In neural stem and progenitor cells (NSCs), the nuclear receptor TLX is a key regulator of proliferation, self-renewal, and differentiation into all three major neural cell types: neurons, astrocytes, and oligodendrocytes. As a transcription factor, TLX functions by inhibiting pro-apoptotic regulators and negative regulators of proliferation. Since the endogenous ligand of TLX has not yet been identified, this nuclear receptor is considered an “orphan.” We previously found that NSCs have a distinct metabolic signature that is highly enriched with fatty acids when compared to other neural cell types, with mono-unsaturated fatty acids (MUFAs) being most abundant. Further profiling also found that these fatty acid concentrations correlate with density of NPCs in the proliferating state. We seek to evaluate ligand binding to human TLX using a biophysical approach. Using the thermofluor assay, cellular thermal shift assay, and bio-layer interferometry, the MUFAs 22:1ω9 and 18:1ω5 were observed to interact with TLX and were identified as candidates for the endogenous ligand. Initial drug screening using these binding assays has also revealed lead compounds with binding comparable to that of the endogenous fatty acids. We are currently optimizing these compounds with the hope of presenting TLX as a promising target for large-scale drug discovery. To observe changes in TLX transcriptional activity, we designed a luciferase reporter system to determine how TLX function can be regulated by ligands that have already demonstrated an ability to interact with the protein. Furthermore, we are currently in the process of validating the downstream targets of TLX using ChIP-qPCR and ChIP-seq, so that we can determine how fatty acids like 22:1ω9, 18:1ω5, and promising synthetic compounds affect their expression. Neurogenesis has been implicated in not only learning and memory formation, but also in the biological response to neurological injury and neurodegenerative diseases. Elucidating the regulatory pathways that govern this phenomenon may provide more robust strategies to counter depression, anxiety, addiction, schizophrenia, and age-related disorders like Alzheimer’s and Parkinson’s.

Discovery of endogenous and synthetic ligands for orphan nuclear receptor TLX

253rd National Meeting (2017)

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