Structure and function of the Trypanosoma brucei bilobe

Trypanosoma brucei is a unicellular protist and a major parasite of mammals. In humans, it is the causative agent of the neglected tropical disease human African trypanosomiasis (also known as Sleeping Sickness), while in livestock it is responsible for nagana. The cytoskeleton of T. brucei is essential both to its viability and to its pathogenesis, but is relatively poorly understood. The proposal here seeks to deepen our understanding of the cytoskeleton of this organism, with a particular focus on a novel structural component known as the bilobe. The bilobe was discovered in 2005, and despite concerted efforts its primary cellular function remains unclear. The focus of the work is TbMORN1, a protein exclusively localised to the bilobe and whose loss from the mammalian-infective bloodstream form of T. brucei is rapidly lethal. The project work outlined here will capitalise on a recent and highly successful screen for novel protein components of the bilobe to provide a full structural, functional, and cell biological appraisal of this object. The proposed experiments feature a combination of molecular cell biology, state-of-the-art biochemical techniques, and structural biology.

This project provided the first high-resolution structures of a ubiquitous family of proteins, and led to new insights into their functions within cells. Proteins are the engines of the cell, responsible for structural, catalytic, and dynamic functions. The polypeptide chains that comprise proteins fold into structured domains, but an alternative mode of assembly is to use repeat motifs. Repeat motifs are short stretches of amino acids that can be arranged in tandem in order to generate complex higher-order structures from simple building blocks. The MORN family of protein repeats is relatively unstudied, and the work in this project led to the first high-resolution structures of MORN repeat proteins. Each MORN repeat was found to form a single hairpin structure, and when assembled side-to-side this produces a twisting fibre with a large surface exposed for interactions. The function of MORN repeats has long been unclear, and by studying a protein composed solely of MORN repeats (TbMORN1 from the parasite Trypanosoma brucei, which causes sleeping sickness), new insights into its cellular functions were obtained. TbMORN1 is essential for the viability of the mammalian-infective form of T. brucei, and appears to regulate the transit of large macromolecular cargo into the cell. Despite MORN repeats being long regarded as membrane-binding modules, no evidence was obtained for an interaction of TbMORN1 with membranes either in vivo or in vitro.