dc.contributor.advisor | Rossner, Moritz Prof. Dr. | |
dc.contributor.author | Herholt, Alexander | |
dc.date.accessioned | 2017-07-24T09:41:51Z | |
dc.date.available | 2017-07-24T09:41:51Z | |
dc.date.issued | 2017-07-24 | |
dc.identifier.uri | http://hdl.handle.net/11858/00-1735-0000-0023-3EB1-0 | |
dc.identifier.uri | http://dx.doi.org/10.53846/goediss-6199 | |
dc.language.iso | eng | de |
dc.rights.uri | http://creativecommons.org/licenses/by-nc-nd/4.0/ | |
dc.subject.ddc | 570 | de |
dc.title | Development of a multiplexed RNAi-coupled sensor assay to study neuronal function on the large-scale | de |
dc.type | doctoralThesis | de |
dc.contributor.referee | Rossner, Moritz Prof. Dr. | |
dc.date.examination | 2016-08-10 | |
dc.description.abstracteng | Psychiatric diseases such as schizophrenia, bipolar disorder and autism spectrum disorders
are considered neurodevelopmental synaptopathies. Compelling evidence obtained from
large-scale genome-wide association studies, identified a plethora of genetic variations
within hundreds of genes encoding components of the post-synaptic compartment and
calcium signaling mediating excitation-transcription-coupling. This synapse-to-nucleus
signaling is thought to be particularly important for synaptic plasticity and learning and
memory. In the last decade, tremendous progress has been made in neuroscience research
when employing an array of molecular and cellular techniques to study the impact of
individual genes on synaptic plasticity. Nevertheless, neuroscience methodology lags behind
the field of cancer research in terms of large scale functional genetic screens, e.g. mediated
by RNA interference (RNAi). The underlying cause is likely due to both the difficulties of
culturing post-mitotic neurons and the phenotypic complexity. In this regard,
comprehensively identifying genes involved in neuronal excitation and synapse-to-nucleus
signaling may not only deepen our understanding of the corresponding biological processes,
but might also be key in unearthing promising targets for psychiatric drug discovery.
I have developed a functional genomics tool that is applicable to primary neurons and
combines the throughput of a pooled RNAi screen with the sensitivity of a pathway reporter
assay based on the synaptic activity-response element, modified from the Arc enhancer.
This thesis describes a proof-of-concept study in which an AAV-based RNAi library was
screened for regulators of neuronal excitation and synapse-to-nucleus signaling. The assay
principle relies on molecular barcodes, which serve as quantitative reporters, while at the
same time also functioning as unique identifiers of the targeted genes. Upon synaptic
stimulation, the screen identified a multitude of known genes involved in glutamatergic
synapse-to-nucleus signaling, as well as previously unknown candidates like the chemokine
receptor XCR1. The technical approach’s reproducibility has been verified by substantial
overlap of gene hits during three independent screens. Later in the thesis, I also present the
principal applicability of CRISPR-Cas9 tools in neurons, which may improve performance for
genetic interference screens in the near future.
This assay seeks to enhance the analytic toolbox used for analyzing regulatory processes
during neuronal signaling and for the identification of novel targets in psychiatric drug
discovery. | de |
dc.contributor.coReferee | Brose, Nils Prof. Dr. | |
dc.subject.eng | Neuroscience | de |
dc.identifier.urn | urn:nbn:de:gbv:7-11858/00-1735-0000-0023-3EB1-0-1 | |
dc.affiliation.institute | Göttinger Graduiertenschule für Neurowissenschaften, Biophysik und molekulare Biowissenschaften (GGNB) | de |
dc.subject.gokfull | Biologie (PPN619462639) | de |
dc.identifier.ppn | 894189883 | |