ALS Sinek Modelinde Nörodejenerasyon Mekanizmasının Proteomik ve Genetik Analizi

Öz

Amaç : TDP-43; RNA bağlanma motifleri içeren ve pre-mRNA ekleme, transkripsiyon, mRNA stabilitesi

ve mRNA transferinde yer aldığı rapor edilen oldukça korunmuş ve her yerde eksprese edilen

nükleer proteindir. Son zamanlarda TDP-43 yapılan çalışmalar ile Amyotrofik Lateral Skleroz (ALS)

hastalarının etkilenmiş beyin bölgelerinde gözlenen hücre içi inklüzyonların ana protein bileşeni olarak

tanımlandı. Bu çalışmada Drosophila melanogaster’de TDP-43’ün analoğunun (TBPH’in) ALS modelindeki

sinekler üzerinde proteomik ve genetik mekanizmaları içerisindeki rolünün araştırılması amaçlanmıştır.

Materyal-Metod : WIII8,UAS GFP, GMR,GMR TBPH genotiplerinde bakire dişi sinekler ve erkekler

uygun besiyeri ortamında çiftleştirilerek farklı genotip ve fenotiplerde ALS için etki gösterebilen

transgenik model sinekler üretildi. Transgenik sineklere TBPH entegre edildi. PCR , Jel Elektroforezi ve

2D kütle spektrofotometri yöntemleri kullanılarak TBPH mutantlarının analizi yapıldı.

Bulgular : TDP-43’ün analoğu olan TBPH’i eksprese eden farklı transgenik sinekler kullanılarak ,

TBPH’in ekspresyonunun kontrol gruplarına kıyasla nörodejenerasyona neden olup olmadığı kontrol

edildi. TPBH’nin ekspresyonunun sineklerin gözlerinde siyah lekelerin (nekrozun) gelişmesine neden

olduğu ve gözlerde nörodejenerasyon oluşturduğu görüldü. Dahası, mutasyon geçirmiş sinekler üzerinde

2D jel analizi ile protein agregasyonunda TBPH’in rolünü düşündüren sinek türlerine kıyasla diğer

sineklerin gözlerinde azalmış yağ vücut proteinleri 1 (Fbp1) seviyeleri gösterildi.

Sonuçlar : Sonuç olarak TBPH’in Fbp1’nın üretimini artırarak nörodejeneratif sürece karıştığı görüldü.

Anahtar Kelimeler

• ALS,Drosophila,TDP-43,Fbp1

Proteomic and Genetic Analysıs of the Mechanism of Neurodegeneration in a Fly Model of ALS

Öz

Objective : TDP-43 is a highly conserved and ubiquitously expressed nuclear protein containing

RNA binding motives and reported to be involved in pre-mRNA splicing, transcription, mRNA stability

and mRNA transport. Recently, TDP-43 was identified as the main protein component of the intracellular

inclusions observed in affected brain areas of patients suffering from Amyotrophic Lateral Sclerosis

(ALS).

Material and Methods : In WIII8, UAS GFP, GMR, GMR TBPH genotypes, virgin and male transgenic

model flies were produced which were able to act for ALS in different genotypes and phenotypes by

mating them in the appropriate medium. TBPH was integrated into the transgenic flies. TBPH mutants

were analyzed using PCR, Gel Electrophoresis and 2D mass spectrophotometry methods.

Results : We have used different transgenic flies expressing TDP-43 analogue TBPH and checked

whether the expression of TBPH is causing the neurodegeneration as compared to the control groups.

Expression of TPBH caused development of black spot (necrosis) in the eyes of the flies clearly suggested

neurodegeneration in the eye. Moreover, 2D gel analysis on TBPH mutated flies showed reduced

fat body proteins 1 (Fbp1) levels in the eyes of flies as compared to wild type, suggesting the role of

TBPH in the protein aggregation.

Conclusion : Our date showed TBPH is involved in the neurodegenerative process by enhancing the

production of Fbp11.

Anahtar Kelimeler

• ALS,Drosophila,TDP-43,TBPH,Fbp1

Kaynakça

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