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Tau is natively an unfolded protein that promotes the assembly and the stability of the axonal microtubules in the central nervous system. Increased formation of Tau protein aggregates has been causatively implicated in several neurodegenerative diseases called tauopathies. In the present study, we used the Drosophila melanogaster system to express the longest isoform of human Tau (2N4R) in the nervous system of adult flies, recreating the main features of the human pathology. Herein, this Tau-mediated neurodegeneration model was used as a platform to perform genetic screenings to identify putative modifiers of Tau toxicity. Our strategy exploited the modulation of genes considered as risk factors of Alzheimer’s disease (AD), Frontotemporal Dementias and other neurodegenerative diseases by RNA interference in vivo. This approach allowed us to identify a new gene which participates in the neuronal response against Tau induced neurotoxicity in Drosophila: D-Reep1, homologue of human REEP1 gene (h-Reep1). D-Reep1 knockout flies showed no apparent phenotypes in physiological growing and developmental conditions, however, they showed peculiar sensitivity to stress conditions. In addition, D-Reep1 knockout enhanced the neurodegeneration mediated by Tau expression in Drosophila eyes. On the contrary, the overexpression of UAS-D-Reep1 and UAS-h-Reep1 abolished the typical rough eye phenotype induced by the presence of Tau. The Co-expression of D-Reep1 in Tau backgrounds did not alter the phosphorylation pattern of this protein while, the presence of D-Reep1 seemed to prevent the formation of Tau aggregates in vivo. Thus, the data support the idea that D-Reep1 exerts a protective role on Tau induced toxicity which is independent of its phosphorylation status. In this work, I analysed the mechanisms behind the neuroprotective role of D-Reep1 and, in particular, I found that REEP1 is involved in the regulation of the unfolded protein response (UPR) through the PERK-ATF4 cascade within the ER. By the activation of this pathway, the neurotoxic aggregates of Tau are removed from Drosophila neuronal tissues rescuing the normal characteristics of the affected tissues. Evidences also suggest that the activation of autophagy was behind the removal of Tau aggregates, providing new molecular information about the physiological role of D Reep1 in the nervous system. V človeškem telesu je Tau prisoten kot nezvit protein in sodeluje pri združevanju in stabilizaciji aksonskih mikrotubulov v centralnem živčnem sistemu. Agregati proteina Tau so karakteristika številnih nevrodegenerativnih bolezni (t.i. tauopatije). Za vzpostavitev simptomov, ki so prisotni pri nevrodegenerativnih boleznih, smo v živčevju odrasle živali D. melanogaster izrazili najdaljšo izoformo humanega proteina Tau (2N4R). Model smo uporabili kot osnovo za genetski pregled domnevnih modifikatorjev toksičnosti proteina Tau. S pomočjo RNA interference smo in vivo identificirali nov gen, kisodeluje v odzivu nevronov na toksično delovanje proteina Tau v D. melanogaster: D-Reep1, homolog človeškega REEP1 gena (h-Reep1). Živali z izbitim D-Reep1 genom niso kazale očitnega fenotipa v fizioloških pogojih. Spremembe so se pokazale šele ob izpostavitvi stresnim dejavnikom. Opazili smo tudi, da odsotnost gena D-Reep1 pospeši nevrodegeneracijo, ki jo povzroči protein Tau v očeh D. melanogaster. Ob izražanju UAS-D-Reep1 in UAS-h-Reep1 kljub toksičnemu delovanju proteina Tau nismo opazili spremenjenega fenotipa na očesu. Ob podrobnejši analizi sicer nismo ugotovili sprememb fosforilacije D-Reep1, protein pa je preprečil nastanek Tau agregatov in vivo. Pri nadaljnih raziskavah smo ugotovili, da je REEP1 vpleten v uravnavanje odziva na nezvite proteine (UPR) preko PERK-ATF4 kaskade znotraj endoplazemskega retikuluma (ER). Aktivacija kaskade je povzročila odstranitev nevrotoksičnih agregatov Tau, živčno tkivo pa ni kazalo patoloških znakov. Rezultati nakazujejo, da je za odstranitev agregatov Tau odgovorna autofagija, kar nam podaja nove informacije o fiziološki vlogi D-Reep1 v živčnem sistemu. |
