To the surprise of many, studies of molecular mechanisms of touch transduction and analyses of epithelial Na+ 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 amiloride‐sensitive Na+ 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+ 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.

Desensitization of the ␣-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) subtype of glutamate receptor channels is an important process shaping the time course of synaptic excitation. Upon desensitization, the receptor channel closes and the agonist affinity increases. So far, the nature of the structural rearrangements leading to these events was unknown. On the basis of the structural homology of the ligand binding domains of AMPA receptors and of the bilobated bacterial periplasmic proteins, we now show that agonist interaction with one lobe of the GluR1 subunit of homomeric AMPA receptors controls channel activation while additional interactions with the other lobe cause channel desensitization. Accordingly, we suggest that the transition of the AMPA receptor channel to the desensitized state involves the agonist-mediated stabilization of the closed lobe conformation of its binding domain and is a process akin to that used by the venus flytrap.The AMPA 1 subtype of glutamate receptors plays a central role in brain excitatory synaptic transmission. Activated by agonists, the receptor channels first open and then desensitize (1-4). The process of desensitization is of physiological importance since it may determine both the duration of excitatory postsynaptic currents as well as the ability to respond to high frequency stimulation (3,(5)(6)(7)(8)(9)(10). Desensitization results from a transition, still unexplained in structural terms, of the receptor channel complex from an open state, which binds the agonist with low affinity, to a closed state, which binds with high affinity (11-15). Recently, it was suggested (16 -19) that the structure of the agonist binding domain of glutamate receptor channels is homologous to that of bacterial periplasmic binding proteins and in particular to the lysine-arginine-ornithine binding protein (LAOBP) (20). For this bi-lobated amino acid binding protein, a substrate binding mechanism akin to that of a venus flytrap was suggested. A single substrate molecule binds to lobe I with low affinity and, by further binding to lobe II, stabilizes a high affinity closed lobe conformation (21-23). Assuming that the structural homology between AMPA receptors and bacterial periplasmic binding proteins extends to a similarity in function, we have investigated here whether the elevation of agonist affinity upon AMPA receptor desensitization might also involve a venus flytrap mechanism. The data presented here strongly suggest that this might indeed be the case. EXPERIMENTAL PROCEDURESMutagenesis and Oocyte Expression-GluR1 amino acid residues possibly involved in agonist binding were identified on the basis of their homology with the residues lining the ligand binding pocket of LAOBP (23), as deduced from amino acid sequence alignments of glutamate receptors (from rat), kainate binding proteins, and bacterial periplasmic proteins (LAOBP, QBP, and HisJ) using the UW-GCG sequence analysis software package (16 -18). Minor shifts were introduced to optimize the homology. The selected ...

Aging is a risk factor for the development of adult-onset neurodegenerative diseases. Although 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 Caenorhabditis 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.

Mechanosensory signaling mediated by mechanically gated ion channels constitutes the basis for the senses of touch and hearing and contributes fundamentally to the development and homeostasis of all organisms. Despite this profound importance in biology, little is known of the molecular identities or functional requirements of mechanically gated ion channels. We report a genetically based structure-function analysis of the candidate mechanotransducing channel subunit MEC-4, a core component of a touch-sensing complex in Caenorhabditis elegans and a member of the DEG/ENaC superfamily. We identify molecular lesions in 40 EMS-induced mec-4 alleles and further probe residue and domain function using site-directed approaches. Our analysis highlights residues and subdomains critical for MEC-4 activity and suggests possible roles of these in channel assembly and/or function. We describe a class of substitutions that disrupt normal channel activity in touch transduction but remain permissive for neurotoxic channel hyperactivation, and we show that expression of an N-terminal MEC-4 fragment interferes with in vivo channel function. These data advance working models for the MEC-4 mechanotransducing channel and identify residues, unique to MEC-4 or the MEC-4 degenerin subfamily, that might be specifically required for mechanotransducing function. Because many other substitutions identified by our study affect residues conserved within the DEG/ENaC channel superfamily, this work also provides a broad view of structure-function relations in the superfamily as a whole. Because the C. elegans genome encodes representatives of a large number of eukaryotic channel classes, we suggest that similar genetic-based structure-activity studies might be generally applied to generate insight into the in vivo function of diverse channel types.

The structure of glutamate receptor-channel (GluR) subunits has recently been shown to differ from that of other ligand-gated channels and to contain a voltage-gated channel-like pore-forming motif. The view that the structure of GluR complexes is similar to the pentameric structure of other ligand-gated channels was questioned here. Studies of the response properties of the GluR1 subunit of the AMPA subtype of GluRs, co-expressed in Xenopus oocytes with its L646A mutant, which differs only by a greatly reduced sensitivity to quisqualate, provide new evidence suggesting that the GluR1 homomeric receptor channel has a tetrameric structure.