Amy K.Y. Fu

Cdk5 regulates EphA4-mediated dendritic spine retraction through an ephexin1-dependent mechanism.

The development of dendritic spines is thought to be crucial for synaptic plasticity. Dendritic spines are retracted upon Eph receptor A4 (EphA4) activation, but the mechanisms that control this process are not well understood. Here we report an important function of cyclin-dependent kinase 5 (Cdk5) in EphA4-dependent spine retraction in mice. We found that blocking Cdk5 activity inhibits ephrin-A1–triggered spine retraction and reduction of mEPSC frequency at hippocampal synapses. The activation of EphA4 resulted in the recruitment of Cdk5 to EphA4, leading to the tyrosine phosphorylation and activation of Cdk5. EphA4 and Cdk5 then enhanced the activation of ephexin1, a guanine-nucleotide exchange factor that regulates activation of the small Rho GTPase RhoA. The association between EphA4 and ephexin1 was significantly reduced in Cdk5−/− brains and Cdk5-dependent phosphorylation of ephexin1 was required for the ephrin-A1–mediated regulation of spine density. These findings suggest that ephrin-A1 promotes EphA4-dependent spine retraction through the activation of Cdk5 and ephexin1, which in turn modulates actin cytoskeletal dynamics.

Regulation of NMDA Receptors by Neuregulin Signaling in Prefrontal Cortex

Recent linkage studies have identified a significant association of the neuregulin gene with schizophrenia, but how neuregulin is involved in schizophrenia is primarily unknown. Aberrant NMDA receptor functions have been implicated in the pathophysiology of schizophrenia. Therefore, we hypothesize that neuregulin, which is present in glutamatergic synaptic vesicles, may affect NMDA receptor functions via actions on its ErbB receptors enriched in postsynaptic densities, hence participating in emotional regulation and cognitive processes that are impaired in schizophrenia. To test this, we examined the regulation of NMDA receptor currents by neuregulin signaling pathways in prefrontal cortex (PFC), a prominent area affected in schizophrenia. We found that bath perfusion of neuregulin significantly reduced whole-cell NMDA receptor currents in acutely isolated and cultured PFC pyramidal neurons and decreased NMDA receptor-mediated EPSCs in PFC slices. The effect of neuregulin was mainly blocked by application of the ErbB receptor tyrosine kinase inhibitor, phospholipase C (PLC) inhibitor, IP3 receptor (IP3R) antagonist, or Ca2+ chelators. The neuregulin regulation of NMDA receptor currents was also markedly attenuated in cultured neurons transfected with mutant forms of Ras or a dominant-negative form of MEK1 (mitogen-activated protein kinase kinase 1). Moreover, the neuregulin effect was prevented by agents that stabilize or disrupt actin polymerization but not by agents that interfere with microtubule assembly. Furthermore, neuregulin treatment increased the abundance of internalized NMDA receptors in cultured PFC neurons, which was also sensitive to agents affecting actin cytoskeleton. Together, our study suggests that both PLC/IP3R/Ca2+ and Ras/MEK/ERK (extracellular signal-regulated kinase) signaling pathways are involved in the neuregulin-induced reduction of NMDA receptor currents, which is likely through enhancing NR1 internalization via an actin-dependent mechanism.

Synaptic Roles of Cdk5: Implications in Higher Cognitive Functions and Neurodegenerative Diseases

Accumulating evidence indicates that cyclin dependent kinase 5 (Cdk5), through phosphorylating a plethora of pre- and postsynaptic proteins, functions as an essential modulator of synaptic transmission. Recent advances in the field of Cdk5 research have not only consolidated the in vivo importance of Cdk5 in neurotransmission but also suggest a pivotal role of Cdk5 in the regulation of higher cognitive functions and neurodegenerative diseases. In this review, we will discuss the recent findings on the emanating role of Cdk5 as a regulator of synaptic functions and plasticity.

Cdk5 is involved in neuregulin-induced AChR expression at the neuromuscular junction

Here we describe an important involvement of Cdk5/p35 in regulating the gene expression of acetylcholine receptor (AChR) at the neuromuscular synapse. Cdk5 and p35 were prominently expressed in embryonic muscle, and concentrated at the neuromuscular junction in adulthood. Neuregulin increased the p35-associated Cdk5 kinase activity in the membrane fraction of cultured C2C12 myotubes. Co-immunoprecipitation studies revealed the association between Cdk5, p35 and ErbB receptors in muscle and cultured myotubes. Inhibition of Cdk5 activity not only blocked the NRG-induced AChR transcription, but also attenuated ErbB activation in cultured myotubes. In light of our finding that overexpression of p35 alone led to an increase in AChR promoter activity in muscle, Cdk5 activation is sufficient to mediate the up-regulation of AChR gene expression. Taken together, these results reveal the unexpected involvement of Cdk5/p35 in neuregulin signaling at the neuromuscular synapse.

APCCdh1 mediates EphA4-dependent downregulation of AMPA receptors in homeostatic plasticity

Homeostatic plasticity is crucial for maintaining neuronal output by counteracting unrestrained changes in synaptic strength. Chronic elevation of synaptic activity by bicuculline reduces the amplitude of miniature excitatory postsynaptic currents (mEPSCs), but the underlying mechanisms of this effect remain unclear. We found that activation of EphA4 resulted in a decrease in synaptic and surface GluR1 and attenuated mEPSC amplitude through a degradation pathway that requires the ubiquitin proteasome system (UPS). Elevated synaptic activity resulted in increased tyrosine phosphorylation of EphA4, which associated with the ubiquitin ligase anaphase-promoting complex (APC) and its activator Cdh1 in neurons in a ligand-dependent manner. APCCdh1 interacted with and targeted GluR1 for proteasomal degradation in vitro, whereas depletion of Cdh1 in neurons abolished the EphA4-dependent downregulation of GluR1. Knockdown of EphA4 or Cdh1 prevented the reduction in mEPSC amplitude in neurons that was a result of chronic elevated activity. Our results define a mechanism by which EphA4 regulates homeostatic plasticity through an APCCdh1-dependent degradation pathway.

Two Cyclin-Dependent Kinase Pathways Are Essential for Polarized Trafficking of Presynaptic Components

Polarized trafficking of synaptic proteins to axons and dendrites is crucial to neuronal function. Through forward genetic analysis in C. elegans, we identified a cyclin (CYY-1) and a cyclin-dependent Pctaire kinase (PCT-1) necessary for targeting presynaptic components to the axon. Another cyclin-dependent kinase, CDK-5, and its activator p35, act in parallel to and partially redundantly with the CYY-1/PCT-1 pathway. Synaptic vesicles and active zone proteins mostly mislocalize to dendrites in animals defective for both PCT-1 and CDK-5 pathways. Unlike the kinesin-3 motor, unc-104/Kif1a mutant, cyy-1 cdk-5 double mutants have no reduction in anterogradely moving synaptic vesicle precursors (SVPs) as observed by dynamic imaging. Instead, the number of retrogradely moving SVPs is dramatically increased. Furthermore, this mislocalization defect is suppressed by disrupting the retrograde motor, the cytoplasmic dynein complex. Thus, PCT-1 and CDK-5 pathways direct polarized trafficking of presynaptic components by inhibiting dynein-mediated retrograde transport and setting the balance between anterograde and retrograde motors.

α2-Chimaerin interacts with EphA4 and regulates EphA4-dependent growth cone collapse

EphA4-dependent growth cone collapse requires reorganization of actin cytoskeleton through coordinated activation of Rho family GTPases. Whereas various guanine exchange factors have recently been identified to be involved in EphA4-mediated regulation of Rho GTPases and growth cone collapse, the functional roles of GTPase-activating proteins in the process are largely unknown. Here we report that EphA4 interacts with α2-chimaerin through its Src homology 2 domain. Activated EphA4 induces a rapid increase of tyrosine phosphorylation of α2-chimaerin and enhances its GTPase-activating protein activity toward Rac1. More importantly, α2-chimaerin regulates the action of EphA4 in growth cone collapse through modulation of Rac1 activity. Our findings have therefore identified a new α2-chimaerin-dependent signaling mechanism through which EphA4 transduces its signals to the actin cytoskeleton and modulates growth cone morphology.

Tyk2/STAT3 Signaling Mediates β-Amyloid-Induced Neuronal Cell Death: Implications in Alzheimer's Disease

One of the pathological hallmarks of Alzheimers disease (AD) is deposition of extracellular amyloid-β (Aβ) peptide, which is generated from the cleavage of amyloid precursor protein (APP). Accumulation of Aβ is thought to associate with the progressive neuronal death observed in AD. However, the precise signaling mechanisms underlying the action of Aβ in AD pathophysiology are not completely understood. Here, we report the involvement of the transcription factor signal transducer and activator of transcription 3 (STAT3) in mediating Aβ-induced neuronal death. We find that tyrosine phosphorylation of STAT3 is elevated in the cortex and hippocampus of APP/PS1 transgenic mice. Treatment of cultured rat neurons with Aβ or intrahippocampal injection of mice with Aβ both induces tyrosine phosphorylation of STAT3 in neurons. Importantly, reduction of either the expression or activation of STAT3 markedly attenuates Aβ-induced neuronal apoptosis, suggesting that STAT3 activation contributes to neuronal death after Aβ exposure. We further identify Tyk2 as the tyrosine kinase that acts upstream of STAT3, as Aβ-induced activation of STAT3 and caspase-3-dependent neuronal death can be inhibited in tyk2−/− neurons. Finally, increased tyrosine phosphorylation of STAT3 is also observed in postmortem brains of AD patients. Our observations collectively reveal a novel role of STAT3 in Aβ-induced neuronal death and suggest the potential involvement of Tyk2/STAT3 signaling in AD pathophysiology.

Expression of Eph receptors in skeletal muscle and their localization at the neuromuscular junction

The participation of ephrins and Eph receptors in guiding motor axons during muscle innervation has been well documented, but little is known about their expression and functional significance in muscle at later developmental stages. Our present study investigates the expression and localization of Eph receptors and ephrins in skeletal muscle. Prominent expression of EphA4, EphA7, and ephrin-A ligands was detected in muscle during embryonic development. More importantly, both EphA4 and EphA7, as well as ephrin-A2, were localized at the neuromuscular junction (NMJ) of adult muscle. Despite their relative abundance, they were not localized at the synapses during embryonic stages. The concentration of EphA4, EphA7, and ephrin-A2 at the NMJ was observed at postnatal stages and the synaptic localization became prominent at later developmental stages. In addition, expression of Eph receptors was increased by neuregulin and after nerve injury. Furthermore, we demonstrated that overexpression of EphA4 led to tyrosine phosphorylation of the actin-binding protein cortactin and that EphA4 was coimmunoprecipitated with cortactin in muscle. Taken together, our findings indicate that EphA4 is associated with the actin cytoskeleton. Since actin cytoskeleton is critical to the formation and stability of NMJ, the present findings raise the intriguing possibility that Eph receptors may have a novel role in NMJ formation and/or maintenance.

α2-chimaerin controls neuronal migration and functioning of the cerebral cortex through CRMP-2

Disrupted cortical neuronal migration is associated with epileptic seizures and developmental delay. However, the molecular mechanism by which disruptions of early cortical development result in neurological symptoms is poorly understood. Here we report α2-chimaerin as a key regulator of cortical neuronal migration and function. In utero suppression of α2-chimaerin arrested neuronal migration at the multipolar stage, leading to accumulation of ectopic neurons in the subcortical region. Mice with such migration defects showed an imbalance between excitation and inhibition in local cortical circuitry and greater susceptibility to convulsant-induced seizures. We further show that α2-chimaerin regulates bipolar transition and neuronal migration through modulating the activity of CRMP-2, a microtubule-associated protein. These findings establish a new α2-chimaerin-dependent mechanism underlying neuronal migration and proper functioning of the cerebral cortex and provide insights into the pathogenesis of seizure-related neurodevelopmental disorders.