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Y N Jan

Publications and source records attributed to Y N Jan.

At least 73 records · Page 4Linked to original sources
PubMed1998

Transcriptional regulation of atonal during development of the Drosophila peripheral nervous system.

atonal is a proneural gene for the development of Drosophila chordotonal organs and photoreceptor cells. We show here that atonal expression is controlled by modular enhancer elements located 5' or 3' to the atonal-coding sequences. During chordotonal organ development, the 3' enhancer directs expression in proneural clusters; whereas successive modular enhancers located in the 5' region drive tissue-specific expression in chordotonal organ precursors in the embryo and larval leg, wing and antennal imaginal discs. Similarly, in the eye disc, the 3' enhancer directs initial expression in a stripe anterior to the morphogenetic furrow. These atonal-expressing cells are then patterned through a Notch-dependent process into initial clusters, representing the earliest patterning event yet identified during eye morphogenesis. A distinct 5' enhancer drives expression in intermediate groups and R8 cells within and posterior to the morphogenetic furrow. Both enhancers are required for normal atonal function in the eye. The 5' enhancer, but not the 3' enhancer, depends on endogenous atonal function, suggesting a switch from regulation directed by other upstream genes to atonal autoregulation during the process of lateral inhibition. The regulatory regions identified in this study can thus account for atonal expression in every tissue and essentially in every stage of its expression during chordotonal organ and photoreceptor development.

Animals↗
PubMed1997

Voltage-gated and inwardly rectifying potassium channels.

This lecture is dedicated to Max Delbrück and Seymour Benzer. Max Delbrück was our graduate advisor. He introduced us to a variety of biophysical problems, and taught us ways of thinking about these problems by example. Potassium channels was one of the topics included in his journal club in the early seventies; Max also carefully considered the feasibility of purifying potassium channels then. It was in Seymour Benzer's laboratory that we began to look for Drosophila mutants that affect synaptic transmission at the larval neuromuscular junction. Shaker was the first behavioural mutant we tested that gave a robust phenotype, a phenotype that could be mimicked by treating wild-type preparations with a potassium channel blocker. This mutant fly has led us to our subsequent molecular studies of potassium channels. Since we settled in the University of California, San Francisco, and began to study neural development as well as potassium channels, we have settled into the pattern of each attending meetings and presenting our studies on one of these two areas so as to avoid both being away from home and our children at the same time. In following this pattern, I will be presenting the studies of potassium channels as part of our long-term collaboration. In this talk I will first briefly take you through the path that led us to the molecular studies of potassium channels and then discuss the diversity and modulation of these potassium channels at the molecular and physiological level.

Animals↗
PubMed1997

Genghis Khan (Gek) as a putative effector for Drosophila Cdc42 and regulator of actin polymerization.

The small GTPases Cdc42 and Rac regulate a variety of biological processes, including actin polymerization, cell proliferation, and JNK/mitogen-activated protein kinase activation, conceivably via distinct effectors. Whereas the effector for mitogen-activated protein kinase activation appears to be p65PAK, the identity of effector(s) for actin polymerization remains unclear. We have found a putative effector for Drosophila Cdc42, Genghis Khan (Gek), which binds to Dcdc42 in a GTP-dependent and effector domain-dependent manner. Gek contains a predicted serine/threonine kinase catalytic domain that is 63% identical to human myotonic dystrophy protein kinase and has protein kinase activities. It also possesses a large coiled-coil domain, a putative phorbol ester binding domain, a pleckstrin homology domain, and a Cdc42 binding consensus sequence that is required for its binding to Dcdc42. To study the in vivo function of gek, we generated mutations in the Drosophila gek locus. Egg chambers homozygous for gek mutations exhibit abnormal accumulation of F-actin and are defective in producing fertilized eggs. These phenotypes can be rescued by a wild-type gek transgene. Our results suggest that this multidomain protein kinase is an effector for the regulation of actin polymerization by Cdc42.

Actins↗
PubMed1997

The N terminus of the Drosophila Numb protein directs membrane association and actin-dependent asymmetric localization.

Drosophila Numb is a membrane associated protein of 557 amino acids (aa) that localizes asymmetrically into a cortical crescent in mitotic neural precursor cells and segregates into one of the daughter cells, where it is required for correct cell fate specification. We demonstrate here that asymmetric localization but not membrane localization of Numb in Drosophila embryos is inhibited by latrunculin A, an inhibitor of actin assembly. We also show that deletion of either the first 41 aa or aa 41-118 of Numb eliminates both localization to the cell membrane and asymmetric localization during mitosis, whereas C-terminal deletions or deletions of central portions of Numb do not affect its subcellular localization. Fusion of the first 76 or the first 119 aa of Numb to beta-galactosidase results in a fusion protein that localizes to the cell membrane, but fails to localize asymmetrically during mitosis. In contrast, a fusion protein containing the first 227 aa of Numb and beta-galactosidase localizes asymmetrically during mitosis and segregates into the same daughter cell as the endogenous Numb protein, demonstrating that the first 227 aa of the Numb protein are sufficient for asymmetric localization.

Animals↗
PubMed1997

Defective gamma-aminobutyric acid type B receptor-activated inwardly rectifying K+ currents in cerebellar granule cells isolated from weaver and Girk2 null mutant mice.

Stimulation of inhibitory neurotransmitter receptors, such as gamma-aminobutyric acid type B (GABAB) receptors, activates G protein-gated inwardly rectifying K+ channels (GIRK) which, in turn, influence membrane excitability. Seizure activity has been reported in a Girk2 null mutant mouse lacking GIRK2 channels but showing normal cerebellar development as well as in the weaver mouse, which has mutated GIRK2 channels and shows abnormal development. To understand how the function of GIRK2 channels differs in these two mutant mice, we compared the G protein-activated inwardly rectifying K+ currents in cerebellar granule cells isolated from Girk2 null mutant and weaver mutant mice with those from wild-type mice. Activation of GABAB receptors in wild-type granule cells induced an inwardly rectifying K+ current, which was sensitive to pertussis toxin and inhibited by external Ba2+ ions. The amplitude of the GABAB receptor-activated current was severely attenuated in granule cells isolated from both weaver and Girk2 null mutant mice. By contrast, the G protein-gated inwardly rectifying current and possibly the agonist-independent basal current appeared to be less selective for K+ ions in weaver but not Girk2 null mutant granule cells. Our results support the hypothesis that a nonselective current leads to the weaver phenotype. The loss of GABAB receptor-activated GIRK current appears coincident with the absence of GIRK2 channel protein and the reduction of GIRK1 channel protein in the Girk2 null mutant mouse, suggesting that GABAB receptors couple to heteromultimers composed of GIRK1 and GIRK2 channel subunits.

Animals↗
PubMed1997

Miranda is required for the asymmetric localization of Prospero during mitosis in Drosophila.

Asymmetric division of Drosophila neuroblasts, sensory organ precursor cells, and cells in the procephalic neurogenic region involves the segregation of Numb and Prospero proteins into one of the two daughter cells. We have isolated a novel gene, miranda, based on the ability of its gene product to interact with the Prospero asymmetric localization domain. miranda expression coincides spatially and temporally with asymmetric cell divisions and asymmetric localization of Prospero. Miranda protein is localized asymmetrically, along with Prospero, to the basal cell membrane during mitosis. Loss of miranda gene function abolishes asymmetric Prospero localization during mitosis. The asymmetric localization of Miranda protein requires inscuteable. Our results suggest that miranda functions downstream of inscuteable and works as an adapter that connects Prospero to the basal cell membrane during asymmetric cell division.

Amino Acid Sequence↗
PubMed1997

Regulation of IRK3 inward rectifier K+ channel by m1 acetylcholine receptor and intracellular magnesium.

Inward rectifier K+ channels control the cell's membrane potential and neuronal excitability. We report that the IRK3 but not the IRK1 inward rectifier K+ channel activity is inhibited by m1 muscarinic acetylcholine receptor. This m1 modulation cannot be accounted for by protein kinase C, Ca2+, or channel phosphorylation, but can be mimicked by Mg2+. Based on quantitative analyses of IRK3 and two different IRK1 mutant channels bestowed with sensitivity to m1 modulation, we suggest that the resting Mg2+ level causes chronic inhibition of IRK3 channels, and m1 receptor stimulation may lead to an increase of cytoplasmic Mg2+ concentration and further channel inhibition, due to the ability of Mg2+ to lead these channels into a prolonged inactivated state.

Amino Acid Sequence↗
PubMed1997

Scanning mutagenesis of the putative transmembrane segments of Kir2.1, an inward rectifier potassium channel.

Structural models of inward rectifier K+ channels incorporate four identical or homologous subunits, each of which has two hydrophobic segments (M1 and M2) which are predicted to span the membrane as alpha helices. Since hydrophobic interactions between proteins and membrane lipids are thought to be generally of a nonspecific nature, we attempted to identify lipid-contacting residues in Kir2.1 as those which tolerate mutation to tryptophan, which has a large hydrophobic side chain. Tolerated mutations were defined as those which produced measurable inwardly rectifying currents in Xenopus oocytes. To distinguish between water-accessible positions and positions adjacent to membrane lipids or within the protein interior we also mutated residues in M1 and M2 individually to aspartate, since an amino acid with a charged side chain should not be tolerated at lipid-facing or interior positions, due to the energy cost of burying a charge in a hydrophobic environment. Surprisingly, 17 out of 20 and 17 out of 22 non-tryptophan residues in M1 and M2, respectively, tolerated being mutated to tryptophan. Moreover, aspartate was tolerated at 15 out of 22 and 15 out of 21 non-aspartate M1 and M2 positions respectively. Periodicity in the pattern of tolerated vs. nontolerated mutations consistent with alpha helices or beta strands did not emerge convincingly from these data. We consider the possibility that parts of M1 and M2 may be in contact with water.

Alanine↗
PubMed1997

Binding of the G protein betagamma subunit to multiple regions of G protein-gated inward-rectifying K+ channels.

We have previously shown that direct binding of the betagamma subunit of G protein (G betagamma) to both the N-terminal domain and the C-terminal domain of a cloned G protein-gated inward-rectifying K+ channel subunit, GIRK1, is important for channel activation. We have now further localized the G betagamma binding region in the N-terminal domain of GIRK1 to amino acids 34-86 and the G betagamma binding region in the C-terminal domain of GIRK1 to two separate fragments of amino acids 318-374 and amino acids 390-462. Of the four cloned mammalian GIRK subunits, GIRK1-4, GIRK1 and 4 form heteromeric K+ channels in the heart and similar channels in the brain include heteromultimers of GIRK1 and 2, and possibly other GIRK homomultimers and heteromultimers. We found that the N-terminal and the C-terminal domains of all four GIRKs bound G betagamma. The G betagamma binding activities for the C-terminal domains of GIRK2-4 were lower than that for the C-terminal domain of GIRK1. The higher G betagamma binding activity for the C-terminal domain of GIRK1 is due to amino acids 390-462 which are unique to GIRK1. We also found that the N-terminal and C-terminal domains of GIRKs interacted with each other, and the N-terminal domain of either GIRK1 or GIRK4 together with the C-terminal domain of GIRK1 exhibited much enhanced binding of G betagamma. These results are consistent with the idea that the N- and C-terminal domains of the cardiac G protein-gated K+ channel subunits may interact with each other to form higher affinity binding site(s) for G betagamma.

Amino Acid Sequence↗
PubMed1997

Requirement for EGF receptor signalling in neural recruitment during formation of Drosophila chordotonal sense organ clusters.

BACKGROUND: Drosophila proneural genes act in the process of selecting neural precursors from undifferentiated ectoderm. The proneural gene atonal is required for the development of precursors of both chordotonal organs (stretch receptors) and photoreceptors. Although these types of sensory element are dissimilar in structure and function, they both occur as organized arrays of neurons. Previous studies have shown that clustering of photoreceptors involves local recruitment, and that signalling by the Drosophila epidermal growth factor receptor (DER) pathway is involved in the recruitment process. We present evidence that a similar mechanism is required for the clustering of embryonic chordotonal organs. RESULTS: We have examined the expression patterns of atonal and genes of the DER pathway in wild-type and mutant backgrounds. Expression of atonal was restricted to a subset of the atonal-requiring chordotonal precursors, which we call founder precursors. The remaining precursors required DER signalling for their selection. Signalling by the founder precursors was initiated by atonal activating, directly or indirectly, rhomboid expression in these cells. Signalling by these founder precursors then provoked a response in the surrounding ectodermal cells, as shown by the activation of expression of the DER target genes pointed and argos. The signal and response then led to recruitment of some of the ectodermal cells to the chordotonal precursor cell fate. DER hyperactivation by misexpression of rhomboid resulted in excessive chordotonal precursor recruitment. CONCLUSIONS: Increased numbers of chordotonal precursors are recruited by homeogenetic induction involving signalling via DER from founder precursors to surrounding ectodermal cells. We suggest that the reason chordotonal organs and photoreceptors share a requirement for the proneural gene atonal is that this gene activates a common pathway leading to neural aggregation.

Animals↗
PubMed1997

Stabilization of ion selectivity filter by pore loop ion pairs in an inwardly rectifying potassium channel.

Ion selectivity is critical for the biological functions of voltage-dependent cation channels and is achieved by specific ion binding to a pore region called the selectivity filter. In voltage-gated K+, Na+ and Ca2+ channels, the selectivity filter is formed by a short polypeptide loop (called the H5 or P region) between the fifth and sixth transmembrane segments, donated by each of the four subunits or internal homologous domains forming the channel. While mutagenesis studies on voltage-gated K+ channels have begun to shed light on the structural organization of this pore region, little is known about the physical and chemical interactions that maintain the structural stability of the selectivity filter. Here we show that in an inwardly rectifying K+ (IRK) channel, IRK1, short range interactions of an ion pair in the H5 pore loop are crucial for pore structure and ion permeation. The two residues, a glutamate and an arginine, appear to form exposed salt bridges in the tetrameric channel. Alteration or disruption of such ion pair interactions dramatically alters ion selectivity and permeation. Since this ion pair is conserved in all IRK channels, it may constitute a general mechanism for maintaining the stability of the pore structure in this channel superfamily.

Amino Acid Sequence↗
PubMed1997

Xath5 participates in a network of bHLH genes in the developing Xenopus retina.

We examined the function of basic-helix-loop-helix (bHLH) transcription factors during retinal neurogenesis. We identified Xath5, a Xenopus bHLH gene related to Drosophila atonal, which is expressed in the developing Xenopus retina. Targeted expression of Xath5 in retinal progenitor cells biased the differentiation of these cells toward a ganglion cell fate, suggesting that Xath5 can regulate the differentiation of retinal neurons. We examined the relationship between the three bHLH genes Xash3, NeuroD, and Xath5 during retinal neurogenesis and found that Xash3 is expressed in early retinoblasts, followed by coexpression of Xath5 and NeuroD in differentiating cells. We provide evidence that the expression of Xash3, NeuroD, and Xath5 is coupled and propose that these bHLH genes regulate successive stages of neuronal differentiation in the developing retina.

Aging↗
PubMed1997

Receptor-regulated ion channels.

Recent advances in the study of receptor-regulated ion channels include the cloning of the genes encoding three types of potassium channel that are favorite targets of receptors for transmitters and hormones. Studies of these channels have also provided a strong indication that G-protein betagamma subunits may gate ion channels via direct protein-protein interactions. Similarities between channel regulation by natriuretic peptides and channel regulation by secreted peptide products of the Alzheimer's beta-amyloid precursor protein offer hints for the existence of a receptor for the latter. There are also other novel examples of channel regulation in excitable and nonexcitable cells, including liver cells and blood cells.

Animals↗
PubMed1997

Rho family GTP-binding proteins in growth cone signalling.

Rho family GTP-binding proteins regulate various aspects of the actin cytoskeleton in a wide variety of organisms. Recent evidence suggests that they may also be important components of the signalling pathways that link the reception of extracellular cues to the regulation of the cytoskeleton in neuronal growth cones.

Actins↗