Role for the flagellum attachment zone in Leishmania anterior cell tip morphogenesis

Abstract

The shape and form of the flagellated eukaryotic parasite Leishmania is sculpted to its ecological niches and needs to be transmitted to each generation with great fidelity. The shape of the Leishmania cell is defined by the sub-pellicular microtubule array and the positioning of the nucleus, kinetoplast and the flagellum within this array. The flagellum emerges from the anterior end of the cell body through an invagination of the cell body membrane called the flagellar pocket. Within the flagellar pocket the flagellum is laterally attached to the side of the flagellar pocket by a cytoskeletal structure called the flagellum attachment zone (FAZ). During the cell cycle single copy organelles duplicate with a new flagellum assembling alongside the old flagellum. These are then segregated between the two daughter cells by cytokinesis, which initiates at the anterior cell tip. Here, we have investigated the role of the FAZ in the morphogenesis of the anterior cell tip. We have deleted the FAZ filament protein, FAZ2 and investigated its function using light and electron microscopy and infection studies. The loss of FAZ2 caused a disruption to the membrane organisation at the anterior cell tip, resulting in cells that were connected to each other by a membranous bridge structure between their flagella. Moreover, the FAZ2 null mutant was unable to develop and proliferate in sand flies and had a reduced parasite burden in mice. Our study provides a deeper understanding of membrane-cytoskeletal interactions that define the shape and form of an individual cell and the remodelling of that form during cell division.

The kinesin of the flagellum attachment zone in Leishmania is required for cell morphogenesis, cell division and virulence in the mammalian host

“…Since the FAZ has been shown to be involved in cell morphogenesis [15,16], we further analysed the cell morphology in promastigote cells. Microscopic observation of the FAZ7B null mutant revealed a greater mean flagellum length than that of the parental and add-back cell lines: 17.2±2.8 μm vs. 11.5±2.4 μm and 13.1±2.3 μm for the FAZ7B null mutant, parental and add-back cell lines, respectively (Fig 4B).…”

“…We then examined the respective localization of FAZ7B and representative components of the FAZ. FAZ7B localized in close proximity with FAZ1, FAZ2, FAZ8, as well as FAZ5 (Fig 1B and 1C, S2B and S2C Fig) . FAZ1, FAZ2, and FAZ8 label the cell body side of the FAZ, while FAZ5 is a component of the 'cell body membrane domain' of the FAZ [4,15,16]. In z-stack images, the FAZ7B signal appeared to be slightly proximal and around or on one side of that of FAZ1 (S2 Movie) which formed a 'horse-shoe' around the flagellar pocket.…”

“…Cell division involves first the duplication and elongation of the flagellum from a mature pro-basal body within a single FP, followed by the duplication of the latter (1N1K2F). Subsequently, nuclear mitosis and kinetoplast division occur, near synchronously in L. mexicana promastigotes, generating 2N2K2F cells [16,25]. FAZ7B null mutant cells were stained with Hoechst and N/K patterns were determined in the promastigote and axenic amastigote stages.…”

Leishmania in 360: Guidelines for Exosomal Research

“…In other way, studies in vitro differentiation of Leishmania (Leishmania) amazonensis amastigotes into promastigotes has been detailed by electron microscopy, gaining a greater knowledge of these important parasite mechanisms [44]. Currently, for high care and caution when defining cell types solely based on cell morphology, exist few molecular markers to identify life cycle forms [36-39-45-46].…”

“…In addition, studies reveal differences in the intramembranous particles (integral proteins) distribution, where particle aggregation on specialized regions of the Leishmania membrane reflect their composition, function, density, also according to the parasitic evolutionary form: promastigote membrane richer in whole proteins than the amastigote membrane [35-40-42-53-118]. The FP is an invagination of the cell membrane at the base of the flagellum, and the flagellum membrane have a smaller amount of intramembranous particles when compared to the parasite body [42][43][44][45]. However, in the region of the membrane around the FP is possible to find specialized domains with a high concentration of these proteins, forming zones of flagellar adhesion [42-45-119].…”