Trk receptors are a family of tyrosine kinases that regulates synaptic strength and plasticity in the mammalian nervous system. Trk receptors affect neuronal survival and differentiation through several signal cascades. However, the activation of these receptors also has significant effects on functional properties of neurons.
The common ligands of trk receptors are neurotrophins, a family of growth factors critical to the functioning of the nervous system. The binding of these molecules is highly specific. Each type of neurotrophin has different binding affinity toward its corresponding trk receptor. The activation of Trk receptors by neurotrophin binding may lead to activation of signal cascades resulting in promoting survival and other functional regulation of cells.
Origin of the name trk
The family of Trk receptors is named for the oncogene trk, whose identifical led to the discovery of its first member, TrkA. trk, initially identified in a colon carcinoma, is frequently (25%) activated in thyroid papillary carcinomas . The oncogene was generated by a mutation in chromosome 1 that resulted in the fusion of the first seven exons of tropomyosin to the transmembrane and cytoplasmic domains of the then-unknown TrkA receptor . Normal Trk receptors do not contain amino acid or DNA sequences related to tropomyosin.
Trk receptor types and corresponding ligands
The three most common types of trk receptors are trkA, trkB, and trkC. Each of these receptors types has different binding affinity to certain types of neurotrophins. The differences in the signaling initiated by these distinct types of receptors are important for generating diverse of biological responses.
Neurotrophin ligands of Trk receptors are processed ligands, meaning that they are synthesized in immature forms and then transformed by protease cleavage. Immature neurotrophins are specific only to one common p75NTR receptor. However, protease cleavage generates neurotrophins that have higher affinity to their corresponding Trk receptors. These processed neurotrophins can still bind to p75NTR, but at a much lower affinity.
TrkA has the highest affinity to the binding nerve growth factor (NGF). NGF is important in both local and nuclear actions, regulating growth cones, motility, and expression of genes encoding the biosynthesis enzymes for neurotransmitters. Nocireceptive sensory neurons express mostly trkA and not trkB or trkC.
TrkB has the highest affinity to the binding brain-derived neurotrophic factor (BDNF) and NT-4. BDNF is growth factor that has important roles in the survival and function of neurons in the central nervous system. The binding of BDNF to TrkB receptor causes many intercellular cascades be activated, which regulate neuronal development and plasticity, long-term potentiation, and apoptosis.
Although both BDNF and NT-4 have high specificity to TrkB, they are not interchangeable. In a mouse model study where BDNF expression was replaced by NT-4, the mouse with NT4 expression appeared to be smaller and exhibited decreased fertility.
TrkC is ordinarily activated by binding with NT-3 and has little activation by other ligands. (TrkA and TrkB also bind NT-3, but to a lesser extent.) TrkC is mostly expressed by proprioceptive sensory neurons. The axons of these proprioceptive sensory neurons are much thicker than those of nocireceptive sensory neurons, which express trkA.
Regulation by P75NTR
P75NTR increases binding affinity and specificity for Trk receptor
p75NTR (p75 neurotrophin receptor) affects the binding affinity and specificity of Trk receptor activation by neurotrophins. The presence of p75NTR is especially important in increasing the binding affinity of NGF to TrkA. Although the dissociation constants of p75NTR and TrkA are remarkably similar, their kinetics are quite different. Reduction and mutation of cytoplasmic and transmembrane domains of either TrkA or P75NTR prevent the formation of high-affinity binding sites on TrkA. However, the binding of ligands in p75NTR is not required to promote high-affinity binding. Therefore, the data suggest that the presence of p75NTR affects the conformation of TrkA, preferentially the state with high-affinity binding site for NGF. Surprisingly, although the presence of p75NTR is essential to promote high-affinity binding, the NT3 binding to the receptor is not required.
Apart from affecting the affinity and specificity for Trk receptor, P75 neurotropins receptor (P75NTR) can also reduce ligand-induced receptor ubiquitination, delays receptor internalization, and degradation. In an experiment by Makkerh in 2005, both TrkA and TrkB receptors were tested using similar method. The only difference is that NGF was used to activate the TrkA receptor whereas BDNF was used for TrkB receptor.
TrkA and TrkB receptors activity are ligand-dependant
To test if TrkA and TrkB receptors activity are ligand-dependant, the receptors are treated with NGF and BDNF correspondingly. In vivo, the activity of these receptors is regulated by ubiquitination, the process in which protein undergoes modification by attachment of ubiquitin monomers. These molecules usually have main function of labeling proteins for proteasomal degradation. After being activated, Trk receptors may undergo endocytosis and will be either recycled or degraded. Two types of cells were used to test the protocol, P12 cells and E16 mouse cortices. An E16 mouse cortex was used in conjunction with BDNF, since it expresses abundant TrkB and lesser p75NTR. On the other hands, P12 cells were treated with NGF, since it expresses more TrkA and p75NTR but not TrkB and TrkC. The treated receptors were then immunoprecipitated and immunoblotted to be analyzed. The result shows that TrkB phosphotyrosine (activated TrkB) content increased rapidly with the presence of BDNF compared with control (absence of BDNF) whereas TrkA phosphotyrosine content increased modestly when treated with NGF.
p75NTR reduces Trk ubiquitination
To test if p75NTR reduces Trk ubiquitination, HEK293 (human embryonic kidney 293 cells) were co-tranfected with plasmids encoding TrkA and Myc-tagged ubiquitin with the presence and absence of plasmid encoding P25NTR. When treated with their corresponding neurotrophins, lower levels of ubiquitinated TrkA and TrkB are present in presence of p75NTR. Vice versa, in the absence of p75NTR, ubiquitination content increases decreases significantly.
p75NTR delays Trk receptor internalization
To test the hypothesis, HEK93 cells were subjected to biotinylation with the presence and absence of P25NTR. Each samples were taken at different time points (0, 2, 5, and 15 mins), immunoblotted, and screened for the level of tyrosine-phosphorylated molecules. The result shows increases in the level of tyrosine-phosphorylated molecules with the presence of p75NTR. In absence of P25NTR, tyrosine-phosphorylated level fell below detection limit in 5 min. The biotinylated protein complexes were lysed to show both tyrosine-phosphorylated molecules in the surface and those that been internalized.
Essential roles in differentiation and function
Precursor cell survival and proliferation
Numerous studies, both in vivo and in vitro, have shown that neurotrophins have proliferation and differentiation effects on CNS neuro-epithelial precursors, neural crest cells, or precursors of the enteric nervous system. TrkA that expresses NGF not only increase the survival of both C and A delta classes of nocireceptor neurons, but also affect the functional properties of these neurons.4 As mentioned before, BDNF improves the survival and function of neurons in CNS, particularly cholinergic neurons of the basal forebrain, as well as neurons in the hippocampus and cortex.
BDNF belongs to the neurotrophin family of growth factors and affects the survival and function of neurons in the central nervous system, particularly in brain regions susceptible to degeneration in AD. BDNF improves survival of cholinergic neurons of the basal forebrain, as well as neurons in the hippocampus and cortex.
Control of target innervation
Each of the neurotrophins mentioned above[vague] promotes neurite outgrowth. NGF/TrkA signaling regulates the advance of sympathetic neuron growth cones; even when neurons received adequate trophic (sustaining and nourishing) support, one experiment showed they did not grow into relating compartments without NGF.[vague] NGF increases the innervation of tissues that receive sympathetic or sensory innervation and induces aberrant innervation in tissues that are normally not innervated.
NGF/TrkA signaling upregulates BDNF, which is transported to both peripheral and central terminals of nocireceptive sensory neurons. In the periphery, TrkB/BDNF binding and TrkB/NT-4 binding acutely sensitizing nocireceptive pathway that require the presence of mast cells.
Sensory neuron function
Trk receptors and their ligands (neurotrophins) also affect neurons' functional properties. Both NT-3 and BDNF are important in the regulation and development of synapses formed between afferent neurons and motor neurons. Increased NT-3/trkC binding results in larger monosynaptic excitatory postsynaptic potentials (EPSPs) and reduced polysynaptic components. On the other hand, increased NT-3 binding to trkB to BDNF[vague] has the opposite effect, reducing the size of monosynaptic excitatory postsynaptic potentials (EPSPs) and increasing polysynaptic signaling.
Formation of ocular dominance column
In the development of mammalian visual system, axons from each eyes crosses through the lateral geniculate nucleus (LGN) and terminate in separate layers of striate cortex. However, axons from each LGN can only be driven by one side of the eye, but not both together. tThese axons that terminate in layer IV of the striate cortex result in ocular dominance columns. A study shows that The density of innervating axons in layer IV from LGN can be increased by exogenous BDNF and reduced by a scavenger of endogenous BDNF. Therefore, it raises the possibility that both of these agents are involved in some sorting mechanism that is not well comprehended yet. Previous studies with cat model has shown that monocular deprivation occurs when input to one of the mammalian eyes is absent during the critical period (critical window). However, A study demonstrated that the infusion of NT-4 (a ligand of trkB) into the visual cortex during the critical period has been shown to prevent many consequences of monocular deprivation. Surprisingly, even after losing responses during the critical period, the infusion of NT-4 has been shown to be able to restore them.
Synaptic strength and plasticity
In mammalian hippocampus, the axons of the CA3 pyramidal cells project into CA1 cells through the Schaffer collaterals. The long-term potentiation (LTP) may induce in either of these pathways, but it is specific only to the one that is stimulated with tetanus.[vague] The stimulated axon does not impact spill over to the other pathway. TrkB receptors are expressed in most of these hippocampal neurons, including dentate granule cells, CA3 and CA1 pyramidal cells, and inhibitory interneurons. Interestingly, LTP can be greatly reduced by BDNF mutants. In a similar study on a mouse mutant with reduced expression of trkB receptors, LTP of CA1 cells reduced significantly. TrkB loss has also been linked to interfere with the memory acquisition and consolidation in many learning paradigm.
Trk receptors dimerize in response to ligand, as do other tyrosine kinase receptors. These dimers phosphorylate each other and enhance catalytic activity of the kinase. Trk receptors affect neuronal growth and differentiation through the activation of different signaling cascades. The three known pathways are PLC, Ras/MAPK (mitogen-activated protein kinase) and the PI3K (phosphatidylinositol 3-kinase) pathways. These pathways involve the interception of nuclear and mitochondrial cell-death programs. These signaling cascades eventually led to the activation of a transcription factor, CREB (cAMP response element-binding), which in turn the activate target genes.
The binding of neutrophin will lead to the phosphorylation PLC by trk receptor. The phosphorylation of PLC induces an enzyme to catalyze the breakdown of lipids to diacyglycerol and inositol (1,4, 5). Diacyglycerol may indirectly activate PI3 kinase or several protein kinases C isoform, whereas inositol (1,4, 5) promotes release of calcium from intracellular stores.
The signaling through Ras/MAPK pathway is important for the neurotrophin-induced differentiation of neuronal and neuroblastoma cells. Phosphorylation of tyrosine residues in the Trk receptors led to the activation of Ras molecules, H-Ras and K-Ras. H-ras is found in lipid rafts, embedded within the plasma membrane, while K-Ras is predominantly found in disordered region of the membrane. RAP, a vesicle bounded molecule that also takes part in the cascading, is localized in the intracellular region.
The activation of these molecules result in two alternative MAP kinase pathways. Erk 1,2 can be stimulated through the activation cascades of K-Ras, Raf1, and MEK 1,2, whereas ERK 5 is stimulated through the activation cascades of B-Raf, MEK5, and Erk 5. However, whether PKC (protein kinase C) could activate MEK5 is not yet known.
PI3 pathway signaling is critical for both mediation of neurotrophin-induced survival and regulation of vesicular trafficking. The trk receptor stimulates P13 heterodimers, which causes the activation of kinases PDK-1 and Akt. Akt in turn stimulates FRK (Forkhead family transcription factor), BAD, and GSK-3.
Some studies have suggested that NGF/TrkA coupling causes preferential activation of the Ras/MAPK pathway, whereas NT3/TrkC coupling causes preferential activation of the PI3 pathway.
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