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

2015 ◽  
Vol 113 (3) ◽  
pp. 632-637 ◽  
Author(s):  
Gerasimos Langousis ◽  
Michelle M. Shimogawa ◽  
Edwin A. Saada ◽  
Ajay A. Vashisht ◽  
Roberto Spreafico ◽  
...  
Keyword(s):  
Membrane Proteins ◽  
Trypanosoma Brucei ◽  
Molecular Mechanisms ◽  
Protozoan Parasite ◽  
Infection Model ◽  
Protein Distribution ◽  
Bardet Biedl Syndrome ◽  
Ciliary Membrane ◽  
Mouse Infection Model ◽  
Endocytic Sorting

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.

scholarly journals Trafficking of ciliary membrane proteins by the intraflagellar transport/BBSome machinery

2018 ◽  
Vol 62 (6) ◽  
pp. 753-763 ◽  
Author(s):  
Jenna L. Wingfield ◽  
Karl-Ferdinand Lechtreck ◽  
Esben Lorentzen
Keyword(s):  
Membrane Proteins ◽  
Molecular Motors ◽  
Intraflagellar Transport ◽  
Inherited Disease ◽  
Cell Organelles ◽  
Bardet Biedl Syndrome ◽  
Ciliary Membrane ◽  
Peripheral Membrane Proteins ◽  
Abnormal Accumulation ◽  
And Function

Bardet–Biedl syndrome (BBS) is a rare inherited disease caused by defects in the BBSome, an octameric complex of BBS proteins. The BBSome is conserved in most organisms with cilia, which are microtubule (MT)-based cell organelles that protrude from the cell surface and function in motility and sensing. Cilia assembly, maintenance, and function require intraflagellar transport (IFT), a bidirectional motility of multi-megadalton IFT trains propelled by molecular motors along the ciliary MTs. IFT has been shown to transport structural proteins, including tubulin, into growing cilia. The BBSome is an adapter for the transport of ciliary membrane proteins and cycles through cilia via IFT. While both the loss and the abnormal accumulation of ciliary membrane proteins have been observed in bbs mutants, recent data converge on a model where the BBSome mainly functions as a cargo adapter for the removal of certain transmembrane and peripheral membrane proteins from cilia. Here, we review recent data on the ultrastructure of the BBSome and how the BBSome recognizes its cargoes and mediates their removal from cilia.

scholarly journals Metabolic reprogramming during the Trypanosoma brucei life cycle

F1000Research ◽  
2017 ◽  
Vol 6 ◽  
pp. 683 ◽  
Author(s):  
Terry K. Smith ◽  
Frédéric Bringaud ◽  
Derek P. Nolan ◽  
Luisa M. Figueiredo
Keyword(s):  
Life Cycle ◽  
Trypanosoma Brucei ◽  
Model Organism ◽  
Protozoan Parasite ◽  
Tsetse Fly ◽  
Metabolic Reprogramming ◽  
Mammalian Host ◽  
Insect Vector ◽  
Environmental Signals ◽  
Cellular Metabolic Activity

Cellular metabolic activity is a highly complex, dynamic, regulated process that is influenced by numerous factors, including extracellular environmental signals, nutrient availability and the physiological and developmental status of the cell. The causative agent of sleeping sickness, Trypanosoma brucei, is an exclusively extracellular protozoan parasite that encounters very different extracellular environments during its life cycle within the mammalian host and tsetse fly insect vector. In order to meet these challenges, there are significant alterations in the major energetic and metabolic pathways of these highly adaptable parasites. This review highlights some of these metabolic changes in this early divergent eukaryotic model organism.

scholarly journals An epitope-based malaria vaccine targeting the junctional region of circumsporozoite protein

npj Vaccines ◽  
2021 ◽  
Vol 6 (1) ◽  
Author(s):  
Lucie Jelínková ◽  
Hugo Jhun ◽  
Allison Eaton ◽  
Nikolai Petrovsky ◽  
Fidel Zavala ◽  
...  
Keyword(s):  
Mouse Model ◽  
Malaria Vaccine ◽  
High Titer ◽  
Circumsporozoite Protein ◽  
Infection Model ◽  
Vaccine Platform ◽  
Junctional Region ◽  
Mouse Infection Model ◽  
Virus Like Particle ◽  
Major Surface

AbstractA malaria vaccine that elicits long-lasting protection and is suitable for use in endemic areas remains urgently needed. Here, we assessed the immunogenicity and prophylactic efficacy of a vaccine targeting a recently described epitope on the major surface antigen on Plasmodium falciparum sporozoites, circumsporozoite protein (CSP). Using a virus-like particle (VLP)-based vaccine platform technology, we developed a vaccine that targets the junctional region between the N-terminal and central repeat regions of CSP. This region is recognized by monoclonal antibodies, including mAb CIS43, that have been shown to potently prevent liver invasion in animal models. We show that CIS43 VLPs elicit high-titer and long-lived anti-CSP antibody responses in mice and is immunogenic in non-human primates. In mice, vaccine immunogenicity was enhanced by using mixed adjuvant formulations. Immunization with CIS43 VLPs conferred partial protection from malaria infection in a mouse model, and passive transfer of serum from immunized macaques also inhibited parasite liver invasion in the mouse infection model. Our findings demonstrate that a Qβ VLP-based vaccine targeting the CIS43 epitope combined with various adjuvants is highly immunogenic in mice and macaques, elicits long-lasting anti-CSP antibodies, and inhibits parasite infection in a mouse model. Thus, the CIS43 VLP vaccine is a promising pre-erythrocytic malaria vaccine candidate.

scholarly journals Membrane recruitment of Atg8 by Hfl1 facilitates turnover of vacuolar membrane proteins in yeast cells approaching stationary phase

BMC Biology ◽  
2021 ◽  
Vol 19 (1) ◽  
Author(s):  
Cheng-Wen He ◽  
Xue-Fei Cui ◽  
Shao-Jie Ma ◽  
Qin Xu ◽  
Yan-Peng Ran ◽  
...  
Keyword(s):  
Membrane Proteins ◽  
Membrane Protein ◽  
Stationary Phase ◽  
Molecular Mechanisms ◽  
Multivesicular Body ◽  
Degradation Process ◽  
Vacuolar Membrane ◽  
Yeast Cells ◽  
Membrane Structures ◽  
Membrane Recruitment

Abstract Background The vacuole/lysosome is the final destination of autophagic pathways, but can also itself be degraded in whole or in part by selective macroautophagic or microautophagic processes. Diverse molecular mechanisms are involved in these processes, the characterization of which has lagged behind those of ATG-dependent macroautophagy and ESCRT-dependent endosomal multivesicular body pathways. Results Here we show that as yeast cells gradually exhaust available nutrients and approach stationary phase, multiple vacuolar integral membrane proteins with unrelated functions are degraded in the vacuolar lumen. This degradation depends on the ESCRT machinery, but does not strictly require ubiquitination of cargos or trafficking of cargos out of the vacuole. It is also temporally and mechanistically distinct from NPC-dependent microlipophagy. The turnover is facilitated by Atg8, an exception among autophagy proteins, and an Atg8-interacting vacuolar membrane protein, Hfl1. Lack of Atg8 or Hfl1 led to the accumulation of enlarged lumenal membrane structures in the vacuole. We further show that a key function of Hfl1 is the membrane recruitment of Atg8. In the presence of Hfl1, lipidation of Atg8 is not required for efficient cargo turnover. The need for Hfl1 can be partially bypassed by blocking Atg8 delipidation. Conclusions Our data reveal a vacuolar membrane protein degradation process with a unique dependence on vacuole-associated Atg8 downstream of ESCRTs, and we identify a specific role of Hfl1, a protein conserved from yeast to plants and animals, in membrane targeting of Atg8.

scholarly journals Polo-like kinase is required for Golgi and bilobe biogenesis in Trypanosoma brucei

2008 ◽  
Vol 181 (3) ◽  
pp. 431-438 ◽  
Author(s):  
Christopher L. de Graffenried ◽  
Helen H. Ho ◽  
Graham Warren
Keyword(s):  
Cell Cycle ◽  
Endoplasmic Reticulum ◽  
Trypanosoma Brucei ◽  
Protozoan Parasite ◽  
Controlled Assembly

A bilobed structure marked by TbCentrin2 regulates Golgi duplication in the protozoan parasite Trypanosoma brucei. This structure must itself duplicate during the cell cycle for Golgi inheritance to proceed normally. We show here that duplication of the bilobed structure is dependent on the single polo-like kinase (PLK) homologue in T. brucei (TbPLK). Depletion of TbPLK leads to malformed bilobed structures, which is consistent with an inhibition of duplication and an increase in the number of dispersed Golgi structures with associated endoplasmic reticulum exit sites. These data suggest that the bilobe may act as a scaffold for the controlled assembly of the duplicating Golgi.

scholarly journals Ciliary membrane proteins traffic through the Golgi via a Rabep1/GGA1/Arl3-dependent mechanism

2014 ◽  
Vol 5 (1) ◽  
Author(s):  
Hyunho Kim ◽  
Hangxue Xu ◽  
Qin Yao ◽  
Weizhe Li ◽  
Qiong Huang ◽  
...  
Keyword(s):  
Membrane Proteins ◽  
Ciliary Membrane ◽  
Dependent Mechanism

scholarly journals Trafficking to the primary cilium membrane

2017 ◽  
Vol 28 (2) ◽  
pp. 233-239 ◽  
Author(s):  
Saikat Mukhopadhyay ◽  
Hemant B. Badgandi ◽  
Sun-hee Hwang ◽  
Bandarigoda Somatilaka ◽  
Issei S. Shimada ◽  
...  
Keyword(s):  
Membrane Proteins ◽  
Signaling Pathways ◽  
Primary Cilium ◽  
Hedgehog Signaling ◽  
Diffusion Barriers ◽  
Disease Pathogenesis ◽  
Ciliary Membrane ◽  
Multiple Organs ◽  
Sensory Reception

The primary cilium has been found to be associated with a number of cellular signaling pathways, such as vertebrate hedgehog signaling, and implicated in the pathogenesis of diseases affecting multiple organs, including the neural tube, kidney, and brain. The primary cilium is the site where a subset of the cell's membrane proteins is enriched. However, pathways that target and concentrate membrane proteins in cilia are not well understood. Processes determining the level of proteins in the ciliary membrane include entry into the compartment, removal, and retention by diffusion barriers such as the transition zone. Proteins that are concentrated in the ciliary membrane are also localized to other cellular sites. Thus it is critical to determine the particular role for ciliary compartmentalization in sensory reception and signaling pathways. Here we provide a brief overview of our current understanding of compartmentalization of proteins in the ciliary membrane and the dynamics of trafficking into and out of the cilium. We also discuss major unanswered questions regarding the role that defects in ciliary compartmentalization might play in disease pathogenesis. Understanding the trafficking mechanisms that underlie the role of ciliary compartmentalization in signaling might provide unique approaches for intervention in progressive ciliopathies.

NgCAM and VAMP2 Reveal that Direct Delivery and Dendritic Degradation Maintain Axonal Polarity

2021 ◽  
Author(s):  
Alec T. Nabb ◽  
Marvin Bentley
Keyword(s):  
Membrane Proteins ◽  
Molecular Mechanisms ◽  
Minor Role ◽  
Axonal Membrane ◽  
Quantitative Analyses ◽  
Direct Delivery ◽  
Novel Strategy ◽  
Extreme Scale ◽  
A Minor

Neurons are polarized cells of extreme scale and compartmentalization. To fulfill their role in electrochemical signaling, axons must maintain a specific complement of membrane proteins. Despite being subject of considerable attention, the trafficking pathway of axonal membrane proteins is not well understood. Two pathways, direct delivery and transcytosis, have been proposed. Previous studies reached contradictory conclusions about which of these mediates delivery of axonal membrane proteins to their destination, in part because they evaluated long-term distribution changes and not vesicle transport. We developed a novel strategy to selectively label vesicles in different trafficking pathways and determined the trafficking of two canonical axonal membrane proteins, NgCAM and VAMP2. Results from detailed quantitative analyses of transporting vesicles differed substantially from previous studies and found that axonal membrane proteins overwhelmingly undergo direct delivery. Transcytosis plays only a minor role in axonal delivery of these proteins. In addition, we identified a novel pathway by which wayward axonal proteins that reach the dendritic plasma membrane are targeted to lysosomes. These results redefine how axonal proteins achieve their polarized distribution, a crucial requirement for elucidating the underlying molecular mechanisms. [Media: see text] [Media: see text] [Media: see text] [Media: see text]

scholarly journals Structure-based drug design against Trypanosoma brucei methionyl-tRNA synthetase

2014 ◽  
Vol 70 (a1) ◽  
pp. C708-C708
Author(s):  
Cho Yeow Koh ◽  
Jasmine Nguyen ◽  
Sayaka Shibata ◽  
Zhongsheng Zhang ◽  
Ranae Ranade ◽  
...  
Keyword(s):  
Crystal Structures ◽  
Trypanosoma Brucei ◽  
High Throughput ◽  
High Throughput Screening ◽  
Protozoan Parasite ◽  
Trna Synthetase ◽  
Structure Based Drug Design ◽  
Chemical Structures ◽  
Ic50 Values ◽  
Methionyl Trna Synthetase

Infection by the protozoan parasite Trypanosoma brucei causes human African trypanosomiasis, commonly known as sleeping sickness. The disease is fatal without treatment; yet, current therapeutic options for the disease are inadequate due to toxicity, difficulty in administration and emerging resistance. Therefore, methionyl-tRNA synthetase of T. brucei (TbMetRS) is targeted for the development of new antitrypanosomal drugs. We have recently completed a high-throughput screening campaign against TbMetRS using a 364,131 compounds library in The Scripps Research Institute Molecular Screening Center. Here we outline our strategy to integrate the power of crystal structures with high-throughput screening in a drug discovery project. We applied the rapid crystal soaking procedure to obtain structures of TbMetRS in complex with inhibitors reported earlier[1] to approximately 70 high-throughput screening hits. This resulted in more than 20 crystal structures of TbMetRS·hit complexes. These hits cover a large diversity of chemical structures with IC50 values between 200 nM and 10 µM. Based on the solved structures and existing knowledge drawn from other in-house inhibitors, the IC50 value of the most promising hit has been improved. Further development of the compounds into potent TbMetRS inhibitors with desirable pharmacokinetic properties is on-going and will continue to benefit from information derived from crystal structures.

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