Role In Neuronal Development Research Articles

Physiological significance of proteolytic processing of Reelin revealed by cleavage-resistant Reelin knock-in mice

Reelin is a secreted protein that plays versatile roles in neuronal development and function. The strength of Reelin signaling is regulated by proteolytic processing, but its importance in vivo is not yet fully understood. Here, we generated Reelin knock-in (PA-DV KI) mice in which the key cleavage site of Reelin was abolished by mutation. As expected, the cleavage of Reelin was severely abrogated in the cerebral cortex and hippocampus of PA-DV KI mice. The amount of Dab1, whose degradation is induced by Reelin signaling, decreased in these tissues, indicating that the signaling strength of Reelin was augmented. The brains of PA-DV KI mice were largely structurally normal, but unexpectedly, the hippocampal layer was disturbed. This phenotype was ameliorated in hemizygote PA-DV KI mice, indicating that excess Reelin signaling is detrimental to hippocampal layer formation. The neuronal dendrites of PA-DV KI mice had more branches and were elongated compared to wild-type mice. These results present the first direct evidence of the physiological importance of Reelin cleavage.

Abstract P2-07-01: Kidins220 (kinase D interacting substrate 220) has a connection with neutrotrophic related factors, BDNF and NGF, in human breast cancer

Abstract Background: Kidins220 (Kinase D Interacting Substrate 220) also known as ARMS (ankyrin repeat-rich membrane spanning) is a transmembrane protein that plays a significant role in neuronal development. It acts as the common downstream effector of neuronal signalling regulated by neurotrophin family including nerve growth factor (NGF) and brain derived neurotrophic factor (BDNF). Expression abnormalities of Kidins220 have been identified in certain solid tumours, but the role played remains controversial. The present study aimed to examine the correlation between Kidins220 and BDNF, factor known to be aberrantly expressed in breast cancer cells (1,2), together with NGF and NGF receptor (BGFR). The involvement of Kidins220 in the chain event of cells response to BDNF, in particular the role played by phospholipase-gamma (PLC- γ) were also been determined. Method: Human breast cancer tissues were collected immediately after surgery. The transcription levels of Kidins220 and the neurotrophic factors and their receptors were determined using quantitative transcript analyses. The correlations between Kidins220 with BDNF, NGF, and NGFR were analysed against the clinical and pathological parameters in the clinical breast cancer cohort. Kidins220 knockdown cell models, using MDA MB-231 and MCF-7 cells, were created by employing anti-Kidins220 shRNA lentiviral technologies. Cell matrix attachment, cellular migration and cell growth were determined in breast cancer cells treated with BDNF and PLC- γ specific inhibitors and analysed using an automated cell analyser (ECIS) and crystal violet method, respectively. Result: The analyses of the clinical cohort demonstrated reduced expression of NGF and NGFR in high grade and node positive breast tumours, which is in contrast to the the reported pattern seen with BDNF expression. Furthermore, the expressions of NGF and NGFR were reduced in patients with poor prognosis and clinical outcomes. Interestingly, Kidins220 was found to have a significant negative correlation with BDNF (r=-0.429, p<0.001), but positive correlation with NGF (r=0.269, p<0.01) and NGFR (r=0.211, p<0.01). Knockdown of Kidins220 increased cellular migration in MDA-MB-231 and MCF-7 breast cells. In vitro, breast cancer cells responded to BDNF and exhibited changes in cellular migration, after knocking down Kidins220 in the cells. Loss of Kidins220 also resulted in cells which were more responsive to BDNF treatment when compared with the controls. Moreover, breast cancer cells exhibited a marked response to PLC-γ inhibitor, in the presence of BDNF, this was particularly so when they lost Kidins220. Conclusion: Our observations suggest that Kidins220 is involved in the metastasis of breast cancer, and it is a potential regulator in the response chain to neurotrophic factors including BDNF. Furthermore, it down-regulates the cells in response to BDNF, which is connected to PLC-γ.

Survival Motor Neuron Protein Participates in Mouse Germ Cell Development and Spermatogonium Maintenance.

The defective human survival motor neuron 1 (SMN1) gene leads to spinal muscular atrophy (SMA), the most common genetic cause of infant mortality. We previously reported that loss of SMN results in rapid differentiation of Drosophila germline stem cells and mouse embryonic stem cells (ESCs), indicating that SMN also plays important roles in germ cell development and stem cell biology. Here, we show that in healthy mice, SMN is highly expressed in the gonadal tissues, prepubertal spermatogonia, and adult spermatocytes, whereas low SMN expression is found in differentiated spermatid and sperm. In SMA-like mice, the growth of testis tissues is retarded, accompanied with gamete development abnormalities and loss of the spermatogonia-specific marker. Consistently, knockdown of Smn1 in spermatogonial stem cells (SSCs) leads to a compromised regeneration capacity in vitro and in vivo in transplantation experiments. In SMA-like mice, apoptosis and accumulation of the R-loop structure were significantly elevated, indicating that SMN plays a critical role in the survival of male germ cells. The present work demonstrates that SMN, in addition to its critical roles in neuronal development, participates in mouse germ cell and spermatogonium maintenance.

Structure of Drosophila melanogaster ARC1 reveals a repurposed molecule with characteristics of retroviral Gag.

The tetrapod neuronal protein ARC and its Drosophila melanogaster homolog, dARC1, have important but differing roles in neuronal development. Both are thought to originate through exaptation of ancient Ty3/Gypsy retrotransposon Gag, with their novel function relying on an original capacity for self-assembly and encapsidation of nucleic acids. Here, we present the crystal structure of dARC1 CA and examine the relationship between dARC1, mammalian ARC, and the CA protein of circulating retroviruses. We show that while the overall architecture is highly related to that of orthoretroviral and spumaretroviral CA, there are substantial deviations in both amino- and carboxyl-terminal domains, potentially affecting recruitment of partner proteins and particle assembly. The degree of sequence and structural divergence suggests that Ty3/Gypsy Gag has been exapted on two separate occasions and that, although mammalian ARC and dARC1 share functional similarity, the structures have undergone different adaptations after appropriation into the tetrapod and insect genomes.

A fluorogenic probe for tracking GSH flux in developing neurons.

Understanding GSH flux in developing neurons is prerequisite to reveal its role in neuronal development but necessiates an ultrasensitive assay. By systematically exploring key structural factors determing probe sensitivity in live cells, we developed a fluorogenic probe capable of imaging subtle GSH fluctuations in developing neurons.

Canonical Wnt Signaling Modulates the Expression of Pre- and Postsynaptic Components in Different Temporal Patterns.

Wnt ligands play critical roles in neuronal development, synapse formation, synaptic activity, and plasticity. Synaptic plasticity requires molecular remodeling of synapses, implying the expression of key synaptic components. Some studies have linked Wnt signaling activity to changes in synaptic protein levels. However, the presynaptic and postsynaptic gene expression profiles of hippocampal neurons exposed to Wnt proteins have not been studied. Hence, we treated rat cultured hippocampal neurons with recombinant Wnt3a, lithium, and the Wnt inhibitor Dkk-1 for different treatment durations and measured the mRNA and protein levels of pre- and postsynaptic components. The ligand Wnt3a promoted the differential temporal expression of genes encoding presynaptic and postsynaptic proteins. Gene expression of the presynaptic proteins Rim1, piccolo (Pclo), Erc2, Ctbp1 and Rimbp2 increased in a specific temporal pattern. Simultaneously, the mRNA and protein levels of postsynaptic components showed a different temporal expression pattern, e.g., the mRNAs for postsynaptic scaffolding components such as postsynaptic density protein-95 (PSD-95/Dlg4), Homer1 and Shank1 were temporally regulated by both Wnt3a and lithium. On the other hand, the mRNA levels of the gene encoding the protein calcium/calmodulin-dependent protein kinase IV (Camk4), canonically upregulated by Wnt, were increased. Our results suggest that Wnt signaling orchestrates expressional changes in genes encoding presynaptic and postsynaptic components, probably as part of a synaptic plasticity mechanism in neurons.

Molecular mechanism of TRPV2 channel modulation by cannabidiol.

Transient receptor potential vanilloid 2 (TRPV2) plays a critical role in neuronal development, cardiac function, immunity, and cancer. Cannabidiol (CBD), the non-psychotropic therapeutically active ingredient of Cannabis sativa, is an activator of TRPV2 and also modulates other transient receptor potential (TRP) channels. Here, we determined structures of the full-length rat TRPV2 channel in apo and CBD-bound states in nanodiscs by cryo-electron microscopy. We show that CBD interacts with TRPV2 through a hydrophobic pocket located between S5 and S6 helices of adjacent subunits, which differs from known ligand and lipid binding sites in other TRP channels. CBD-bound TRPV2 structures revealed that the S4-S5 linker plays a critical role in channel gating upon CBD binding. Additionally, nanodiscs permitted us to visualize two distinct TRPV2 apo states in a lipid environment. Together these results provide a foundation to further understand TRPV channel gating, their divergent physiological functions, and to accelerate structure-based drug design.

The Rac3 GTPase in Neuronal Development, Neurodevelopmental Disorders, and Cancer.

Rho family small guanosine triphosphatases (GTPases) are important regulators of the cytoskeleton, and are critical in many aspects of cellular and developmental biology, as well as in pathological processes such as intellectual disability and cancer. Of the three members of the family, Rac3 has a more restricted expression in normal tissues compared to the ubiquitous member of the family, Rac1. The Rac3 polypeptide is highly similar to Rac1, and orthologues of the gene for Rac3 have been found only in vertebrates, indicating the late appearance of this gene during evolution. Increasing evidence over the past few years indicates that Rac3 plays an important role in neuronal development and in tumor progression, with specificities that distinguish the functions of Rac3 from the established functions of Rac1 in these processes. Here, results highlighting the importance of Rac3 in distinct aspects of neuronal development and tumor cell biology are presented, in support of the non-redundant role of different members of the two Rac GTPases in physiological and pathological processes.

De novo GRIN variants in NMDA receptor M2 channel pore-forming loop are associated with neurological diseases.

N-methyl-D-aspartate receptors (NMDARs) mediate slow excitatory postsynaptic transmission in the central nervous system, thereby exerting a critical role in neuronal development and brain function. Rare genetic variants in the GRIN genes encoding NMDAR subunits segregated with neurological disorders. Here, we summarize the clinical presentations for 18 patients harboring 12 de novo missense variants in GRIN1, GRIN2A, and GRIN2B that alter residues in the M2 re-entrant loop, a region that lines the pore and is intolerant to missense variation. These de novo variants were identified in children with a set of neurological and neuropsychiatric conditions. Evaluation of the receptor cell surface expression, pharmacological properties, and biophysical characteristics show that these variants can have modest changes in agonist potency, proton inhibition, and surface expression. However, voltage-dependent magnesium inhibition is significantly reduced in all variants. The NMDARs hosting a single copy of a mutant subunit showed a dominant reduction in magnesium inhibition for some variants. These variant NMDARs also show reduced calcium permeability and single-channel conductance, as well as altered open probability. The data suggest that M2 missense variants increase NMDAR charge transfer in addition to varied and complex influences on NMDAR functional properties, which may underlie the patients' phenotypes.

Bcl-2 Overexpression Induces Neurite Outgrowth via the Bmp4/Tbx3/NeuroD1 Cascade in H19-7 Cells.

Bcl-2 is overexpressed in the nervous system during neural development and plays an important role in modulating cell survival. In addition to its anti-apoptotic function, it has been suggested previously that Bcl-2 might act as a mediator of neuronal differentiation. However, the mechanism by which Bcl-2 might influence neurogenesis is not sufficiently understood. In this study, we aimed to determine the non-apoptotic functions of Bcl-2 during neuronal differentiation. First, we used microarrays to analyze the whole-genome expression patterns of rat neural stem cells overexpressing Bcl-2 and found that Bcl-2 overexpression induced the expression of various neurogenic genes. Moreover, Bcl-2 overexpression increased the neurite length as well as expression of Bmp4, Tbx3, and proneural basic helix-loop-helix genes, such as NeuroD1, NeuroD2, and Mash1, in H19-7 rat hippocampal precursor cells. To determine the hierarchy of these molecules, we selectively depleted Bmp4, Tbx3, and NeuroD1 in Bcl-2-overexpressing cells. Bmp4 depletion suppressed the upregulation of Tbx3 and NeuroD1 as well as neurite outgrowth, which was induced by Bcl-2 overexpression. Although Tbx3 knockdown repressed Bcl-2-mediated neurite elaboration and downregulated NeuroD1 expression, it did not affect Bcl-2-induced Bmp4 expression. While the depletion of NeuroD1 had no effect on the expression of Bcl-2, Bmp4, or Tbx3, Bcl-2-mediated neurite outgrowth was suppressed. Taken together, these results demonstrate that Bcl-2 regulates neurite outgrowth through the Bmp4/Tbx3/NeuroD1 cascade in H19-7 cells, indicating that Bcl-2 may have a direct role in neuronal development in addition to its well-known anti-apoptotic function in response to environmental insults.

Influence of Vitamin B12 Deficiency on Autonomic Nervous System Activity in Children

: Vitamin B12 has an important role in neuronal development of children. Autonomic nervous dysfunction which can be hidden by compensating mechanisms also accompanies vitamin B12 deficiency. However, in children, there is still limited data about this effect of vitamin B12 deficiency. The aim of this study was to evaluate the autonomic activity in vitamin B12 deficient children by analyzing the heart rate variability (HRV) with Holter monitoring. HRV parameters of 37 children who were diagnosed with vitamin B12 deficiency were compared those of 25 age and sex matched healthy controls. All of the heart rate variability parameters (time-domain and frequency-domain) were reduced in the vitamin B12 deficient group compared with controls. The search for factors modifying heart rate variability yielded that serum vitamin B12 level was significantly effective on both time-domain and frequency-domain HRV parameters. A positive correlation was also found between vitamin B12 levels and the mean of the 5 minute RR interval standard deviations (SDNNi), the square root of the mean of the squared differences of two consecutive RR intervals (RMSSD), low-frequency power (LF) parameters (P < 0.05, r: 0.285, 0.262, 0.246, respectively). In conclusion, vitamin B12 deficiency can cause autonomic dysfunction in children.

Interaction specificity of clustered protocadherins inferred from sequence covariation and structural analysis

Clustered protocadherins, a large family of paralogous proteins that play important roles in neuronal development, provide an important case study of interaction specificity in a large eukaryotic protein family. A mammalian genome has more than 50 clustered protocadherin isoforms, which have remarkable homophilic specificity for interactions between cellular surfaces. A large antiparallel dimer interface formed by the first 4 extracellular cadherin (EC) domains controls this interaction. To understand how specificity is achieved between the numerous paralogs, we used a combination of structural and computational approaches. Molecular dynamics simulations revealed that individual EC interactions are weak and undergo binding and unbinding events, but together they form a stable complex through polyvalency. Strongly evolutionarily coupled residue pairs interacted more frequently in our simulations, suggesting that sequence coevolution can inform the frequency of interaction and biochemical nature of a residue interaction. With these simulations and sequence coevolution, we generated a statistical model of interaction energy for the clustered protocadherin family that measures the contributions of all amino acid pairs at the interface. Our interaction energy model assesses specificity for all possible pairs of isoforms, recapitulating known pairings and predicting the effects of experimental changes in isoform specificity that are consistent with literature results. Our results show that sequence coevolution can be used to understand specificity determinants in a protein family and prioritize interface amino acid substitutions to reprogram specific protein-protein interactions.

Mutations in PIGU Impair the Function of the GPI Transamidase Complex, Causing Severe Intellectual Disability, Epilepsy, and Brain Anomalies

The glycosylphosphatidylinositol (GPI) anchor links over 150 proteins to the cell surface and is present on every cell type. Many of these proteins play crucial roles in neuronal development and function. Mutations in 18 of the 29 genes implicated in the biosynthesis of the GPI anchor have been identified as the cause of GPI biosynthesis deficiencies (GPIBDs) in humans. GPIBDs are associated with intellectual disability and seizures as their cardinal features. An essential component of the GPI transamidase complex is PIGU, along with PIGK, PIGS, PIGT, and GPAA1, all of which link GPI-anchored proteins (GPI-APs) onto the GPI anchor in the endoplasmic reticulum (ER). Here, we report two homozygous missense mutations (c.209T>A [p.Ile70Lys] and c.1149C>A [p.Asn383Lys]) in five individuals from three unrelated families. All individuals presented with global developmental delay, severe-to-profound intellectual disability, muscular hypotonia, seizures, brain anomalies, scoliosis, and mild facial dysmorphism. Using multicolor flow cytometry, we determined a characteristic profile for GPI transamidase deficiency. On granulocytes this profile consisted of reduced cell-surface expression of fluorescein-labeled proaerolysin (FLAER), CD16, and CD24, but not of CD55 and CD59; additionally, B cells showed an increased expression of free GPI anchors determined by T5 antibody. Moreover, computer-assisted facial analysis of different GPIBDs revealed a characteristic facial gestalt shared among individuals with mutations in PIGU and GPAA1. Our findings improve our understanding of the role of the GPI transamidase complex in the development of nervous and skeletal systems and expand the clinical spectrum of disorders belonging to the group of inherited GPI-anchor deficiencies.

Magnesium Is a Key Player in Neuronal Maturation and Neuropathology.

Magnesium (Mg) is the second most abundant cation in mammalian cells, and it is essential for numerous cellular processes including enzymatic reactions, ion channel functions, metabolic cycles, cellular signaling, and DNA/RNA stabilities. Because of the versatile and universal nature of Mg2+, the homeostasis of intracellular Mg2+ is physiologically linked to growth, proliferation, differentiation, energy metabolism, and death of cells. On the cellular and tissue levels, maintaining Mg2+ within optimal levels according to the biological context, such as cell types, developmental stages, extracellular environments, and pathophysiological conditions, is crucial for development, normal functions, and diseases. Hence, Mg2+ is pathologically involved in cancers, diabetes, and neurodegenerative diseases, such as Parkinson’s disease, Alzheimer’s disease, and demyelination. In the research field regarding the roles and mechanisms of Mg2+ regulation, numerous controversies caused by its versatility and complexity still exist. As Mg2+, at least, plays critical roles in neuronal development, healthy normal functions, and diseases, appropriate Mg2+ supplementation exhibits neurotrophic effects in a majority of cases. Hence, the control of Mg2+ homeostasis can be a candidate for therapeutic targets in neuronal diseases. In this review, recent results regarding the roles of intracellular Mg2+ and its regulatory system in determining the cell phenotype, fate, and diseases in the nervous system are summarized, and an overview of the comprehensive roles of Mg2+ is provided.

Regulatory networks between Polycomb complexes and non-coding RNAs in the central nervous system.

High-throughput sequencing has facilitated the identification of many types of non-coding RNAs (ncRNAs) involved in diverse cellular processes. NcRNAs as epigenetic mediators play key roles in neuronal development, maintenance, and dysfunction by controlling gene expression at multiple levels. NcRNAs may not only target specific DNA or RNA for gene silence but may also directly interact with chromatin-modifying proteins like Polycomb group (PcG) proteins to drive orchestrated transcriptional programs. Recent significant progress has been made in characterizing ncRNAs and PcG proteins involved in transcriptional, post-transcriptional, and epigenetic regulation. More importantly, dysregulation of ncRNAs, PcG proteins, and interplay among them is closely associated with the pathogenesis of central nervous system (CNS) disorders. In this review, we focus on the interplay between ncRNAs and PcG proteins in the CNS and highlight the functional roles of the partnership during neural development and diseases.

IQSEC2-Associated Intellectual Disability and Autism.

Mutations in IQSEC2 cause intellectual disability (ID), which is often accompanied by seizures and autism. A number of studies have shown that IQSEC2 is an abundant protein in excitatory synapses and plays an important role in neuronal development as well as synaptic plasticity. Here, we review neuronal IQSEC2 signaling with emphasis on those aspects likely to be involved in autism. IQSEC2 is normally bound to N-methyl-D-aspartate (NMDA)-type glutamate receptors via post synaptic density protein 95 (PSD-95). Activation of NMDA receptors results in calcium ion influx and binding to calmodulin present on the IQSEC2 IQ domain. Calcium/calmodulin induces a conformational change in IQSEC2 leading to activation of the SEC7 catalytic domain. GTP is exchanged for GDP on ADP ribosylation factor 6 (ARF6). Activated ARF6 promotes downregulation of surface α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)-type glutamate receptors through a c-jun N terminal kinase (JNK)-mediated pathway. NMDA receptors, AMPA receptors, and PSD-95 are all known to be adversely affected in autism. An IQSEC2 transgenic mouse carrying a constitutively active mutation (A350V) shows autistic features and reduced levels of surface AMPA receptor subunit GluA2. Sec7 activity and AMPA receptor recycling are presented as two targets, which may respond to drug treatment in IQSEC2-associated ID and autism.

Both GSK-3β/CRMP2 and CDK5/CRMP2 Pathways Participate in the Protection of Dexmedetomidine Against Propofol-Induced Learning and Memory Impairment in Neonatal Rats.

Dexmedetomidine has been reported to ameliorate propofol-induced neurotoxicity in neonatal animals. However, the underlying mechanism is still undetermined. Glycogen synthase kinase-3β (GSK-3β), cycline-dependent kinase-5 (CDK5), and Rho-kinase (RhoA) pathways play critical roles in neuronal development. The present study is to investigate whether GSK-3β, CDK5, and RhoA pathways are involved in the neuroprotection of dexmedetomidine. Seven-day-old (P7) Sprague Dawley rats were anesthetized with propofol for 6 h. Dexmedetomidine at various concentrations were administered before propofol exposure. Neuroapoptosis, the neuronal proliferation, and the level of neurotransmitter in the hippocampus were evaluated. The effects of GSK-3β inhibitor SB415286, CDK5 inhibitor roscovitine, or RhoA inhibitor Y276321 on propofol-induced neurotoxicity were assessed. Propofol-induced apoptosis in the hippocampal neurons and astrocytes, inhibited neuronal proliferation in the dentate gyrus region, down-regulated the level of γ-aminobutyric acid and glutamate in the hippocampus, and impaired long-term cognitive function. These harmful effects were reduced by pretreatment with 50 μg·kg-1 dexmedetomidine. Moreover, propofol-activated GSK-3β and CDK5 pathways, but not RhoA pathway, by reducing the phosphorylation of GSK-3β (ser 9), increasing the expression of CDK5 activator P25 and increasing the phosphorylation of their target sites on collapsin response mediator protein 2 (CRMP2) shortly after exposure. These effects were reversed by pretreatment with 50 μg·kg-1 dexmedetomidine. Furthermore, SB415286 and roscovitine, not Y276321, attenuated the propofol-induced neuroapoptosis, brain cell proliferation inhibition, γ-aminobutyric acid and glutamate downregulation, and learning and memory dysfunction. Our results indicate that dexmedetomidine reduces propofol-induced neurotoxicity and neurocognitive impairment via inhibiting activation of GSK-3β/CRMP2 and CDK5/CRMP2 pathways in the hippocampus of neonatal rats.

A Short Isoform of Coagulation Factor XII mRNA Is Expressed by Neurons in the Human Brain

Coagulation factor XII (FXII) is synthesized in the liver and secreted into the circulation, where it initiates the contact activation system. Although typically thought to be restricted to the circulation, FXII protein has been found in the brain of Alzheimer's disease (AD) and multiple sclerosis patients. Moreover, activation of the contact system has been detected in the cerebrospinal fluid of these patients as well as in the brain of healthy and AD individuals. While FXII protein has been detected in the brain, its source and its potential role in brain physiology and/or pathology have not been elucidated. Using in situ hybridization, we show that a shorter FXII mRNA isoform is expressed by neurons in human brain and in the brain of FXII humanized mice, with the highest expression observed in pyramidal neurons. This shorter FXII transcript contains an open reading frame coding for the portion of FXII that spans its proline-rich and catalytic domains (FXII297–596). We show that a recombinant version of this shorter FXII protein is activated by plasma kallikrein, reciprocally activates prekallikrein, and converts pro-hepatocyte growth factor (HGF) to active HGF in vitro. HGF-Met signaling plays a role in neuronal development and survival, and its dysregulation has been implicated in neurodevelopmental disorders and neurodegeneration. Taken together, our results show that a short isoform of FXII mRNA is expressed in the brain and raise the possibility that brain-derived FXII may be involved in HGF-Met signaling in neurons.

RNA editing alterations in a multi-ethnic Alzheimer disease cohort converge on immune and endocytic molecular pathways.

Little is known about the post-transcriptional mechanisms that modulate the genetic effects in the molecular pathways underlying Alzheimer disease (AD), and even less is known about how these changes might differ across diverse populations. RNA editing, the process that alters individual bases of RNA, may contribute to AD pathogenesis due to its roles in neuronal development and immune regulation. Here, we pursued one of the first transcriptome-wide RNA editing studies in AD by examining RNA sequencing data from individuals of both African-American (AA) and non-Hispanic White (NHW) ethnicities. Whole transcriptome RNA sequencing and RNA editing analysis were performed on peripheral blood specimens from 216 AD cases (105 AA, 111 NHW) and 212 gender matched controls (105 AA, 107 NHW). 449 positions in 254 genes and 723 positions in 371 genes were differentially edited in AA and NHW, respectively. While most differentially edited sites localized to different genes in AA and NHW populations, these events converged on the same pathways across both ethnicities, especially endocytic and inflammatory response pathways. Furthermore, these differentially edited sites were preferentially predicted to disrupt miRNA binding and induce nonsynonymous coding changes in genes previously associated with AD in molecular studies, including PAFAH1B2 and HNRNPA1. These findings suggest RNA editing is an important post-transcriptional regulatory program in AD pathogenesis.

Investigating the Role of Cdk5 in Alzheimer's Disease and Type‐2 Diabetes

Alzheimer's disease (AD), a progressive, age‐related neurodegenerative disorder, affects more than 30 million people worldwide. The etiology of this disease remains unclear, although risks factors such as aging, type‐2 diabetes and obesity have been identified. Hallmarks of AD include extracellular amyloid‐b plaques and interneuronal neurofibrillary tangles of hyperphosphorylated tau protein. Our lab investigates the role of Cyclin dependent kinase 5 (Cdk5) in mammalian nervous system development using the well‐established neuronal cell model PC12. PC12 cells are derived pheochromcytoma of a rat adrenal medulla. Our early findings have shown that Cdk5 is essential for the establishment of crucial neuronal cell morphology and action. This is in good agreement with other published reports that show Cdk5 is required for proper central nervous system development and for neuronal patterning and migration. Given Cdk5's role in neuronal development and that one of its substrates is tau, we decided to explore a role for Cdk5 in neurodegeneration. Chemical inhibition of Cdk5 revealed extreme aberrations in nerve cell morphology, architecture and function in our PC12 cell line in the presence of neurotrophic factors. To characterize and quantify these physical changes, we devised a simple method to convert these qualitative results to quantitative results, allowing rigorous comparison of our data. Our findings align well with the reports of other labs demonstrating a role for Cdk5 in the formation of neurofibrillary tangles and other protein aggregation and misfolding events in neurodegenerative diseases like AD. Cdk5 has also been implicated in the differentiation of extra neural tissues, including the pancreas. Since some of the risk factors for AD includes obesity and Type 2 diabetes, we also investigated the effect of Cdk5 inhibition in pancreatic cell models. Our findings suggest a role for Cdk5 in insulin secretion, supporting published reports of a role for Cdk5 in exocytosis in the pancreas. Recent evidence that deregulation of brain insulin plays a role in the pathology of AD may explain the associated risk factors of obesity and Type 2 diabetes with AD. Therefore, we are exploring the neuronal phenotype under hyperglycemic conditions while manipulating Cdk5 activity. We are extending our research using another well‐known neuronal cell model, N2a, a murine neuroblastoma. Here we report our test results of the impact of normal and aberrant Cdk5 activity on the response of differentiated N2a cells to varying concentrations of glucose in culture. This has allowed us to draw broader conclusions regarding the role of Cdk5 in neuronal development and degeneration and the connection between type II diabetes and AD. Since a number of reports suggest that dysfunction in cerebral glucose metabolism are early abnormalities in AD, this model system provides a means to examine the potential role of Cdk5 as a link between AD and Type II diabetes.Support or Funding InformationThe Rose M. Badgely Charitable Trust, Joseph Alexander Foundation, Sister Colette Mahoney Science Research Grant and the Marymount Manhattan College Faculty Award.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.