@phdthesis{Gupta2021,
  author    = {Gupta, Rohini},
  title     = {Intracellular self-activation of the TrkB kinase domain causes FAK phosphorylation and disrupts actin filopodia dynamics},
  doi       = {10.25972/OPUS-23382},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-233829},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2021},
  abstract  = {The tropomysin receptor kinase B (TrkB), the receptor for the neurotrophin brain-derived neurotrophic factor (BDNF), plays an important role in neuronal survival, neuronal differentiation, and cellular plasticity. Conventionally, TrkB activation is induced by binding of BDNF at extracellular sites and subsequent dimerization of receptor monomers. Classical Trk signaling concepts have failed to explain ligand-independent signaling of intracellular TrkB or oncogenic NTRK-fusion proteins. The intracellular activation domain of TrkB consists of a tyrosine kinase core, with three tyrosine (Y) residues at positions 701, 705 and 706, that catalyzes the phosphorylation reaction between ATPγ and tyrosine. The release of cisautoinhibition of the kinase domain activates the kinase domain and tyrosine residues outside of the catalytic domain become phosphorylated. The aim of this study was to find out how ligand-independent activation of TrkB is brought about. With the help of phosphorylation mutants of TrkB, it has been found that a high, local abundance of the receptor is sufficient to activate TrkB in a ligand-independent manner. This self-activation of TrkB was blocked when either the ATP-binding site or Y705 in the core domain was mutated. The vast majority of this self-active TrkB was found at intracellular locations and was preferentially seen in roundish cells, lacking filopodia. Live cell imaging of actin dynamics showed that self-active TrkB changed the cellular morphology by reducing actin filopodia formation. Signaling cascade analysis confirmed that self-active TrkB is a powerful activator of focal adhesion kinase (FAK). This might be the reason why self-active TrkB is able to disrupt actin filopodia formation. The signaling axis from Y705 to FAK could be mimicked by expression of the soluble, cytosolic TrkB kinase domain. However, the signaling pathway was inactive, when the TrkB kinase domain was targeted to the plasmamembrane with the help of artificial myristoylation membrane anchors. A cancer-related intracellular NTRK2-fusion protein (SQSTM1-NTRK2) also underwent constitutive kinase activation. In glioblastoma-like U87MG cells, self-active TrkB kinase reduced cell migration. These constitutive signaling pathways could be fully blocked within minutes by clinically approved, anti-tumorigenic Trk inhibitors. Moreover, this study found evidences for constitutively active, intracellular TrkB in tissue of human grade IV glioblastoma. In conclusion, the data provide an explanation and biological function for selfactive, constitutive TrkB kinase domain signaling, in the absence of a ligand.},
  language  = {en}
}
@phdthesis{Andreska2021,
  author    = {Andreska, Thomas},
  title     = {Effects of dopamine on BDNF / TrkB mediated signaling and plasticity on cortico-striatal synapses},
  doi       = {10.25972/OPUS-17431},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-174317},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2021},
  abstract  = {Progressive loss of voluntary movement control is the central symptom of Parkinson's disease (PD). Even today, we are not yet able to cure PD. This is mainly due to a lack of understanding the mechanisms of movement control, network activity and plasticity in motor circuits, in particular between the cerebral cortex and the striatum. Brain-derived neurotrophic factor (BDNF) has emerged as one of the most important factors for the development and survival of neurons, as well as for synaptic plasticity. It is thus an important target for the development of new therapeutic strategies against neurodegenerative diseases. Together with its receptor, the Tropomyosin receptor kinase B (TrkB), it is critically involved in development and function of the striatum. Nevertheless, little is known about the localization of BDNF within presynaptic terminals in the striatum, as well as the types of neurons that produce BDNF in the cerebral cortex. Furthermore, the influence of midbrain derived dopamine on the control of BDNF / TrkB interaction in striatal medium spiny neurons (MSNs) remains elusive so far. Dopamine, however, appears to play an important role, as its absence leads to drastic changes in striatal synaptic plasticity. This suggests that dopamine could regulate synaptic activity in the striatum via modulation of BDNF / TrkB function. To answer these questions, we have developed a sensitive and reliable protocol for the immunohistochemical detection of endogenous BDNF. We find that the majority of striatal BDNF is provided by glutamatergic, cortex derived afferents and not dopaminergic inputs from the midbrain. In fact, we found BDNF in cell bodies of neurons in layers II-III and V of the primary and secondary motor cortex as well as layer V of the somatosensory cortex. These are the brain areas that send dense projections to the dorsolateral striatum for control of voluntary movement. Furthermore, we could show that these projection neurons significantly downregulate the expression of BDNF during the juvenile development of mice between 3 and 12 weeks. In parallel, we found a modulatory effect of dopamine on the translocation of TrkB to the cell surface in postsynaptic striatal Medium Spiny Neurons (MSNs). In MSNs of the direct pathway (dMSNs), which express dopamine receptor 1 (DRD1), we observed the formation of TrkB aggregates in the 6-hydroxydopamine (6-OHDA) model of PD. This suggests that DRD1 activity controls TrkB surface expression in these neurons. In contrast, we found that DRD2 activation has opposite effects in MSNs of the indirect pathway (iMSNs). Activation of DRD2 promotes a rapid decrease in TrkB surface expression which was reversible and depended on cAMP. In parallel, stimulation of DRD2 led to induction of phospho-TrkB (pTrkB). This effect was significantly slower than the effect on TrkB surface expression and indicates that TrkB is transactivated by DRD2. Together, our data provide evidence that dopamine triggers dual modes of plasticity on striatal MSNs by acting on TrkB surface expression in DRD1 and DRD2 expressing MSNs. This surface expression of the receptor is crucial for the binding of BDNF, which is released from corticostriatal afferents. This leads to the induction of TrkB-mediated downstream signal transduction cascades and long-term potentiation (LTP). Therefore, the dopamine-mediated translocation of TrkB could be a mediator that modulates the balance between dopaminergic and glutamatergic signaling to allow synaptic plasticity in a spatiotemporal manner. This information and the fact that TrkB is segregated to persistent aggregates in PD could help to improve our understanding of voluntary movement control and to develop new therapeutic strategies beyond those focusing on dopaminergic supply.},
  subject      = {Brain-derived neurotrophic factor},
  language  = {en}
}