Investigating Unkempt in mTOR signalling and mammalian neurogenesis

Student thesis: Doctoral ThesisDoctor of Philosophy


The insulin receptor (InR)/mechanistic target of rapamycin (mTOR) pathway is a key regulator of neurogenesis, and mTOR dysregulation is associated with neurological diseases such as epilepsy, autism and attention deficit hyperactivity disorder (ADHD). The downstream mechanism by which mTOR regulates neurogenesis is poorly understood. Unkempt (Unk) was the first neurogenic component of the mTOR pathway identified. Unk is a zinc finger/RING domain protein that negatively regulates neuronal differentiation in Drosophila, and cortical migration in mice. However, the role of Unk in mammalian neurogenesis is largely uncharacterised. In this thesis, I aim to investigate the mechanistic relationship between Unk and mTOR, and to characterise the role of Unk in mammalian neurogenesis. To investigate the mechanistic relationship between Unk and mTOR, I manipulated the InR/mTOR pathway in mammalian cell lines. I found that Unk is phosphorylated downstream of mTOR complex I (mTORC1), but in a divergent pathway from the cellular growth regulator S6 kinase. Instead, inhibition of mTORC1 leads to the ubiquitination of Unk. I then used liquid-chromatography tandem mass spectrometry (LC-MS/MS) to identify phosphorylated residues in Unk. This led to the identification of novel phosphorylation sites in Unk, and one potential mTOR substrate residue. To investigate the role of Unk in mammalian neurogenesis, I created and characterised the first Unk nervous system conditional knock-out (cKO) mouse. A behavioural battery assessing the motor function, sociability, anxiety, memory and cognition in Unk cKO mice was performed. The results demonstrated that Unk cKO mice are hyperactive and have enhanced learning and cognitive flexibility. This may be related to preliminary neuroanatomical phenotypes observed. Namely, that Unk cKO mice have more proliferating cells in the embryonic neurogenic niche, and altered alignment of cell bodies in the adult hippocampus. These data are consistent with existing knowledge of InR/mTOR signalling in mammalian neurogenesis and have opened new avenues for future research to investigate the downstream functions of Unk.
Date of Award1 Apr 2019
Original languageEnglish
Awarding Institution
  • King's College London
SupervisorJoseph Bateman (Supervisor), Giovanna Lalli (Supervisor) & Albert Basson (Supervisor)

Cite this