A novel Approach to Target Trypanosome, Leishmania and Superbugs to the Lysosomal Compartment

这项研究的长期目标是开发一种新的治疗方法，通过提供抗病毒药物，抗生素和疫苗的替代疗法，彻底改变传染性疾病的治疗方式。锥虫（锥虫属），利什曼病（利什曼原虫属），布鲁氏菌病（布鲁氏菌属），和结核病（分枝杆菌属），目前还没有有效的治愈或治疗方法，是野生物种、家畜和人类的主要传染源。众所周知，大多数病原体通过附着于现有的细胞表面受体而经由内吞作用侵入细胞。一旦进入细胞，病原体就会欺骗吞噬体和溶酶体，并优先在其中分裂。在真核细胞中，将新合成的蛋白质递送至内体、吞噬体和溶酶体依赖于高尔基体，即，将蛋白质分类并定位到这些目的地的细胞器。递送至内体、吞噬体和溶酶体的蛋白质包括酶和激活蛋白。它们通过特殊的分选受体被引导到其中。我们发现，在溶酶体激活蛋白“Prosaposin”（PSAP）的区域包含一个基序，其结合到一个名为“分拣蛋白”的分选受体，因此其运输到内溶酶体隔室所需的。在我们的最后一次“NSERC资助”期间，我们证明了与PSAP的溶酶体靶向基序连接的抗微生物蛋白-防御素和Tachyplesin被递送到内体和溶酶体。在该提议中，我们想要测试以下假设：1）-防御素-PSAP和鲎素-PSAP经由分拣蛋白被导向内溶酶体系统; 2）鲎素-PSAP在感染细胞的溶酶体区室中具有杀锥虫和杀利什曼原虫的作用; 3）-防御素-PSAP在感染细胞的溶酶体区室中具有杀菌作用。因此，本研究将评估这些抗微生物融合蛋白杀死锥虫、利什曼原虫和吞噬体和溶酶体内的细菌的有效性。我们的研究将允许建立一个新的策略，消除病原体的内/溶酶体隔室的概念证明。我们的研究结果将与野生，濒危和家养动物物种以及人类的传染病治疗相关，并将开辟无数途径来检查病毒，细菌和原生动物感染中抗病原肽的递送和作用。

The long term objective of this research is to develop a novel therapeutic approach that will revolutionize the way transmissible diseases are treated, by offering an alternative therapy to antiviral drugs, antibiotics and vaccines. Trypanosomiasis (Trypanosoma spp.), leishmaniosis (Leishmania spp.), brucellosis (Brucella spp.), and tuberculosis (Mycobacterium spp.), for which there are no effective cures or treatments are major infectious agents in wild species, domestic animals and humans. It is well known that the majority of pathogens invade the cell via endocytosis, by attaching to existing cell surface receptors. Once inside the cell, the pathogens trick the phagosomes and lysosomes and preferentially divide within them. In eukaryotic cells the delivery of newly synthesized proteins to endosomes, phagosomes and lysosomes is dependent on the Golgi apparatus, i.e., the organelle that sorts and targets proteins to these destinations. Proteins delivered to endosomes, phagosomes and lysosomes include enzymes and activator proteins. They are directed therein via special sorting receptors. We found that a region in the lysosomal activator protein “Prosaposin” (PSAP) contains a motif required for its binding to a sorting receptor called “Sortilin” and consequently for its transport to the endolysosomal compartment. During our last “NSERC Grant” we demonstrated that the antimicrobial proteins -Defensin and Tachyplesin linked to the lysosomal targeting motif of PSAP were delivered to endosomes and lysosomes. In this proposal, we would like to test the following hypotheses: 1) -Defensin-PSAP and Tachyplesin-PSAP are directed to the endolysosomal system via Sortilin; 2)Tachyplesin-PSAP has trypanocidal and leishmanocidal effects in the lysosomal compartment of infected cells; 3) -Defensin-PSAP has a bactericidal effect in the lysosomal compartment of infected cells. Thus, the present study will assess the effectiveness of these antimicrobial fusion proteins to kill trypanosomes, leishmania and bacteria within phagosomes and lysosomes. Our study will permit to establish a proof of concept of a new strategy for the elimination of pathogens in the endo/lysosomal compartment. The outcome of our research will be relevant for the treatment of infectious diseases in wild, endangered, and domestic animal species, as well as in humans and will open innumerable avenues to examine the delivery and effects of antipathogenic peptides in viral, bacterial and protozoal infections.***