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1.
Genes Brain Behav ; 4(2): 77-88, 2005 Mar.
Article in English | MEDLINE | ID: mdl-15720404

ABSTRACT

Shaker-type potassium (K+) channels are composed of pore-forming alpha subunits associated with cytoplasmic beta subunits. Kv beta2 is the predominant Kv beta subunit in the mammalian nervous system, but its functions in vivo are not clear. Kv beta2-null mice have been previously characterized in our laboratory as having reduced lifespans, cold swim-induced tremors and occasional seizures, but no apparent defect in Kv alpha-subunit trafficking. To test whether strain differences might influence the severity of this phenotype, we analyzed Kv beta2-null mice in different strain backgrounds: 129/SvEv (129), C57BL/6J (B6) and two mixed B6/129 backgrounds. We found that strain differences significantly affected survival, body weight and thermoregulation in Kv beta2-null mice. B6 nulls had a more severe phenotype than 129 nulls in these measures; this dramatic difference did not reflect alterations in seizure thresholds but may relate to strain differences we observed in cerebellar Kv1.2 expression. To specifically test whether Kv beta1 is a genetic modifier of the Kv beta2-null phenotype, we generated Kv beta1.1-deficient mice by gene targeting and bred them to Kv beta2-null mice. Kv beta1.1/Kv beta2 double knockouts had significantly increased mortality compared with either single knockout but still maintained surface expression of Kv1.2, indicating that trafficking of this alpha subunit does not require either Kv beta subunit. Our results suggest that genetic differences between 129/SvEv and C57Bl/6J are key determinants of the severity of defects seen in Kv beta2-null mice and that Kv beta1.1 is a specific although not strain-dependent modifier.


Subject(s)
Epilepsy/genetics , Mice, Knockout/genetics , Potassium Channels, Voltage-Gated/genetics , Animals , Body Temperature Regulation , Cerebellum/physiology , Cold Temperature , Epilepsy/mortality , Exons , Kv1.1 Potassium Channel , Mice , Mice, Inbred C57BL , Myoclonus/genetics , Myoclonus/mortality , Phenotype , Severity of Illness Index , Shaker Superfamily of Potassium Channels , Species Specificity , Survival Rate , Swimming
2.
Dev Cell ; 1(3): 351-61, 2001 Sep.
Article in English | MEDLINE | ID: mdl-11702947

ABSTRACT

Segregation Distorter (SD) is a meiotic drive system in Drosophila that causes preferential transmission of the SD chromosome from SD/SD+ males owing to dysfunction of SD+ spermatids. The Sd locus, which is essential for distortion, encodes a truncated RanGAP (Ran GTPase activating protein), a key nuclear transport factor. Here, we show that Sd-RanGAP retains normal enzyme activity but is mislocalized to nuclei. Distortion is abolished when enzymatic activity or nuclear localization of Sd-RanGAP is perturbed. Overexpression of Ran or RanGEF (Ran GTPase exchange factor) in the male germline fully suppresses distortion. We conclude that mislocalization of Sd-RanGAP causes distortion by reducing nuclear RanGTP, thereby disrupting the Ran signaling pathway. Nuclear transport of a GFP reporter in salivary glands is impaired by SD, suggesting that a defect in nuclear transport may underlie sperm dysfunction.


Subject(s)
Cell Nucleus/metabolism , Chromosome Segregation , Drosophila Proteins , Drosophila melanogaster/metabolism , GTPase-Activating Proteins/metabolism , Active Transport, Cell Nucleus/physiology , Animals , Drosophila melanogaster/enzymology , Drosophila melanogaster/genetics , Female , GTPase-Activating Proteins/genetics , Genes, Reporter , Immunohistochemistry , Insect Proteins/genetics , Insect Proteins/metabolism , Male , Meiosis/physiology , Recombinant Fusion Proteins/genetics , Recombinant Fusion Proteins/metabolism , Salivary Glands/cytology , Salivary Glands/physiology , Spermatocytes/metabolism , Testis/metabolism , Transgenes
3.
J Neurosci ; 21(23): 9255-64, 2001 Dec 01.
Article in English | MEDLINE | ID: mdl-11717359

ABSTRACT

Rabphilin, a putative rab effector, interacts specifically with the GTP-bound form of the synaptic vesicle-associated protein rab3a. In this study, we define in vivo functions for rabphilin through the characterization of mutants that disrupt the Caenorhabditis elegans rabphilin homolog. The mutants do not display the general synaptic defects associated with rab3 lesions, as assayed at the pharmacological, physiological, and ultrastructural level. However, rabphilin mutants exhibit severe lethargy in the absence of mechanical stimulation. Furthermore, rabphilin mutations display strong synergistic interactions with hypomorphic lesions in the syntaxin, synaptosomal-associated protein of 25 kDa, and synaptobrevin soluble N-ethylmaleimide sensitive factor attachment protein receptor (SNARE) genes; double mutants were nonresponsive to mechanical stimulation. These synergistic interactions were independent of rab3 function and were not observed in rab3-SNARE double mutants. Our data reveal rab3-independent functions for rabphilin in the potentiation of SNARE function.


Subject(s)
Carrier Proteins/metabolism , Membrane Proteins/metabolism , Nerve Tissue Proteins/metabolism , Vesicular Transport Proteins , rab GTP-Binding Proteins/metabolism , rab3 GTP-Binding Proteins/metabolism , Adaptor Proteins, Signal Transducing , Animals , Behavior, Animal/physiology , Biomarkers , Caenorhabditis elegans , Caenorhabditis elegans Proteins/genetics , Caenorhabditis elegans Proteins/metabolism , Electrophysiology , Guanosine Triphosphate/metabolism , Helminth Proteins/genetics , Helminth Proteins/metabolism , Locomotion/genetics , Membrane Proteins/genetics , Molecular Sequence Data , Mutagenesis, Site-Directed , Nerve Tissue Proteins/genetics , Phenotype , Physical Stimulation , Protein Structure, Tertiary/physiology , SNARE Proteins , Sequence Deletion , Sequence Homology, Amino Acid , Sleep Stages/genetics , Soluble N-Ethylmaleimide-Sensitive Factor Attachment Proteins , Synapses/metabolism , Synaptic Vesicles/metabolism , Synaptosomal-Associated Protein 25 , rab GTP-Binding Proteins/genetics , rab3 GTP-Binding Proteins/genetics , Rabphilin-3A
4.
Proc Natl Acad Sci U S A ; 98(21): 12233-8, 2001 Oct 09.
Article in English | MEDLINE | ID: mdl-11593041

ABSTRACT

Soluble N-ethylmaleimide-sensitive fusion attachment protein receptor (SNARE)-mediated fusion of synaptic vesicles with the presynaptic-plasma membrane is essential for communication between neurons. Disassembly of the SNARE complex requires the ATPase N-ethylmaleimide-sensitive fusion protein (NSF). To determine where in the synaptic-vesicle cycle NSF functions, we have undertaken a genetic analysis of comatose (dNSF-1) in Drosophila. Characterization of 16 comatose mutations demonstrates that NSF mediates disassembly of SNARE complexes after synaptic-vesicle fusion. Hypomorphic mutations in NSF cause temperature-sensitive paralysis, whereas null mutations result in lethality. Genetic-interaction studies with para demonstrate that blocking evoked fusion delays the accumulation of assembled SNARE complexes and behavioral paralysis that normally occurs in comatose mutants, indicating NSF activity is not required in the absence of vesicle fusion. In addition, the entire vesicle pool can be depleted in shibire comatose double mutants, demonstrating that NSF activity is not required for the fusion step itself. Multiple rounds of vesicle fusion in the absence of NSF activity poisons neurotransmission by trapping SNAREs into cis-complexes. These data indicate that NSF normally dissociates and recycles SNARE proteins during the interval between exocytosis and endocytosis. In the absence of NSF activity, there are sufficient fusion-competent SNAREs to exocytose both the readily released and the reserve pool of synaptic vesicles.


Subject(s)
Carrier Proteins/metabolism , Membrane Proteins/metabolism , Synaptic Vesicles/metabolism , Vesicular Transport Proteins , Alleles , Animals , Carrier Proteins/genetics , Drosophila melanogaster , Female , Male , Membrane Fusion/physiology , Mutagenesis , N-Ethylmaleimide-Sensitive Proteins , SNARE Proteins , Synaptic Vesicles/physiology
5.
Neuron ; 31(3): 421-37, 2001 Aug 16.
Article in English | MEDLINE | ID: mdl-11516399

ABSTRACT

Calcium-activated protein for secretion (CAPS) is proposed to play an essential role in Ca2+-regulated dense-core vesicle exocytosis in vertebrate neuroendocrine cells. Here we report the cloning, mutation, and characterization of the Drosophila ortholog (dCAPS). Null dCAPS mutants display locomotory deficits and complete embryonic lethality. The mutant NMJ reveals a 50% loss in evoked glutamatergic transmission, and an accumulation of synaptic vesicles at active zones. Importantly, dCAPS mutants display a highly specific 3-fold accumulation of dense-core vesicles in synaptic terminals, which was not observed in mutants that completely arrest synaptic vesicle exocytosis. Targeted transgenic CAPS expression in identified motoneurons fails to rescue dCAPS neurotransmission defects, demonstrating a cell nonautonomous role in synaptic vesicle fusion. We conclude that dCAPS is required for dense-core vesicle release and that a dCAPS-dependent mechanism modulates synaptic vesicle release at glutamatergic synapses.


Subject(s)
Calcium-Binding Proteins/physiology , Drosophila melanogaster/physiology , Synaptic Vesicles/physiology , Amino Acid Sequence , Animals , Animals, Genetically Modified , Calcium-Binding Proteins/chemistry , Calcium-Binding Proteins/genetics , Cloning, Molecular , Conserved Sequence , Drosophila melanogaster/embryology , Drosophila melanogaster/genetics , Embryo, Nonmammalian/physiology , Exocytosis , Genes, Essential , Glutamic Acid/physiology , Membrane Fusion/physiology , Molecular Sequence Data , Motor Activity , Motor Neurons/physiology , Neuromuscular Junction/physiology , Rats , Sequence Alignment , Sequence Homology, Amino Acid , Synaptic Transmission/physiology , Vesicular Transport Proteins
6.
J Neurosci ; 21(5): 1421-33, 2001 Mar 01.
Article in English | MEDLINE | ID: mdl-11222632

ABSTRACT

Synaptotagmin has been proposed to function as a Ca(2+) sensor that regulates synaptic vesicle exocytosis, whereas the soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) complex is thought to form the core of a conserved membrane fusion machine. Little is known concerning the functional relationships between synaptotagmin and SNAREs. Here we report that synaptotagmin can facilitate SNARE complex formation in vitro and that synaptotagmin mutations disrupt SNARE complex formation in vivo. Synaptotagmin oligomers efficiently bind SNARE complexes, whereas Ca(2+) acting via synaptotagmin triggers cross-linking of SNARE complexes into dimers. Mutations in Drosophila that delete the C2B domain of synaptotagmin disrupt clathrin AP-2 binding and endocytosis. In contrast, a mutation that blocks Ca(2+)-triggered conformational changes in C2B and diminishes Ca(2+)-triggered synaptotagmin oligomerization results in a postdocking defect in neurotransmitter release and a decrease in SNARE assembly in vivo. These data suggest that Ca(2+)-driven oligomerization via the C2B domain of synaptotagmin may trigger synaptic vesicle fusion via the assembly and clustering of SNARE complexes.


Subject(s)
Calcium-Binding Proteins , Calcium/metabolism , Membrane Fusion/physiology , Membrane Glycoproteins/genetics , Membrane Glycoproteins/metabolism , Nerve Tissue Proteins/genetics , Nerve Tissue Proteins/metabolism , Synaptic Vesicles/metabolism , Vesicular Transport Proteins , Adaptor Protein Complex alpha Subunits , Adaptor Proteins, Vesicular Transport , Animals , Biopolymers/biosynthesis , Biopolymers/chemistry , Calcium/pharmacology , Dimerization , Drosophila , Endocytosis/physiology , Exocytosis/physiology , Macromolecular Substances , Membrane Fusion/drug effects , Membrane Glycoproteins/chemistry , Membrane Proteins/chemistry , Membrane Proteins/metabolism , Mutation , Nerve Tissue Proteins/chemistry , Precipitin Tests , Protein Conformation , Protein Structure, Tertiary/genetics , Rats , SNARE Proteins , Structure-Activity Relationship , Synaptotagmins
7.
Genetics ; 155(3): 1149-60, 2000 Jul.
Article in English | MEDLINE | ID: mdl-10880477

ABSTRACT

Post-transcriptional editing of pre-mRNAs through the action of dsRNA adenosine deaminases results in the modification of particular adenosine (A) residues to inosine (I), which can alter the coding potential of the modified transcripts. We describe here three sites in the para transcript, which encodes the major voltage-activated Na(+) channel polypeptide in Drosophila, where RNA editing occurs. The occurrence of RNA editing at the three sites was found to be developmentally regulated. Editing at two of these sites was also conserved across species between the D. melanogaster and D. virilis. In each case, a highly conserved region was found in the intron downstream of the editing site and this region was shown to be complementary to the region of the exonic editing site. Thus, editing at these sites would appear to involve a mechanism whereby the edited exon forms a base-paired secondary structure with the distant conserved noncoding sequences located in adjacent downstream introns, similar to the mechanism shown for A-to-I RNA editing of mammalian glutamate receptor subunits (GluRs). For the third site, neither RNA editing nor the predicted RNA secondary structures were evolutionarily conserved. Transcripts from transgenic Drosophila expressing a minimal editing site construct for this site were shown to faithfully undergo RNA editing. These results demonstrate that Na(+) channel diversity in Drosophila is increased by RNA editing via a mechanism analogous to that described for transcripts encoding mammalian GluRs.


Subject(s)
Conserved Sequence/genetics , Drosophila/genetics , RNA Editing/genetics , RNA, Messenger/metabolism , Sodium Channels/genetics , Alternative Splicing , Animals , Base Pairing , Base Sequence , Binding Sites , Drosophila/metabolism , Gene Expression Regulation, Developmental , Insect Proteins/genetics , Insect Proteins/metabolism , Introns , Molecular Sequence Data , Nucleic Acid Conformation , Protein Structure, Secondary , Reverse Transcriptase Polymerase Chain Reaction , Sodium Channels/metabolism
9.
Neuron ; 25(1): 139-49, 2000 Jan.
Article in English | MEDLINE | ID: mdl-10707979

ABSTRACT

The mle(napts) mutation causes temperature-dependent blockade of action potentials resulting from decreased abundance of para-encoded Na+ channels. Although maleless (mle) encodes a double-stranded RNA (dsRNA) helicase, exactly how mle(napts) affects para expression remained uncertain. Here, we show that para transcripts undergo adenosine-to-inosine (A-to-I) RNA editing via a mechanism that apparently requires dsRNA secondary structure formation encompassing the edited exon and the downstream intron. In an mle(napts) background, >80% of para transcripts are aberrant, owing to internal deletions that include the edited exon. We propose that the Mle helicase is required to resolve the dsRNA structure and that failure to do so in an mle(napts) background causes exon skipping because the normal splice donor is occluded. These results explain how mlen(napts) affects Na+ channel expression and provide new insights into the mechanism of RNA editing.


Subject(s)
Chromosomal Proteins, Non-Histone , DNA Helicases , DNA-Binding Proteins , Drosophila Proteins , RNA Editing/genetics , RNA Helicases/genetics , RNA Splicing/genetics , Sodium Channels/genetics , Transcription Factors/genetics , Action Potentials/physiology , Animals , Animals, Genetically Modified , Base Sequence , Conserved Sequence , DNA, Complementary , Drosophila , Evolution, Molecular , Gene Dosage , Introns/genetics , Molecular Sequence Data , Neurons/chemistry , Neurons/enzymology , Nucleic Acid Conformation , Phenotype , RNA, Double-Stranded/chemistry , RNA, Double-Stranded/genetics , RNA, Messenger/chemistry , RNA, Messenger/genetics , Sequence Homology, Amino Acid , Sodium Channels/metabolism , Transcription, Genetic/genetics
10.
Kidney Int ; 57(3): 766-71, 2000 Mar.
Article in English | MEDLINE | ID: mdl-10720927

ABSTRACT

To dissect the molecular mechanisms of electrical activity in the nervous system, an extensive collection of mutations affecting various types of voltage-gated ion channels was identified and characterized in Drosophila. Most of these mutations were generated by chemical mutagenesis and were recognized on the basis of defects in motor behavior. These were the first genetically determined ion channelopathies to be characterized in any multicellular organism. Drosophila is a particularly attractive model system for such studies because of the availability of powerful genetic, electrophysiological, and molecular techniques for generating new mutations, characterizing their phenotypes, and cloning the genes thus defined. Consequently, a number of ion channels, including various types of K+ channels that had not yielded previously to biochemical approaches, were first identified via a genetic strategy in Drosophila. Evolutionary conservation of these genes enabled subsequent isolation of the corresponding genes from various mammals, including humans. Several of these human homologues have been found to be associated with heritable neuromuscular disorders. Studies of ion channel mutations in Drosophila have thus provided important biological information concerning the molecular and functional diversity of ion channels, their evolutionary relationships, and their in vivo functions in the nervous system. Similar studies of additional new mutations should now facilitate the analysis of ion channel regulatory mechanisms.


Subject(s)
Cation Transport Proteins , DNA-Binding Proteins , Drosophila/genetics , Drosophila/metabolism , Ion Channels/genetics , Ion Channels/metabolism , Mutation , Potassium Channels, Voltage-Gated , Trans-Activators , Animals , Arrhythmias, Cardiac/genetics , Drosophila Proteins , ERG1 Potassium Channel , Ether-A-Go-Go Potassium Channels , Humans , Potassium Channels/genetics , Transcriptional Regulator ERG
11.
Recept Channels ; 7(3): 237-43, 2000.
Article in English | MEDLINE | ID: mdl-11342391

ABSTRACT

Voltage-gated K(+) channel alpha subunits (K(V) alpha) have been previously identified in pancreatic islet beta-cells where it has been suggested they have a role in membrane repolarization and insulin secretion. Here we report the cloning of the three mammalian K(V) beta subunits, including splice variants of these subunits, from both human and rat pancreatic islets and from the rat insulinoma cell line INS-1. Two of the splice variants, K(V) beta1a and K(V) beta3, previously reported to be neuronal tissue specific, are expressed in islets and INS-1 cells. In addition, a splice variant of K(V) beta2 that lacks two potential protein kinase C phosphorylation sites at the amino terminus is present. Immunoblot analysis suggests a high level of K(V) beta2 subunit protein in rat pancreatic islets and immunoprecipitation with anti-K(V) beta2 antibody pulls down a protein from INS-1 cells that reacts with anti-aldose reductase antibody. The K(V) beta subunits, which are attached to the cytoplasmic face of the alpha subunits and are members of the aldose reductase superfamily of NADPH oxidoreductases, may have an as yet undetermined role in the regulation of insulin secretion by the intracellular redox potential. Finally, we suggest that a systematic nomenclature for K(V) beta subunits first proposed by McCormack et al. be adopted for this family of potassium channel subunits as it corresponds with the nomenclature used for their cognate K(V) alpha subunits.


Subject(s)
Aldehyde Reductase/genetics , Islets of Langerhans/physiology , NADH, NADPH Oxidoreductases/genetics , Potassium Channels/genetics , Alternative Splicing , Amino Acid Sequence , Animals , Base Sequence , Cloning, Molecular , Humans , Ion Channel Gating , Molecular Sequence Data , Protein Subunits , Rats , Sequence Analysis, DNA , Sequence Analysis, Protein , Tumor Cells, Cultured
12.
Mol Psychiatry ; 4(5): 408-9, 1999 Sep.
Article in English | MEDLINE | ID: mdl-10523808

ABSTRACT

The newly identified axotactin (AXO) protein in Drosophila, a member of the neurexin superfamily, appears to be a key element that mediates the effects of glial cells on neuronal membrane excitability and synaptic plasticity.


Subject(s)
Cell Communication/genetics , Nerve Tissue Proteins/genetics , Neuroglia/cytology , Neurons/cytology , Animals , Drosophila , Insect Proteins/genetics
13.
Nature ; 400(6746): 757-60, 1999 Aug 19.
Article in English | MEDLINE | ID: mdl-10466723

ABSTRACT

Communication within the nervous system is mediated by Ca2+-triggered fusion of synaptic vesicles with the presynaptic plasma membrane. Genetic and biochemical evidence indicates that synaptotagmin I may function as a Ca2+ sensor in neuronal exocytosis because it can bind Ca2+ and penetrate into lipid bilayers. Chronic depolarization or seizure activity results in the upregulation of a distinct and unusual isoform of the synaptotagmin family, synaptotagmin IV. We have identified a Drosophila homologue of synaptotagmin IV that is enriched on synaptic vesicles and contains an evolutionarily conserved substitution of aspartate to serine that abolishes its ability to bind membranes in response to Ca2+ influx. Synaptotagmin IV forms hetero-oligomers with synaptotagmin I, resulting in synaptotagmin clusters that cannot effectively penetrate lipid bilayers and are less efficient at coupling Ca2+ to secretion in vivo: upregulation of synaptotagmin IV, but not synaptotagmin I, decreases evoked neurotransmission. These findings indicate that modulating the expression of synaptotagmins with different Ca2+-binding affinities can lead to heteromultimers that can regulate the efficiency of excitation-secretion coupling in vivo and represent a new molecular mechanism for synaptic plasticity.


Subject(s)
Calcium-Binding Proteins , Membrane Glycoproteins/physiology , Nerve Tissue Proteins/physiology , Synapses/physiology , Amino Acid Sequence , Animals , Animals, Genetically Modified , Caenorhabditis elegans , Calcium/metabolism , Drosophila , Electrophysiology , Liposomes/metabolism , Membrane Glycoproteins/chemistry , Membrane Glycoproteins/genetics , Membrane Glycoproteins/metabolism , Molecular Sequence Data , Nerve Tissue Proteins/chemistry , Nerve Tissue Proteins/genetics , Nerve Tissue Proteins/metabolism , Protein Binding , Recombinant Fusion Proteins/metabolism , Sequence Homology, Amino Acid , Synaptic Vesicles/metabolism , Synaptotagmin I , Synaptotagmins , Tissue Distribution
15.
Ann N Y Acad Sci ; 868: 356-69, 1999 Apr 30.
Article in English | MEDLINE | ID: mdl-10414305

ABSTRACT

Mutations of eag, first identified in Drosophila on the basis of their leg-shaking phenotype, cause repetitive firing and enhanced transmitter release in motor neurons. The encoded EAG polypeptide is related both to voltage-gated K+ channels and to cyclic nucleotide-gated cation channels. Homology screens identified a family of eag-related channel polypeptides, highly conserved from nematodes to humans, comprising three subfamilies: EAG, ELK, and ERG. When expressed in frog oocytes, EAG channels behave as voltage-dependent, outwardly rectifying K(+)-selective channels. Mutations of the human eag-related gene (HERG) result in a form of cardiac arrhythmia that can lead to ventricular fibrillation and sudden death. Electrophysiological and pharmacological studies have provided evidence that HERG channels specify one component of the delayed rectifier, IKr, that contributes to the repolarization phase of cardiac action potentials. An important role for HERG channels in neuronal excitability is also suggested by the expression of these channels in brain tissue. Moreover, mutations of ERG-type channels in the Drosophila sei mutant cause temperature-induced convulsive seizures associated with aberrant bursting activity in the flight motor pathway. The in vivo function of ELK channels has not yet been established, but when these channels are expressed in frog oocytes, they display properties intermediate between those of EAG- and ERG-type channels. Coexpression of the K(+)-channel beta subunit encoded by Hk with EAG in oocytes dramatically increases current amplitude and also affects the gating and modulation of these currents. Biochemical evidence indicates a direct physical interaction between EAG and HK proteins. Overall, these studies highlight the diverse properties of the eag family of K+ channels, which are likely to subserve diverse functions in vivo.


Subject(s)
Cation Transport Proteins , DNA-Binding Proteins , Drosophila/genetics , Potassium Channels, Voltage-Gated , Potassium Channels/genetics , Trans-Activators , Alternative Splicing , Amino Acid Sequence , Animals , Arrhythmias, Cardiac/genetics , Drosophila/metabolism , Drosophila Proteins , ERG1 Potassium Channel , Electrophysiology , Ether-A-Go-Go Potassium Channels , Evolution, Molecular , Gene Expression Regulation , Humans , Molecular Sequence Data , Mutation , Potassium Channels/metabolism , Receptor Protein-Tyrosine Kinases/genetics , Receptor, EphB4 , Receptors, Eph Family , Sequence Alignment , Transcriptional Regulator ERG
18.
J Neurosci ; 19(8): 2906-18, 1999 Apr 15.
Article in English | MEDLINE | ID: mdl-10191308

ABSTRACT

Members of the Ether à go-go (Eag) K+ channel subfamilies Eag, Erg, and Elk are widely expressed in the nervous system, but their neural functions in vivo remain largely unknown. The biophysical properties of channels from the Eag and Erg subfamilies have been described, and based on their characteristic features and expression patterns, Erg channels have been associated with native currents in the heart. Little is known about the properties of channels from the Elk subfamily. We have identified a mouse gene, Melk2, that encodes a predicted polypeptide with 48% amino acid identity to Drosophila Elk but only 40 and 36% identity with mouse Erg (Merg) and Eag (Meag), respectively. Melk2 RNA appears to be expressed at high levels only in brain tissue. Functional expression of Melk2 in Xenopus oocytes reveals large, transient peaks of current at the onset of depolarization. Like Meag currents, Melk2 currents activate relatively quickly, but they lack the nonsuperimposable Cole-Moore shift characteristic of the Eag subfamily. Melk2 currents are insensitive to E-4031, a class III antiarrhythmic compound that blocks the Human Ether-à-go-go-Related Gene (HERG) channel and its counterpart in native tissues, IKr. Melk2 channels exhibit inward rectification because of a fast C-type inactivation mechanism, but the slower rate of inactivation and the faster rate of activation results in less inward rectification than that observed in HERG channels. This characterization of Melk currents should aid in identification of native counterparts to the Elk subfamily of channels in the nervous system.


Subject(s)
Brain/physiology , Nerve Tissue Proteins/physiology , Potassium Channels/physiology , Protein Serine-Threonine Kinases/physiology , Action Potentials/physiology , Amino Acid Sequence , Animals , Ether-A-Go-Go Potassium Channels , Humans , Kinetics , Mice , Molecular Sequence Data , Multigene Family , Oocytes/physiology , Potassium Channels/genetics , Sequence Homology, Amino Acid , Xenopus
19.
Science ; 283(5408): 1742-5, 1999 Mar 12.
Article in English | MEDLINE | ID: mdl-10073941

ABSTRACT

Segregation Distorter (SD) in Drosophila melanogaster is a naturally occurring meiotic drive system in which the SD chromosome is transmitted from SD/SD+ males in vast excess over its homolog owing to the induced dysfunction of SD+-bearing spermatids. The Sd locus is the key distorting gene responsible for this phenotype. A genomic fragment from the Sd region conferred full distorting activity when introduced into the appropriate genetic background by germline transformation. The only functional product encoded by this fragment is a truncated version of the RanGAP nuclear transport protein. These results demonstrate that this mutant RanGAP is the functional Sd product.


Subject(s)
Carrier Proteins/genetics , Drosophila Proteins , Drosophila melanogaster/genetics , GTPase-Activating Proteins , Genes, Insect , Meiosis , Nuclear Proteins/genetics , Animals , Carrier Proteins/chemistry , Carrier Proteins/physiology , Cell Nucleus/metabolism , Crosses, Genetic , DNA, Complementary , Drosophila melanogaster/physiology , Female , Gene Duplication , Gene Expression , Male , Nuclear Proteins/chemistry , Nuclear Proteins/physiology , RNA, Messenger/genetics , Spermatids/physiology , Sulfotransferases/chemistry , Sulfotransferases/genetics , Transcription, Genetic , Transformation, Genetic
20.
Science ; 283(5406): 1343-5, 1999 Feb 26.
Article in English | MEDLINE | ID: mdl-10037607

ABSTRACT

In the nervous system, glial cells greatly outnumber neurons but the full extent of their role in determining neural activity remains unknown. Here the axotactin (axo) gene of Drosophila was shown to encode a member of the neurexin protein superfamily secreted by glia and subsequently localized to axonal tracts. Null mutations of axo caused temperature-sensitive paralysis and a corresponding blockade of axonal conduction. Thus, the AXO protein appears to be a component of a glial-neuronal signaling mechanism that helps to determine the membrane electrical properties of target axons.


Subject(s)
Axons/physiology , Drosophila/physiology , Insect Proteins/physiology , Neuroglia/physiology , Signal Transduction , Action Potentials , Animals , DNA, Complementary , Drosophila/embryology , Drosophila/genetics , Embryo, Nonmammalian/physiology , Gene Expression Regulation, Developmental , Genes, Insect , Insect Proteins/genetics , Ion Channels/physiology , Mutation , Neuromuscular Junction/physiology , Synaptic Transmission , Temperature
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