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B Ganetzky

Publications and source records attributed to B Ganetzky.

At least 55 records · Page 3Linked to original sources
PubMed1995

Conserved alternative splicing patterns and splicing signals in the Drosophila sodium channel gene para.

We cloned genomic DNA corresponding to the Drosophila virilis homologue of para, a gene encoding a sodium channel alpha-subunit, and obtained many partial cDNA clones from embryos and adults. Para protein has been well conserved, and the optional elements at six different sites of alternative splicing in D. melanogaster are present in D. virilis, in addition to one new optional exon. Among 31 different splice-types observed in D. virilis, the stage-specific pattern of alternative splicing seen in D. melanogaster is also conserved. Comparison of genomic DNA sequence revealed three aspects that vary between alternatively and constitutively used exon sequences. Sixteen short blocks (10-75 bp), the only recognizably conserved intron sequence, were disproportionately associated with alternatively used splice sites. Silent site substitutions were found much less frequently in alternative than constitutive exon elements, and the degree of match to the Drosophila splice site consensus tended to be lower at less frequently selected alternative splice junctions. This study shows that the developmentally regulated variability of para products is highly conserved and therefore likely to be of functional significance and suggests that a variety of different sequence-dependent mechanisms may regulate this pattern of alternative splicing.

Alternative Splicing↗
PubMed1994

Neurally expressed Drosophila genes encoding homologs of the NSF and SNAP secretory proteins.

Several lines of investigation have now converged to indicate that the neurotransmitter release apparatus is formed by assembly of cytosolic proteins with proteins of the synaptic vesicle and presynaptic terminal membranes. We are undertaking a genetic approach in Drosophila melanogaster to investigate the functions of two types of cytosolic proteins thought to function in this complex: N-ethylmaleimide-sensitive fusion protein (NSF) and the soluble NSF attachment proteins (SNAPs). We have identified Drosophila homologs of the vertebrate and yeast NSF and SNAP genes. Both Drosophila genes encode polypeptides that closely resemble their vertebrate counterparts and are expressed in the nervous system; neither appears to be in a family of closely related Drosophila genes. These results indicate that the Drosophila NSF and SNAP genes are excellent candidates for mutational analysis of neurotransmitter release.

Amino Acid Sequence↗
PubMed1994

A family of potassium channel genes related to eag in Drosophila and mammals.

We have identified a conserved family of genes related to Drosophila eag, which encodes a distinct type of voltage-activated K+ channel. Three related genes were recovered in screens of cDNA libraries from Drosophila, mouse, and human tissues. One gene is the mouse counterpart of eag; the other two represent additional subfamilies. The human gene maps to chromosome 7. Family members share at least 47% amino acid identity in their hydrophobic cores and all contain a segment homologous to a cyclic nucleotide-binding domain. Sequence comparisons indicate that members of this family are most closely related to vertebrate cyclic nucleotide-gated cation channels and plant inward-rectifying K+ channels. The existence of another family of K+ channel structural genes further extends the known diversity of K+ channels and has important implications for the structure, function, and evolution of the superfamily of voltage-sensitive ion channels.

Amino Acid Sequence↗
PubMed1994

Cysteine strings, calcium channels and synaptic transmission.

Multidisciplinary studies have led to the discovery and characterization of cysteine string proteins (csps) in both Drosophila and Torpedo. Phenotypic analysis of csp mutants in Drosophila demonstrates a crucial role for csp in synaptic transmission. Expression studies of Torpedo csp (Tcsp) in Xenopus oocytes suggests that the protein has some role in the function of presynaptic Ca2+ channels. However, biochemical purification of Tcsp indicates that is associated with synaptic vesicles rather than with the plasma membrane of presynaptic terminals where Ca2+ channels reside. These results suggest a model in which csps serve as a link by which docked synaptic vesicles could modulate the activity of presynaptic Ca2+ channels.

Animals↗
PubMed1994

Evidence that the Drosophila olfactory mutant smellblind defines a novel class of sodium channel mutation.

The smellblind (sbl) gene of Drosophila is associated with olfactory defects, and the paralytic (para) gene encodes a voltage-gated sodium channel. sbl and para have similar genetic map positions, many combinations of sbl and para mutations fail to complement, and two sbl mutations contain molecular lesions within the para transcription unit. sbl mutations also behave like para mutations in that they are enhanced by the mutation no action potential temperature-sensitive (mlenapts1). The simplest interpretation of these results is that sbl and para are the same gene. Two sbl mutations produce olfactory defects not characteristic of classic sodium channel mutations and do not show typical heat-sensitive paralysis, suggesting that these sbl mutants define a novel class of sodium channel mutation.

Alleles↗
PubMed1994

Functional identification of the Segregation distorter locus of Drosophila melanogaster by germline transformation.

Segregation Distorter (SD) is a meiotic drive system in D. melanogaster that results in the failure of SD/SD+ males to transmit SD+ homologs owing to the induced dysfunction of spermatids carrying the normal chromosome. Segregation distorter (Sd), the gene primarily responsible for this distorted transmission, is associated with a novel 12-kb restriction fragment containing a tandem duplication of a 5-kb wild-type segment of genomic DNA. When introduced into appropriate genetic backgrounds by germline transformation, this 12-kb fragment causes full levels of distortion and directs the expression of an SD-specific 4-kb transcript. Transformants that have lost part of this segment are unable to cause distortion and do not express the 4-kb transcript. These results identify the tandem duplication as Sd.

Animals↗
PubMed1994

Cloning and characterization of human and mouse homologs of the Drosophila calcium-activated potassium channel gene, slowpoke.

Potassium channels play important roles in a wide variety of physiological processes. Although several genes encoding voltage-activated potassium channels have been analyzed at the molecular level, no calcium-activated potassium channel gene has yet been characterized in humans. In an effort to provide the foundation for functional analysis of such polypeptides we report the cloning of mouse and human homologs of the Drosophila melanogaster calcium-activated potassium channel gene, slowpoke. Both the human and mouse genes encode polypeptides that have more than 50% amino acid identifies with their Drosophila counterpart. In addition, like the Drosophila slowpoke gene, both the mouse and human genes generate multiple transcripts by alternative splicing. The human gene maps to chromosome 10 based on the results of polymerase chain reaction analysis of genomic DNA from human-hamster hybrid cell lines. Because calcium-activated potassium channels participate in wide variety of cellular functions including neuromuscular communication, secretion and cellular immunity, their continued analysis promises to have broad biological and medical significance.

Amino Acid Sequence↗
PubMed1994

Spatial and temporal expression patterns of two sodium channel genes in Drosophila.

Genetic and molecular studies have identified two different sodium channel genes in Drosophila, para and DSC1. The functional contributions of the para-encoded channel have been inferred from analysis of mutant phenotypes. However, no mutations of DSC1 have been identified, so the in vivo functions of the channel it encodes are not yet known. To learn more about the possible functions of DSC1 in the Drosophila nervous system compared with those of para, we have characterized the expression patterns of these two sodium channel genes at embryonic, larval, pupal, and adult stages by tissue in situ hybridization, para encodes the predominant type of sodium channel and is ubiquitously transcribed throughout the CNS and PNS at all developmental stages. The expression pattern of DSC1 is very different from para during embryonic and larval stages during which there are very few DSC1-expressing cells in either the CNS or PNS. Double-labeling studies suggest that some of these cells are non-neuronal. However, in pupal and adult stages, para and DSC1 have completely overlapping patterns of expression in the CNS and retina. In the pupal and adult, PNS expression of these genes is still distinct because only para transcripts are detected in wing sensory neurons. The strong and widespread expression of DSC1 in the CNS of pupae and adults suggests that the DSC1 channels are likely to provide an important function in neurons during these stages. Since most, if not all, neurons in the pupal and adult CNS express both para and DSC1, these two sodium channel genes probably subserve distinct functions within these cells. Our results provide the background for elucidating the respective in vivo contributions of para and DSC1 to neuronal excitability and for dissecting the regulatory mechanisms that underlie their different patterns of expression.

Animals↗
PubMed1994

Developmentally regulated alternative splicing generates a complex array of Drosophila para sodium channel isoforms.

The para locus encodes the predominant class of sodium channels expressed in Drosophila neurons. Previous sequence analysis of para cDNAs indicated the occurrence of alternative splicing at several sites within the open reading frame. Here we report a detailed analysis of this alternative splicing and its regulation during development. We have used a combination of RNA-PCR and sequence analysis to examine a 1.7 kilobase region of the para mRNA that encompasses the previously reported sites of alternative splicing. Five sites of alternative splicing were identified; 48 different splice variants could be generated by the differential exon usage observed. The number of splice forms and their relative frequency in vivo were characterized in RNA samples of both embryos and adults. The range of splice types was found to be much more diverse in adults than in embryos; of a total of 19 different combinations of alternative exons, 11 splice types were found in embryos and 18 in adults. Usage of some individual alternative exons changed during development; a newly identified exon, which is found in one of two forms either 24 or 30 base pairs long, was present in about 85% of para transcripts from embryos but only 7% of those in adults. These data suggest that a wide variety of subtly distinct Na channel isoforms are present in Drosophila, and that these may provide a range of voltage-gated sodium channel functions. Although multiple sodium channel genes have already been described in both Drosophila and mammalian systems, this study provides a clear indication that sodium channel variability may be much greater than previously thought.

Aging↗
PubMed1992

Effects of kinesin mutations on neuronal functions.

Kinesin is believed to generate force for the movement of organelles in anterograde axonal transport. The identification of genes that encode kinesin-like proteins suggests that other motors may provide anterograde force instead of or in addition to kinesin. To gain insight into the specific functions of kinesin, the effects of mutations in the kinesin heavy chain gene (khc) on the physiology and ultrastructure of Drosophila larval neurons were studied. Mutations in khc impair both action potential propagation in axons and neurotransmitter release at nerve terminals but have no apparent effect on the concentration of synaptic vesicles in nerve terminal cytoplasm. Thus kinesin is required in vivo for normal neuronal function and may be active in the transport of ion channels and components of the synaptic release machinery to their appropriate cellular locations. Kinesin appears not to be required for the anterograde transport of synaptic vesicles or their components.

Action Potentials↗
PubMed1992

Analysis of repolarization of presynaptic motor terminals in Drosophila larvae using potassium-channel-blocking drugs and mutations.

In Drosophila melanogaster muscles and neuronal cell bodies at least four different potassium currents have been identified whose activity shapes the electrical properties of these cells. Potassium currents also control repolarization of presynaptic terminals and, therefore, exert a major effect on transmitter release and synaptic plasticity. However, because of the small size of presynaptic terminals in Drosophila, it has not been possible to analyze the potassium currents they express. As a first approach to characterizing the ionic currents present at presynaptic motor terminals of Drosophila larvae, we recorded synaptic currents at the neuromuscular junction. From the alterations in evoked synaptic currents caused by various drugs and by mutations known to affect potassium currents in other tissues, we suggest that the repolarizing mechanism in presynaptic terminals consists of at least four distinct currents. One is affected by aminopyridines or Sh mutations, a second component is affected by the slo mutation, a third is sensitive to quinidine and one or more additional components are blocked by tetraethylammonium. Depolarization depends on a presynaptic calcium current, which displays only slight voltage-dependent inactivation. Because the mechanism of repolarization exerts a major effect on synaptic activity, this analysis provides a framework for further genetic and molecular dissection of the basic processes involved in the regulation of transmitter release.

Animals↗
PubMed1992

Identification and characterization of inebriated, a gene affecting neuronal excitability in Drosophila.

On the basis of behavioral interactions with mutations in a potassium channel gene of Drosophila--Shaker (Sh)--we have isolated mutations in a new gene called inebriated (ine). In a wildtype background, ine mutants display no observable behavioral defects. However, in a Sh mutant background, ine mutations cause downturned wings and an indented thorax. This distinctive phenotype is also exhibited by flies of other genotypes that cause extreme neuronal hyperexcitability. We utilized the potassium channel blocking drugs quinidine and dideoxy forskolin (DDF) to test the effects of ine on synaptic transmission. DDF and ine mutations each potentiated the effects of quinidine on synaptic transmission, but neither had any observable effects in the absence of quinidine. Application of DDF to ine mutants had no effects either in the presence or absence of quinidine. We conclude that ine mutations increase neuronal membrane excitability and perhaps block a DDF-sensitive potassium channel.

Animals↗
PubMed1991

The maleless protein associates with the X chromosome to regulate dosage compensation in Drosophila.

The maleless (mle) gene is one of four known regulatory loci required for increased transcription (dosage compensation) of X-linked genes in D. melanogaster males. A predicted mle protein (MLE) contains seven short segments that define a superfamily of known and putative RNA and DNA helicases. MLE, while present in the nuclei of both male and female cells, differs in its association with polytene X chromosomes in the two sexes. MLE is associated with hundreds of discrete sites along the length of the X chromosome in males and not in females. The predominant localization of MLE to the X chromosome in males makes it a strong candidate to be a direct regulator of dosage compensation.

Amino Acid Sequence↗
PubMed1991

napts, a mutation affecting sodium channel activity in Drosophila, is an allele of mle, a regulator of X chromosome transcription.

napts is a recessive mutation that affects the level of sodium channel activity and, at high temperature, causes paralysis associated with a loss of action potentials. We show, by genetic complementation tests, germline transformation, and analysis of mutations, that napts is a gain-of-function mutation of mle, a gene required for X chromosome dosage compensation and male viability. Molecular analyses of nap and mle mutations indicate that mle+, nap+, and napts activities are encoded by the same open reading frame and suggest that napts is due to a single amino acid substitution. Although napts is known to act via para+, an X-linked sodium channel structural gene, its effect is not due to a simple defect in para+ dosage compensation.

Action Potentials↗