Characterization of leishmania RNA virus infected leishmania major exosomes and evaluation of the impact of cytosolic DNA sensing related pathways in L. Major infection

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2020

Author

Yılmaz, İsmail Cem

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Leishmaniasis is a neglected infectious disease caused by Leishmaniaparasite. The disease represents a significant health problem, necessitating urgent development of effective treatment strategies. New World Leishmaniaspecies harboringLeishmania RNA virus (LRV) causesevere,metastatic disease. In this thesis, we compared LRV2-1 deficient and proficient Old World L. majorstrains in terms of their exosomal content and infectivity. PCR and mass spectrometry based analysis revealed that virus infectedLeishmaniaexosomes containedat least 64%of the viralRNA but not the viral proteins. LRV2-1positive and negative L.majorstrains had similar infectivity in macrophages, the positive strain exhibited delayed infection in a cutaneous leishmaniasis model in mice.Next,we aimedto investigatethe role ofcGAS-STING DNA sensing pathwayinLeishmania infection. Infection rates inSTING and TBK-1 knockout cell lines were significantly lower than wild type THP-1 cells and this difference wasrelated to decreased phagocytic capacities of the knockout lines. Effect of the TBK1 inhibitor amlexanox was investigated in in vitroand in vivoinfection models. The drug blocked parasite phagocytosis in macrophages and delayed footpad swellingin parasite infected mice, suggesting that amlexanox could be of interest asa candidate drug in cutaneous leishmaniasis treatment. We alsoaimed to determine the virole of ISG15 in L. majorinfectionand autophagy by mutating the ISG15 using a LentiCRISPRv2 system.Our findings revealed apro-parasitic role ofISG15 inearlystages of Leishmaniainfection. Furthermore, we found that autophagy was enhanced in ISG15 KO cells,suggestinga regulatory role forISG15 in autophagy. Collectively,our findings might benefit future work related to Leishmaniaimmunity in defining new targets and developing effective treatment strategies.

Subject Keywords

Leishmaniasis., Leishmania RNA virus, Leishmania, amlexanox, ISG15, cGAS, STING, TBK-1, exosome, autophagy

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http://etd.lib.metu.edu.tr/upload/12625226/index.pdf
https://hdl.handle.net/11511/45059

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Graduate School of Natural and Applied Sciences, Thesis

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İ. C. Yılmaz, “Characterization of leishmania RNA virus infected leishmania major exosomes and evaluation of the impact of cytosolic DNA sensing related pathways in L. Major infection,” Thesis (Ph.D.) -- Graduate School of Natural and Applied Sciences. Biology., Middle East Technical University, 2020.