Browsing by Subject "CDK5"

Now showing 1 - 2 of 2
  • Thumbnail Image
    Item
    Involvement of Collapsin Response Mediator Protein 2 in Posttraumatic Sprouting in Acquired Epilepsy
    (2014) Wilson, Sarah Marie; Oxford, Gerry S.; Khanna, Rajesh; Jen, Joanna; Xu, Zao C.; Jin, Xiao-Ming
    Posttraumatic epilepsy, the development of temporal lobe epilepsy (TLE) following traumatic brain injury, accounts for 20% of symptomatic epilepsy. Reorganization of mossy fibers within the hippocampus is a common pathological finding of TLE. Normal mossy fibers project into the CA3 region of the hippocampus where they form synapses with pyramidal cells. During TLE, mossy fibers are observed to innervate the inner molecular layer where they synapse onto the dendrites of other dentate granule cells, leading to the formation of recurrent excitatory circuits. To date, the molecular mechanisms contributing to mossy fiber sprouting are relatively unknown. Recent focus has centered on the involvement of tropomycin-related kinase receptor B (TrkB), which culminates in glycogen synthase kinase 3β (GSK3β) inactivation. As the neurite outgrowth promoting collapsin response mediator protein 2 (CRMP2) is rendered inactive by GSK3β phosphorylation, events leading to inactivation of GSK3β should therefore increase CRMP2 activity. To determine the involvement of CRMP2 in mossy fiber sprouting, I developed a novel tool ((S)-LCM) for selectively targeting the ability of CRMP2 to enhance tubulin polymerization. Using (S)-LCM, it was demonstrated that increased neurite outgrowth following GSK3β inactivation is CRMP2 dependent. Importantly, TBI led to a decrease in GSK3β-phosphorylated CRMP2 within 24 hours which was secondary to the inactivation of GSK3β. The loss of GSK3β-phosphorylated CRMP2 was maintained even at 4 weeks post-injury, despite the transience of GSK3β-inactivation. Based on previous work, it was hypothesized that activity-dependent mechanisms may be responsible for the sustained loss of CRMP2 phosphorylation. Activity-dependent regulation of GSK3β-phosphorylated CRMP2 levels was observed that was attributed to a loss of priming by cyclin dependent kinase 5 (CDK5), which is required for subsequent phosphorylation by GSK3β. It was confirmed that the loss of GSK3β-phosphorylated CRMP2 at 4 weeks post-injury was likely due to decreased phosphorylation by CDK5. As TBI resulted in a sustained increase in CRMP2 activity, I attempted to prevent mossy fiber sprouting by targeting CRMP2 in vivo following TBI. While (S)-LCM treatment dramatically reduced mossy fiber sprouting following TBI, it did not differ significantly from vehicle-treated animals. Therefore, the necessity of CRMP2 in mossy fiber sprouting following TBI remains unknown.
  • Thumbnail Image
    Item
    Transcription factor regulation of amyloid-beta pathway genes by SP1-Modulating compounds : a novel approach in Alzheimer's Disease
    (2017-07-07) Bayon, Baindu L.; Lahiri, Debomoy K.; Herbert, Brittney-Shea; Nho, Kwangsik; Nurnberger, John I.; Zhou, Feng C.
    Alzheimer's disease (AD) is a neurodegenerative disorder characterized by the presence of neuritic plaques consisting of extracellular amyloid-beta (Aβ) and neurofibrillary tangles comprised of hyperphosphorylated microtubule associated tau. Aβ is produced following the cleavage of amyloid precursor protein (APP) by the enzyme BACE1. Transcription factors (TFs) are proteins involved in the regulation of gene transcription. Expression levels of some TFs are perturbed in AD. SP1 binding sites on both the APP and BACE1 promoters implicate its potential role in AD. Aβ peptide itself mediates activation of cyclindependent kinase 5 (CDK5), an enzyme which phosphorylates the FOXO (Forkhead Box) TFs. In order to study mechanisms of TF regulation of Aβ production in human models, neuronally differentiated cells as well as a primary human neurosphere culture were used to test the effects of TF-modulating compounds. Our hypothesis is that by targeting relevant TFs via pharmacological inhibitors in human cells, BACE1 activity or APP expression will decrease and Aβ production will be reduced as a result. To test the involvement of TFs in the regulation of APP, we treated several mammalian cells lines and post-mitotic human neuronal cells with roscovitine, mithramycin A (MTM), MTM analogs (MTM-SDK, MTM-SK), and tolfenamic acid (TA). MTM and TA treatment of neurons differentially activated several TFs implicated in AD. Treatment of differentiated neurospheres with MTM led to a significant decrease in APP and SP1 expression along with Aβ40 levels. Epigenetic mechanisms involve alteration of the binding affinity between DNA and transcription factors. We predict that modulation of these TFs may be influenced by epigenetic modifications. To test the effects of drugs on epigenetic markers, histone deacetylase (HDAC) and DNA methyltransferase (DNMT) activity was measured. MTM-SDK significantly decreased DNMT activity in differentiated neuroblastoma cells, this may enhance or decrease the ability of SP1 to bind to target DNA and affect transcription of BACE1 or APP. Targeting TF activity is a novel means to manipulate the amyloid pathway. Compounds modifying TF binding to sites on the BACE1 or APP promoters may provide a means to limit the production of amyloid-beta and slow the symptoms of AD.