Shin Jung Kang

Caspase-11 regulates cell migration by promoting Aip1-Cofilin-mediated actin depolymerization

Coordinated regulation of cell migration, cytokine maturation and apoptosis is critical in inflammatory responses. Caspases, a family of cysteine proteases, are known to regulate cytokine maturation and apoptosis. Here, we show that caspase-11, a mammalian pro-inflammatory caspase, regulates cell migration during inflammation. Caspase-11-deficient lymphocytes exhibit a cell-autonomous migration defect in vitro and in vivo. We demonstrate that caspase-11 interacts physically and functionally with actin interacting protein 1 (Aip1), an activator of cofilin-mediated actin depolymerization. The caspase-recruitment domain (CARD) of caspase-11 interacts with the carboxy-terminal WD40 propeller domain of Aip1 to promote cofilin-mediated actin depolymerization. Cells with Aip1 or caspase-11 deficiency exhibit defects in actin dynamics. Using in vitro actin depolymerization assays, we found that caspase-11 and Aip1 work cooperatively to promote cofilin-mediated actin depolymerization. These data demonstrate a novel cell autonomous caspase-mediated mechanism that regulates actin dynamics and mammalian cell migration distinct from the receptor mediated Rho–Rac–Cdc42 pathway.

Superoxide dismutase 1 mutants related to amyotrophic lateral sclerosis induce endoplasmic stress in neuro2a cells

One of the common features of damaged neurons in many neurodegenerative diseases is the presence of abnormal aggregates of the disease‐related proteins. In amyotrophic lateral sclerosis (ALS) of both sporadic and familial forms, protein aggregates are found in the affected spinal cords. In familial ALS with mutations in copper–zinc superoxide dismutase 1 (SOD1), the propensity of SOD1 for aggregation is known to increase with the mutation. In the present study, we examined whether the aggregate‐prone SOD1 mutants induce endoplasmic reticulum (ER) stress and the inhibition of the ER stress protects the cells. The ALS‐related mutant G85R SOD1 and G93A SOD1 formed visible aggregates and caused cell death possibly by apoptosis when over‐expressed in neuro2a cells. Interestingly, the rate of the mutant SOD1‐induced cell death was greater than that of the visible aggregate formation. Expression of the mutant SOD1 caused signs of both early and late ER stress responses, namely, RNA‐dependent protein kinase‐like ER kinase and eukaryotic initiation factor α phosphorylation, Jun amino‐terminal kinase activation, activating transcription factor 6‐translocation, X‐box binding protein 1 mRNA splicing, and caspase 12 activation. The X‐box binding protein 1 mRNA splicing activation was also detected in the mutant SOD1‐expressing cells even without the visible aggregates. The cell death induced by the mutant SOD1 over‐expression looked like apoptosis as evidenced by nuclear morphology and terminal deoxynucleotidyl transferase‐mediated deoxyuridine triphosphate (dUTP) nick end labeling. Importantly, an ER stress inhibitor, salubrinal delayed the formation of insoluble aggregates of the mutant SOD1 and suppressed the mutant‐induced cell death. In addition, over‐expression of the ER‐targeted Bcl‐xL protected the cells from the mutant SOD1‐induced cytotoxicity. These results suggest that the misfolding of ALS‐related mutant SOD1 induces ER stress possibly prior to the formation of visible aggregates, which may contribute to the motor neuron degeneration in ALS pathogenesis.

(ADP-ribose) polymerase 1 and AMP-activated protein kinase mediate progressive dopaminergic neuronal degeneration in a mouse model of Parkinson's disease

Genetic and epidemiologic evidence suggests that cellular energy homeostasis is critically associated with Parkinson’s disease (PD) pathogenesis. Here we demonstrated that genetic deletion of Poly (ADP-ribose) polymerase 1 completely blocked 6-hydroxydopamine-induced dopaminergic neurodegeneration and related PD-like symptoms. Hyperactivation of PARP-1 depleted ATP pools in dopaminergic (DA) neurons, thereby activating AMP-activated protein kinase (AMPK). Further, blockade of AMPK activation by viral infection with dominant-negative AMPK strongly inhibited DA neuronal atrophy with moderate suppression of nuclear translocation of apoptosis-inhibiting factor (AIF), whereas overactivation of AMPK conversely strengthened the 6-OHDA-induced DA neuronal degeneration. Collectively, these results suggest that manipulation of PARP-1 and AMPK signaling is an effective therapeutic approach to prevent PD-related DA neurodegeneration.

Resveratrol upregulated heat shock proteins and extended the survival of G93A-SOD1 mice.

In the present study, we investigated whether resveratrol, a SIRT1 activator, can suppress the motor neuron degeneration in a transgenic mouse model of amyotrophic lateral sclerosis. Chronic intraperitoneal injection of resveratrol delayed the disease onset and extended survival of the transgenic mice overexpressing G93A-SOD1. The number of surviving motor neurons increased in the resveratrol-injected G93A mice. Importantly, the levels of Hsp25 and Hsp70 were elevated while the level of heat shock factor 1 (HSF1) acetylation decreased in the spinal cords of the resveratrol-injected G93A mice. Our data suggest that resveratrol may protect motor neurons from the mutant SOD1-induced neurotoxicity by promoting SIRT1-mediated deacetylation of HSF1 and subsequent upregulation of Hsps.

Human α-synuclein modulates vesicle trafficking through its interaction with prenylated Rab acceptor protein 1

α-Synuclein has been implicated in the pathogenesis of Parkinsons disease. Although it is highly conserved, its physiological function has not yet been elucidated in detail. In an effort to define the function of α-synuclein, interacting proteins were screened in phage display assays. Prenylated Rab acceptor protein 1 (PRA1) was identified as an interacting partner. A selective interaction between α-synuclein and PRA1 was confirmed by coimmunoprecipitation and GST pull-down assays. PRA1 and α-synuclein were colocalized in N2a neuronal cells. Cotransfection of α-synuclein and PRA1 caused vesicles to accumulate in the periphery of the cytosol in neuronal cells, suggesting that overexpression of α-synuclein hinders proper vesicle trafficking and recycling as a result of the interaction between α-synuclein and PRA1.

AIF translocates to the nucleus in the spinal motor neurons in a mouse model of ALS

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease characterized by selective loss of motor neurons in the brain stem and the spinal cords. One of the causes for the familial ALS has been attributed to the mutations in copper-zinc superoxide dismutase (SOD1). Although the toxic function of the mutant enzyme has not been fully understood, the final cell death pathway has been suggested as caspase-dependent. In the present study, we present evidence that the activation of apoptosis inducing factor (AIF) may play a role to induce motor neuron death during ALS pathogenesis. In the spinal cord of SOD1 G93A transgenic mice, expression of AIF was detected in the motor neurons and astrocytes. The level of AIF expression increased as the disease progressed. In the symptomatic SOD1 G93A transgenic mice, AIF released from the mitochondria and translocated into the nucleus in the motor neurons as evidenced by confocal microscopy and biochemical analysis. These results suggest that AIF may play a role to induce motor neuron death in a mouse model of ALS.

HCN channel activity-dependent modulation of inhibitory synaptic transmission in the rat basolateral amygdala

Hyperpolarization-activated cyclic nucleotide-gated (HCN) channels are expressed in the central nervous system and play a regulatory role in neuronal excitability. In the present study, we examined a physiological role of HCN channels in the rat basolateral amygdala (BLA). In vitro electrophysiological studies showed that ZD7288 decreased spontaneous inhibitory postsynaptic current (sIPSC) without changing miniature IPSC (mIPSC). HCN channel blockade also attenuated feedback inhibitions in BLA principal neurons. However, blockade of HCN channel had little effects on spontaneous excitatory postsynaptic current (sEPSC) and mEPSC. Therefore, HCN channel appeared to decrease BLA excitability by increasing the action potential-dependent inhibitory control over the BLA principal neurons. Anxiety is reported to be influenced by neuronal excitability in the BLA and inhibitory synaptic transmission is thought to play a pivotal role in regulating overall excitability of the amygdala. As expected, blockade of HCN channels by targeted injection of ZD7288 to the BLA increased anxiety-like behavior under elevated plus maze test. Our results suggest that HCN channel activity can modulate the GABAergic synaptic transmission in the BLA, which in turn control the amygdala-related emotional behaviors such as anxiety.

Heptachlor induced nigral dopaminergic neuronal loss and Parkinsonism-like movement deficits in mice.

Epidemiological studies have suggested an association between pesticide exposure and Parkinsons disease. In this study, we examined the neurotoxicity of an organochlorine pesticide, heptachlor, in vitro and in vivo. In cultured SH-SY5Y cells, heptachlor induced mitochondria-mediated apoptosis. When injected into mice intraperitoneally on a subchronic schedule, heptachlor induced selective loss of dopaminergic neurons in the substantia nigra pars compacta. In addition, the heptachlor injection induced gliosis of microglia and astrocytes selectively in the ventral midbrain area. When the general locomotor activities were monitored by open field test, the heptachlor injection did not induce any gross motor dysfunction. However, the compound induced Parkinsonism-like movement deficits when assessed by a gait and a pole test. These results suggest that heptachlor can induce Parkinsons disease-related neurotoxicities in vivo.

Suppression of caspase-11 expression by histone deacetylase inhibitors

It has been well documented that histone deacetylase inhibitors suppress inflammatory gene expression. Therefore, we investigated whether histone deacetylase inhibitors modulate the expression of caspase-11 that is known as an inducible caspase regulating both inflammation and apoptosis. In the present study, we show that sodium butyrate and trichostatin A, two structurally unrelated inhibitors of histone deacetylase (HDAC), effectively suppressed the induction of caspase-11 in mouse embryonic fibroblasts stimulated with lipopolysaccharides. Sodium butyrate inhibited the activation of upstream signaling events for the caspase-11 induction such as activation of p38 mitogen-activated protein kinase and c-Jun N-terminal kinase, degradation of inhibitor of kappaB, and activation of nuclear factor-kappaB. These results suggest that the HDAC inhibitor suppressed cytosolic signaling events for the induction of caspase-11 by inhibiting the deacetylation of non-histone proteins.

β-Sheet-breaking peptides inhibit the fibrillation of human α-synuclein

alpha-Synuclein is the major components of the intracellular protein-aggregates, found in the dopaminergic neurons of Parkinsons disease patients. Previously, we screened for alpha-synuclein substitution mutants that prevent fibril formation of both wild-type and Parkinsons disease-linked alpha-synuclein variants. In the present study, we show that short synthetic peptides derived from these mutant sequences not only prevented alpha-synuclein fibrillation but also dissolved preformed alpha-synuclein aggregates in vitro. The hexapeptide PGVTAV, which was the shortest peptide that retained the ability to block alpha-synuclein fibrillation, may serve as a lead compound for the development of therapeutics for Parkinsons disease.