic cell through elongation of the flagellum and cell body. (ii) Long epimastigotes are formed via repositioning of the nucleus and basal body. (iii – iv) An asymmetric division gives rise to one long and one short epimastigote. The short epimastigote arises via cleavage furrow initiation at the tip of a shortened flagellum and FAZ (arrow in iii). (v) Metacyclic trypomastigotes are formed through an asymmetric division that again repositions the nucleus and basal body.

Figure 4. The flagellum as a platform for host-pathogen interactions

Boxes highlight flagellum features that contribute to host-parasite interaction. In the mammalian host, receptors on the trypanosome flagellar pocket mediate uptake of host factors that are essential for parasite survival, such as transferrin (Tf), as well as those that dictate host-range specificity, such as trypanosome lytic factor (TLF). Several flagellar proteins contribute to virulence in the mammalian host. Loss of flagellar proteins GPI-PLC, calflagins (CFs)or metacaspase 4 (MCA4)reduces parasite virulence and prolongs mouse survival through unknown mechanisms. Flagellar adenylate cyclase (ESAG4) interferes with the host’s early innate immune response to promote parasite virulence. Flagellum-dependent motility facilitates immune evasion through clearance of host immunoglobulin from the parasite surface. In the tsetse fly, the flagellum-mediated attachment to the salivary gland epithelium enables parasite persistence and marks the final stage of differentiation into mammalian infectious forms.