Establishing a readout for validating genetic interactions of C. elegans PINK-1, LRK-1 and SGK-1 and their putative roles in regulating endomembrane trafficking
Endomembrane trafficking (ET) is required for maintaining cell homeostasis in eukaryotes. Endocytic recycling, endomembrane vesicular transport, autophagy, mitophagy are considered important aspects of ET process. LRRK2/LRK-1, PINK1/PINK-1, two genes associated with hereditary Parkinson?s Disease, a...
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| Formato: | Tesis de maestría acceptedVersion |
| Lenguaje: | Inglés |
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Facultad de Farmacia y Bioquímica
2015
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| Acceso en línea: | http://repositoriouba.sisbi.uba.ar/gsdl/cgi-bin/library.cgi?a=d&c=afamaster&cl=CL1&d=HWA_2893 http://repositoriouba.sisbi.uba.ar/gsdl/collect/afamaster/index/assoc/HWA_2893.dir/2893.PDF |
| Aporte de: |
| Sumario: | Endomembrane trafficking (ET) is required for maintaining cell homeostasis in eukaryotes. Endocytic recycling, endomembrane vesicular transport, autophagy, mitophagy are considered important aspects of ET process. LRRK2/LRK-1, PINK1/PINK-1, two genes associated with hereditary Parkinson?s Disease, and SGK1/SGK-1, in distinct experimental settings, have been suggested to play either direct or indirect roles in regulating these processes. Previous work of my lab had indicated that these genes and their encoded proteins may be mechanistically linked. In addition, C. elegans PINK-1 and LRK-1 have been shown to function antagonistically in stress response and neurite outgrowth and also interaction between SGK-1 and hLRRK2/CeLRK-1 has been suggested in controlling cytoskeletal dynamics which may affect endocytosis and intracellular cell sorting. Our hypothesis is that PINK1, LRRK2, and SGK1 may share a common function in ET. Currently available readouts for the function of these genes do not relate to ET. Studying genetic interaction of these genes in C. elegans requires screenable phenotype associated with ET, and this was the focus of my master project.\nIn this work with mutant analysis and pharmacological studies I have found several new phenotypic readouts using pink-1 mutant animals. Vacuole phenotype observed in pink-1 animals was found to be rescued by loss of lrk-1 in consistency with the anatagonistic function of PINK-1 and LRK-1, although present results suggest that this vacuole phenotype may be independent of ET perturbations. In another readout, modulation of RAB-10 marker protein (rab-10::gfp) in the intestinal cells of pink-1 animals suggested the potential role of PINK-1 in controlling transport of early endosomes during endomembrane trafficking. Furthermore I have also used the vacuole phenotype in SGK-1 transgene for an RNAi test to study putative interaction between LRK-1, PINK-1 and SGK-1, resulting in the identification of several modulators of this phenotype. Moreover lrk-1 downregulation was sufficient to reduce the levels of transgenic SGK-1 expression, suppressing its phenotypic consequences. This data further support the existence of a mechanistic connection between LRK-1 and SGK-1. |
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