DEG/ENaC channels: a touchy superfamily that watches its salt

Summary To the surprise of many, studies of molecular mechanisms of touch transduction and analyses of epithelial Na ϩ transport have converged to define a new class of ion channel subunits. Based on the names of the first two identified subfamilies, the Caenorhabditis elegans degenerins and the vertebrate epithelial amiloridesensitive Na ϩ channel, this ion channel class is called the DEG/ENaC superfamily. Members of the DEG/ENaC superfamily have been found in nematodes, flies, snails, and vertebrates. Family members share common topology, such that they span the membrane twice and have intracellular N-and C-termini; a large extracellular loop includes a conserved cysteine-rich region. DEG/ENaC channels have been implicated a broad spectrum of cellular functions, including mechanosensation, proprioception, pain sensation, gametogenesis, and epithelial Na ϩ transport. These channels exhibit diverse gating properties, ranging from near constitutive opening to rapid inactivation. We discuss working understanding of DEG/ENaC functions, channel properties, structure/activity correlations and possible evolutionary relationship to other channel classes. BioEssays 21:568-578, 1999. 1999 John Wiley & Sons, Inc. Introduction What do touch sensation, coordination, gametogenesis, pain perception, and blood pressure regulation have in common? Recent investigations indicate that each of these fundamentally important processes is mediated by members of the DEGenerin/Epithelial Na ϩ Channel (DEG/ENaC) superfamily. Members of the DEG/ENaC superfamily have been identified from nematodes, snails, flies, and several vertebrates, including humans, and are expressed in tissues as diverse as kidney epithelia, muscle, and neurons. Although DEG/ENaC channels participate in strikingly diverse biological processes and are activated in response to markedly distinct stimuli, the overall secondary structure of channe

The Insulin/IGF Signaling Regulators Cytohesin/GRP-1 and PIP5K/PPK-1 Modulate Susceptibility to Excitotoxicity in C. elegans

During ischemic stroke, malfunction of excitatory amino acid transporters and reduced synaptic clearance causes accumulation of Glutamate (Glu) and excessive stimulation of postsynaptic neurons, which can lead to their degeneration by excitotoxicity. The balance between cell death-promoting (neurotoxic) and survival-promoting (neuroprotective) signaling cascades determines the fate of neurons exposed to the excitotoxic insult. The evolutionary conserved Insulin/IGF Signaling (IIS) cascade can participate in this balance, as it controls cell stress resistance in nematodes and mammals. Blocking the IIS cascade allows the transcription factor FoxO3/DAF-16 to accumulate in the nucleus and activate a transcriptional program that protects cells from a range of insults. We study the effect of IIS cascade on neurodegeneration in a C. elegans model of excitotoxicity, where a mutation in a central Glu transporter (glt-3) in a sensitizing background causes Glu-Receptor –dependent neuronal necrosis. We expand our studies on the role of the IIS cascade in determining susceptibility to excitotoxic necrosis by either blocking IIS at the level of PI3K/AGE-1 or stimulating it by removing the inhibitory effect of ZFP-1 on the expression of PDK-1. We further show that the components of the Cytohesin/GRP-1, Arf, and PIP5K/PPK-1 complex, known to regulate PIP2 production and the IIS cascade, modulate nematode excitotoxicity: mutations that are expected to reduce the complex’s ability to produce PIP2 and inhibit the IIS cascade protect from excitotoxicity, while overstimulation of PIP2 production enhances neurodegeneration. Our observations therefore affirm the importance of the IIS cascade in determining the susceptibility to necrotic neurodegeneration in nematode excitotoxicity, and demonstrate the ability of Cytohesin/GRP-1, Arf, and PIP5K/PPK-1 complex to modulate neuroprotection

Death Associated Protein Kinase (DAPK) -mediated neurodegenerative mechanisms in nematode excitotoxicity

Background: Excitotoxicity (the toxic overstimulation of neurons by the excitatory transmitter Glutamate) is a central process in widespread neurodegenerative conditions such as brain ischemia and chronic neurological diseases. Many mechanisms have been suggested to mediate excitotoxicity, but their significance across diverse excitotoxic scenarios remains unclear. Death Associated Protein Kinase (DAPK), a critical molecular switch that controls a range of key signaling and cell death pathways, has been suggested to have an important role in excitotoxicity. However, the molecular mechanism by which DAPK exerts its effect is controversial. A few distinct mechanisms have been suggested by single (sometimes contradicting) studies, and a larger array of potential mechanisms is implicated by the extensive interactome of DAPK. Results: Here we analyze a well-characterized model of excitotoxicity in the nematode C. elegans to show that DAPK is an important mediator of excitotoxic neurodegeneration across a large evolutionary distance. We further show that some proposed mechanisms of DAPK’s action (modulation of synaptic strength, involvement of the DANGER-related protein MAB-21, and autophagy) do not have a major role in nematode excitotoxicity. In contrast, Pin1/PINN-1 (a DAPK interaction-partner and a peptidyl-prolyl isomerase involved in chronic neurodegenerative conditions) suppresses neurodegeneration in our excitotoxicity model. Conclusions: Our studies highlight the prominence of DAPK and Pin1/PINN-1 as conserved mediators of cell death processes in diverse scenarios of neurodegeneration

FOXO3a is broadly neuroprotective in vitro and in vivo against insults implicated in motor neuron diseases

Aging is a risk factor for the development of adult-onset neuro-degenerative diseases. While some of the molecular pathways regulating longevity and stress resistance in lower organisms are defined (i.e., those activating the transcriptional regulators DAF-16 and HSF-1 in C. elegans), their relevance to mammals and disease susceptibility are unknown. We studied the signaling controlled by the mammalian homolog of DAF-16, FOXO3a, in model systems of motor neuron disease. Neuron death elicited in vitro by excitotoxic insult or the expression of mutant SOD1, mutant p150(glued) or polyQ expanded androgen receptor was abrogated by expression of nuclear-targeted FOXO3a. We identify a compound (Psammaplysene A, PA) that increases nuclear localization of FOXO3a in vitro and in vivo and show that PA also protects against these insults in vitro. Administration of PA to invertebrate model systems of neurodegeneration similarly blocked neuron death in a DAF-16/FOXO3a-dependent manner. These results indicate that activation of the DAF-16/FOXO3a pathway, genetically or pharmacologically, confers protection against the known causes of motor neuron diseases

The insulin/IGF signaling regulators cytohesin/GRP-1 and PIP5K/PPK-1 modulate susceptibility to excitotoxicity in C. elegans.

During ischemic stroke, malfunction of excitatory amino acid transporters and reduced synaptic clearance causes accumulation of Glutamate (Glu) and excessive stimulation of postsynaptic neurons, which can lead to their degeneration by excitotoxicity. The balance between cell death-promoting (neurotoxic) and survival-promoting (neuroprotective) signaling cascades determines the fate of neurons exposed to the excitotoxic insult. The evolutionary conserved Insulin/IGF Signaling (IIS) cascade can participate in this balance, as it controls cell stress resistance in nematodes and mammals. Blocking the IIS cascade allows the transcription factor FoxO3/DAF-16 to accumulate in the nucleus and activate a transcriptional program that protects cells from a range of insults. We study the effect of IIS cascade on neurodegeneration in a C. elegans model of excitotoxicity, where a mutation in a central Glu transporter (glt-3) in a sensitizing background causes Glu-Receptor -dependent neuronal necrosis. We expand our studies on the role of the IIS cascade in determining susceptibility to excitotoxic necrosis by either blocking IIS at the level of PI3K/AGE-1 or stimulating it by removing the inhibitory effect of ZFP-1 on the expression of PDK-1. We further show that the components of the Cytohesin/GRP-1, Arf, and PIP5K/PPK-1 complex, known to regulate PIP2 production and the IIS cascade, modulate nematode excitotoxicity: mutations that are expected to reduce the complex's ability to produce PIP2 and inhibit the IIS cascade protect from excitotoxicity, while overstimulation of PIP2 production enhances neurodegeneration. Our observations therefore affirm the importance of the IIS cascade in determining the susceptibility to necrotic neurodegeneration in nematode excitotoxicity, and demonstrate the ability of Cytohesin/GRP-1, Arf, and PIP5K/PPK-1 complex to modulate neuroprotection

A model for regulation of IIS-mediated neuroprotection in nematode excitotoxicity.

<p>When the Cytohesin/GRP-1, Arf, and PIP5K/PPK-1 complex is active, it stimulates the IIS cascade, resulting in phosphorylation and cytoplasmic sequestration of FoxO/DAF-16. If the IIS cascade is less active, un-phosphorylated FoxO/DAF-16 accumulates in the nucleus and activates a transcriptional program that results in neuroprotection from the excitotoxic insult.</p

Blocking canonical apoptosis using a <i>ced-4</i> mutation does not suppress cell death in nematode excitotoxicity.

<p>Blocking canonical apoptosis using a <i>ced-4</i> mutation does not suppress cell death in nematode excitotoxicity.</p

KO of <i>arf-1.2</i> provides neuroprotection.

<p>**: p<0.05; ***: p<0.01.</p

Epistasis analysis suggests that <i>grp-1</i> works in the same pathway as <i>age-1</i>.

<p><i>grp-1</i> was inactivated using a KO strain. <i>age-1</i> was inhibited using the drug LY294002. If these two factors worked in separate pathways, their ability to suppress neurodegeneration would be (at least partially) additive, a concept not supported by our observations. The levels of neurodegeneration seen in our original excitotoxicity strain (under ethanol conditions needed to be used in this experiment) is equally different from the reduced neurodegeneration seen with inhibition of <i>grp-1</i>, <i>age-1</i>, or both (***: p<0.01).</p