Vladana Vukojevic

This interdisciplinary research proposal at the interface between biophysical chemistry, optics/photonics, cell biology and molecular medicine, aims to solidify instrumentation and further advance a new methodology for quantitative spatio-temporally resolved fluorescence microscopy imaging via massively parallel Fluorescence Correlation Spectroscopy (mpFCS) that we have recently developed. The methodology, which has the ultimate, single-molecule sensitivity, will be used in basic and translational studies to:Aim 1a. Dissect in live cells kinetic mechanisms through which gene transcription networks self-adjust their essential variables and control their own biological function.Aim 1b. Provide in live cells quantitative biophysical data and mechanistic insights on how the functional dynamics of oligodendrocyte transcription factor (OLIG2) can be rescued by treatment with novel prospective drug candidates. Aim 2.   Implement this new technology in a translational research study aiming to develop a reliable and affordable non-immune, blood-based assay for early diagnosis of amyloid diseases, such as Alzheimer’s disease (AD).Aim 3a. Development of a system for massively parallel dual-color Fluorescence Cross-Correlation Spectroscopy (mpFCCS).Aim 3b. Develop a fiber optic probe to allow parallelized FCS recordings in behaving animals.

The most common neurodegenerative disease is Alzheimer´s disease, in which the amyloid-ß (Aß) peptide aggregates to amyloid fibers that accumulate in plaques in the human brain. Key aspects of the disease are still unclear of which the following are addressed in this proposal:(i) Aß oligomers have been correlated with the severity of the disease but their structure is controversial. Here, we will generate structural models of several oligomers in aqueous solu tion and in interaction with membranes using isotope-edited infrared (IR) spectroscopy. (ii) Lipids are important modulators of Aß aggregation and are elevated in and around plaques. However, their role in plaque development is unclear. Therefore, we will map the biochemical composition of plaques from human brain with higher (20 nm) resolution than before using nanoscale IR spectroscopy.(iii) Little is known about Aß´s interaction with other amyloidogenic polypeptides. This will be studied with a particular focus on structure and morphology of the co-aggregates. For this, we will combine the above approaches.(iv) There is presently no cure for AD but promising anti-amyloid peptides are known. We will study their interaction with Aß as under (iii) for a further knowledge-based improvement of their design.In addition to pioneering methodological development, our approach will further the molecular understanding of AD in areas that are difficult to study with other methods.