Loss of the BBSome perturbs endocytic trafficking and disrupts virulence of Trypanosoma brucei

Abstract

Cilia (eukaryotic flagella) are present in diverse eukaryotic lineages and have essential motility and sensory functions. The cilium’s capacity to sense and transduce extracellular signals depends on dynamic trafficking of ciliary membrane proteins. This trafficking is often mediated by the Bardet–Biedl Syndrome complex (BBSome), a protein complex for which the precise subcellular distribution and mechanisms of action are unclear. In humans, BBSome defects perturb ciliary membrane protein distribution and manifest clinically as Bardet–Biedl Syndrome. Cilia are also important in several parasites that cause tremendous human suffering worldwide, yet biology of the parasite BBSome remains largely unexplored. We examined BBSome functions in Trypanosoma brucei, a flagellated protozoan parasite that causes African sleeping sickness in humans. We report that T. brucei BBS proteins assemble into a BBSome that interacts with clathrin and is localized to membranes of the flagellar pocket and adjacent cytoplasmic vesicles. Using BBS gene knockouts and a mouse infection model, we show the T. brucei BBSome is dispensable for flagellar assembly, motility, bulk endocytosis, and cell viability but required for parasite virulence. Quantitative proteomics reveal alterations in the parasite surface proteome of BBSome mutants, suggesting that virulence defects are caused by failure to maintain fidelity of the host–parasite interface. Interestingly, among proteins altered are those with ubiquitination-dependent localization, and we find that the BBSome interacts with ubiquitin. Collectively, our data indicate that the BBSome facilitates endocytic sorting of select membrane proteins at the base of the cilium, illuminating BBSome roles at a critical host–pathogen interface and offering insights into BBSome molecular mechanisms.

Ubiquitin Links Smoothened to Intraflagellar Transport to Regulate Hedgehog Signaling

“…Trypanosomes, Ub co-purifies with the BBSome and BBSome mutations affect cellular localization of ubiquitinated receptors (Langousis et al, 2016). In C. elegans, attachment of Ub to the ciliary membrane protein Pkd2, reduced its level in cilia and similar to our findings, the BBSome appeared to be required (Hu et al, 2007;Xu et al, 2015).…”

IFT25 is required for the construction of the trypanosome flagellum

“…If the later two contribute to the trafficking of BBS proteins, one could envisage that the spermatozoa phenotype is due to BBS perturbation. In trypanosomes, BBS proteins are not found in the flagellum but are located at the flagellar pocket membrane and the endocytic network (Langousis et al, 2016). Knockout of BBS genes does not impact flagellum construction and parasites even infect mice normally (Langousis et al, 2016).…”

Endocytic Control of Cellular Signaling at the Primary Cilium

“…In C. elegans, the periciliary membrane compartment constitutes a specialized domain for the coordinated sorting of clathrin-positive vesicles, which are exocytosed in a BBS8-and Rab8-dependent manner to regulate ciliary and periciliary membrane homeostasis, as well as ciliary targeting of GPCRs and transient receptor potential (TRP) ion channels that are involved in behavioral responses and dauer formation [67,69,70]. Similarly, endocytosis of signaling complexes at the periciliary membrane of the photoreceptor is linked to the coordination of vesicular cargo delivery to the connecting cilium [71] and, in Trypanosoma brucei, the BBSome associates with the flagellar pocket to facilitate endocytic sorting of membrane proteins which are required for parasite virulence [72].…”