Title Aggregation-resistant mutants of schizophrenia-related protein TRIOBP-1
Title (croatian) Otpornost na agregaciju mutanata proteina TRIOBP-1 povezanog sa schizofrenijom
Author Anja Hart
Mentor Nicholas James Bradshaw (mentor)
Committee member Antonija Jurak Begonja (predsjednik povjerenstva)
Committee member Christian Andrew Reynolds (član povjerenstva)
Granter University of Rijeka (Faculty of Biotechnology and Drug Development) Rijeka
Defense date and country 2020-09-22, Croatia
Scientific / art field, discipline and subdiscipline BIOTECHNICAL SCIENCES Biotechnology Molecular Biotechnology
Abstract Schizophrenia is a chronic mental illness, characterized by severe symptoms such as delusions, hallucinations, disorganized speech, lack of motivation, and unpredictable behavior. The development of schizophrenia is caused by genetic and environmental factors. The complexity of these factors has made it difficult to develop new methods for the successful treatment of patients. Recent research has shown that disruption of proteostasis may also impact on the progression of schizophrenia. Disruption in proteostasis causes certain proteins to misfold and therefore aggregate if the cell fails to degrade them. Aggregation is a process in which large insoluble structures known as aggregates are formed. So far, five proteins have been identified that may aggregate in schizophrenia. Among these five is TRIO and F-actin-binding protein (TRIOBP-1), the protein of interest in this thesis. Using C-terminally truncated constructs, it was discovered that the critical region for aggregation of TRIOBP-1 is located in the central section of the protein. As a next step, different truncated variants of TRIOBP-1 were expressed in neuroblastoma cells and immunofluorescent microscopy was used as a way to visualize aggregation. In this way, the critical region for aggregation was narrowed down to a sequence of less than 10 amino acids. To expand and verify this, the next step was to use full-length constructs with a small deletion from their center. In this thesis, four such constructs were used. Unexpectedly, all four of them formed aggregates in cells. This finding indicates that either the PH domain, or an optional 59 disordered amino acids at the N-terminus have effect on TRIOBP-1’s aggregation propensity which was previously thought to be very unlikely. The next step is to generate plasmids lacking the optionally translated N-terminal unstructured region, and look into the effect it may have on aggregation. By generating a TRIOBP-1 mutant with the minimal number of mutations required to prevent aggregation, we will be able to generate model systems for studying TRIOBP-1 aggregation. This will allow us to better understand the role of TRIOBP-1 in the progression of schizophrenia.
Abstract (croatian) Shizofrenija je kronična mentalna bolest koju karakteriziraju teški simptomi kao što su zabluda, halucinacije, neorganiziran govor, nedostatak motivacije i nepredvidivo ponašanje. Razvoj shizofrenije uzrokovan je genetskim i okolišnim čimbenicima. Složenost ovih čimbenika otežala je razvoj novih metoda za uspješno liječenje pacijenata. Nedavna istraživanja pokazala su da poremećaj proteostaze također može utjecati na napredovanje shizofrenije. Poremećaj proteostaze uzrokuje pogrešno savijanje određenih proteina i s time stvaranje agregata ako ih stanica ne razgradi. Agregacija je proces u kojem se formiraju netopive velike strukture poznate kao agregati. Do sada je identificirano pet proteina koji mogu stvarati agregate u shizofreniji. Među tih pet je i TRIO and F-actin-binding protein (TRIOBP-1), protein od interesa u ovom radu. Koristeći C-terminalno skraćene konstrukte, otkriveno je da se kritična regija za agregaciju TRIOBP-1 nalazi u središnjoj regiji proteina. Kao sljedeći korak, različite skraćene varijante TRIOBP-1 izražene su u stanicama neuroblastoma, a agregacija je vizualizirana pomoću imunofluorescentne mikroskopije. Na taj se način kritična regija za agregaciju suzila na niz od manje od 10 aminokiselina. Da bi se ovo potvrdilo, sljedeći korak je bio upotreba konstrukata pune duljine s malim delecijama iz njihovog središta. U ovom su radu korištena 4 takva konstrukta. Neočekivano, sva 4 konstrukta su tvorila agregate u stanicama. Ovo otkriće ukazuje da ili PH domena ili 59 opcinalnih nestrukturiranih aminokiselina na N-terminalnom kraju utječu na sklonost agregacije TRIOBP-1, za koju se ranije smatralo obrnuto. Sljedeći korak je stvaranje plazmida kojima nedostaje opcionalno translatirana N-terminalna nestrukturirana regija, te razmotriti učinak koji ona može imati na agregaciju. Generiranjem TRIOBP-1 mutanta s minimalnim brojem mutacija potrebnih da se spriječi agregacija, moći ćemo generirati modelne sustave za proučavanje TRIOBP-1 agregacije. To će nam omogućiti da bolje razumijemo ulogu TRIOBP-1 u napredovanju shizofrenije.
Keywords
schizophrenia
TRIOBP-1
protein aggregation
mental illness
Keywords (croatian)
shizofrenija
TRIOBP-1
agregacija proteina
mentalne bolesti
Language english
URN:NBN urn:nbn:hr:193:419958
Study programme Title: Biotechnology and drug research Study programme type: university Study level: undergraduate Academic / professional title: sveučilišni/a prvostupnik/prvostupnica (baccalaureus/baccalaurea) biotehnologije i istraživanja lijekova (sveučilišni/a prvostupnik/prvostupnica (baccalaureus/baccalaurea) biotehnologije i istraživanja lijekova)
Type of resource Text
File origin Born digital
Access conditions Open access
Terms of use
Repository Repository of the University of Rijeka, Faculty of Biotechnology and Drug Development
Created on 2020-09-28 09:54:30