bluuebunny/crossref_metadata_2025 huggingface dataset:
The Invulnerability of Adult Neurons: A Critical Role for p73
Authors: Walsh Gregory S. , Orike Nina , Kaplan David R. , Miller Freda D.
Published: October 2004

Here, we investigated the intracellular mechanisms that underlie the relative invulnerability of adult versus developing dorsal root ganglion (DRG) sensory neurons. In culture, adult neurons were resistant to stimuli that caused apoptosis of their neonatal counterparts. In both adult and neonatal neurons, death stimuli induced the apoptotic c-Jun N-terminal protein kinase (JNK) pathway, but JNK activation only caused death of neonatal neurons, indicating that adult neurons have a downstream block to apoptosis. Expression of the dominant-inhibitory p53 family member, ΔNp73, rescued JNK-induced apoptosis of neonatal neurons, suggesting that it might participate in the downstream apoptotic block in adult neurons. To test this possibility, we examined adult DRG neurons cultured from p73+/- mice. Adult p73+/- DRG neurons were more vulnerable to apoptotic stimuli than their p73+/+ counterparts, and invulnerability could be restored to the p73+/- neurons by increased expression of ΔNp73. Moreover, although DRG neuron development was normal in p73+/- animalsin vivo, axotomy caused death of adult p73+/- but not p73+/+ DRG neurons. Thus, one way adult neurons become invulnerable is to enhance endogenous survival pathways, and one critical component of these survival pathways is p73.
Neurofilament Transport Is Dependent on Actin and Myosin
Authors: Jung Cheolwha , Chylinski Teresa M. , Pimenta Aurea , Ortiz Daniela , Shea Thomas B.
Published: October 2004

Real-time analyses have revealed that some newly synthesized neurofilament (NF) subunits translocate into and along axonal neurites by moving along the inner plasma membrane surface, suggesting that they may translocate against the submembrane actin cortex. We therefore examined whether or not NF axonal transport was dependent on actin and myosin. Perturbation of filamentous actin in NB2a/d1 cells with cytochalasin B inhibited translocation of subunits into axonal neurites and inhibited bidirectional translocation of NF subunits within neurites. Intravitreal injection of cytochalasin B inhibited NF axonal transport in optic axons in a dose-response manner. NF subunits were coprecipitated from NB2a/d1 cells by an anti-myosin antibody, and myosin colocalized with NFs in immunofluorescent analyses. The myosin light chain kinase inhibitor ML-7 and the myosin ATPase inhibitor 2,3-butanedione-2-monoxime perturbed NF translocation within NB2a/d1 axonal neurites. These findings suggest that some NF subunits may undergo axonal transport via myosin-mediated interactions with the actin cortex.
Enhanced Accuracy in Novel Mirror Drawing after Repetitive Transcranial Magnetic Stimulation-Induced Proprioceptive Deafferentation
Authors: Balslev Daniela , Christensen Lars O. D. , Lee Ji-Hang , Law Ian , Paulson Olaf B. , Miall R. Christopher
Published: October 2004

When performing visually guided actions under conditions of perturbed visual feedback, e.g., in a mirror or a video camera, there is a spatial conflict between visual and proprioceptive information. Recent studies have shown that subjects without proprioception avoid this conflict and show a performance benefit. In this study, we tested whether deafferentation induced by repetitive transcranial magnetic stimulation (rTMS) can improve mirror tracing skills in normal subjects. Hand trajectory error during novel mirror drawing was compared across two groups of subjects that received either 1 Hz rTMS over the somatosensory cortex contralateral to the hand or sham stimulation. Mirror tracing was more accurate after rTMS than after sham stimulation. Using a position-matching task, we confirmed that rTMS reduced proprioceptive acuity and that this reduction was largest when the coil was placed at an anterior parietal site. It is thus possible, with rTMS, to enhance motor performance in tasks involving a visuoproprioceptive conflict, presumably by reducing the excitability of somatosensory cortical areas that contribute to the sense of hand position.
Metamodulation of a Spinal Locomotor Network by Nitric Oxide
Authors: McLean David L. , Sillar Keith T.
Published: October 2004

Flexibility in the output of spinal networks can be accomplished by the actions of neuromodulators; however, little is known about how the process of neuromodulation itself may be modulated. Here we investigate the potential “meta”-modulatory hierarchy between nitric oxide (NO) and noradrenaline (NA) inXenopus laevistadpoles. NO and NA have similar effects on fictive swimming; both potentiate glycinergic inhibition to slow swimming frequency and GABAergic inhibition to reduce episode durations. In addition, both modulators have direct effects on the membrane properties of motor neurons. Here we report that antagonism of noradrenergic pathways with phentolamine dramatically influences the effect of the NO donorS-nitroso-N-acetylpenicillamine (SNAP) on swimming frequency, but not its effect on episode durations. In contrast, scavenging extracellular NO with 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (PTIO) does not influence any of the effects of NA on fictive swimming. These data place NO above NA in the metamodulatory hierarchy, strongly suggesting that NO works via a noradrenergic pathway to control glycine release but directly promotes GABA release. We confirmed this possibility using intracellular recordings from motor neurons. In support of a natural role for NO in theXenopuslocomotor network, PTIO not only antagonized all of the effects of SNAP on swimming but also, when applied on its own, modulated both swimming frequency and episode durations in addition to the underlying glycinergic and GABAergic pathways. Collectively, our results illustrate that NO and NA have parallel effects on motor neuron membrane properties and GABAergic inhibition, but that NO serially metamodulates glycinergic inhibition via NA.
Cross Talk between Metabotropic and Ionotropic Glutamate Receptor-Mediated Signaling in Parallel Fiber-Induced Inositol 1,4,5-Trisphosphate Production in Cerebellar Purkinje Cells
Authors: Okubo Yohei , Kakizawa Sho , Hirose Kenzo , Iino Masamitsu
Published: October 2004

In many excitatory glutamatergic synapses, both ionotropic glutamate receptors (iGluRs) and metabotropic glutamate receptors (mGluRs) are closely distributed on the postsynaptic membrane. However, the functional significance of the close distribution of the two types of glutamate receptors has not been fully clarified. In this study, we examined the functional interaction between iGluR and mGluR at parallel fiber (PF)→ Purkinje cell synapses in the generation of inositol 1,4,5-trisphosphate (IP3), a key second messenger that regulates many important cellular functions. We visualized local IP3dynamics in Purkinje cells using the green fluorescent protein-tagged pleckstrin homology domain (GFP-PHD) as a fluorescent IP3probe. Purkinje cells were transduced with Sindbis virus encoding GFP-PHD and imaged with a two-photon laser scanning microscope. Translocation of GFP-PHD from the plasma membrane to the cytoplasm attributable to an increase in IP3concentration was observed on PF stimulation in fine dendrites of Purkinje cells. Surprisingly, this PF-induced IP3production was blocked not only by the group I mGluR antagonist but also by the AMPA receptor (AMPAR) antagonist. The PF-induced IP3production was blocked by either the inhibition of G-protein activation by GDP-βS or intracellular Ca2+buffering by BAPTA. These results show that IP3production is mediated cooperatively by group I mGluR and AMPAR through G-protein activation and Ca2+influx at PF→ Purkinje cell synapses, identifying the robust cross talk between iGluR and mGluR for the generation of IP3signals.
Experience and Activity-Dependent Maturation of Perisomatic GABAergic Innervation in Primary Visual Cortex during a Postnatal Critical Period
Authors: Chattopadhyaya Bidisha , Di Cristo Graziella , Higashiyama Hiroyuki , Knott Graham W. , Kuhlman Sandra J. , Welker Egbert , Huang Z. Josh
Published: October 2004

The neocortical GABAergic network consists of diverse interneuron cell types that display distinct physiological properties and target their innervations to subcellular compartments of principal neurons. Inhibition directed toward the soma and proximal dendrites is crucial in regulating the output of pyramidal neurons, but the development of perisomatic innervation is poorly understood because of the lack of specific synaptic markers. In the primary visual cortex, for example, it is unknown whether, and to what extent, the formation and maturation of perisomatic synapses are intrinsic to cortical circuits or are regulated by sensory experience. Using bacterial artificial chromosome transgenic mice that label a defined class of perisomatic synapses with green fluorescent protein, here we show that perisomatic innervation developed during a protracted postnatal period after eye opening. Maturation of perisomatic innervation was significantly retarded by visual deprivation during the third, but not the fifth, postnatal week, implicating an important role for sensory input. To examine the role of cortical intrinsic mechanisms, we developed a method to visualize perisomatic synapses from single basket interneurons in cortical organotypic cultures. Characteristic perisomatic synapses formed through a stereotyped process, involving the extension of distinct terminal branches and proliferation of perisomatic boutons. Neuronal spiking in organotypic cultures was necessary for the proliferation of boutons and the extension, but not the maintenance, of terminal branches. Together, our results suggest that although the formation of perisomatic synapses is intrinsic to the cortex, visual experience can influence the maturation and pattern of perisomatic innervation during a postnatal critical period by modulating the level of neural activity within cortical circuits.
Glutamate Mediates Acute Glucose Transport Inhibition in Hippocampal Neurons
Authors: Porras Omar H. , Loaiza Anitsi , Barros L. Felipe
Published: October 2004

Although it is known that brain activity is fueled by glucose, the identity of the cell type that preferentially metabolizes the sugar remains elusive. To address this question, glucose uptake was studied simultaneously in cultured hippocampal neurons and neighboring astrocytes using a real-time assay based on confocal epifluorescence microscopy and fluorescent glucose analogs. Glutamate, although stimulating glucose transport in astrocytes, strongly inhibited glucose transport in neurons, producing in few seconds a 12-fold increase in the ratio of astrocytic-to-neuronal uptake rate. Neuronal transport inhibition was reversible on removal of the neurotransmitter and displayed an IC50of 5 μm, suggesting its occurrence at physiological glutamate concentrations. The phenomenon was abolished by CNQX and mimicked by AMPA, demonstrating a role for the cognate subset of ionotropic glutamate receptors. Transport inhibition required extracellular sodium and calcium and was mimicked by veratridine but not by membrane depolarization with high K+or by calcium overloading with ionomycin. Therefore, glutamate inhibits glucose transport via AMPA receptor-mediated sodium entry, whereas calcium entry plays a permissive role. This phenomenon suggests that glutamate redistributes glucose toward astrocytes and away from neurons and represents a novel molecular mechanism that may be important for functional imaging of the brain using positron emission tomography.
Cell-Specific Expression and Lipopolysaccharide-Induced Regulation of Tumor Necrosis Factor α (TNFα) and TNF Receptors in Rat Dorsal Root Ganglion
Authors: Li Yanzhang , Ji Ailing , Weihe Eberhard , Schäfer Martin K.-H.
Published: October 2004

The proinflammatory and lipopolysaccharide (LPS)-inducible cytokine tumor necrosis factor α (TNFα) has been shown to enhance primary sensory nociceptive signaling. However, the precise cellular sites of TNFα and TNF receptor synthesis are still a matter of controversy. Therefore, we differentiated the neuronal and non-neuronal sites of TNFα, TNFR1, and TNFR2 mRNA synthesis in dorsal root ganglion (DRG) of control rats and evaluated how their expression is altered under systemic challenge with LPS.In situhybridization (ISH), RT-PCR analysis of laser-microdissected cells, and immunocytochemistry revealed absence of TNFα from DRG neurons and LPS-induced expression of TNFα exclusively in a subpopulation of non-neuronal DRG cells. Using RT-PCR and Northern blotting TNFR1 and TNFR2 mRNAs were found to be constitutively expressed and increased after LPS. TNFR1 mRNA was expressed in virtually all neurons and in non-neuronal cells with increased levels after LPS in both. TNFR2 was exclusively expressed and regulated in non-neuronal cells. RT-PCR analysis of microdissected DRG neurons and of the sensory neuronal cell line F11 confirmed the neuronal expression of TNFR1 and excluded that of TNFR2. Double ISH revealed varying levels of TNFR1 mRNA in virtually all DRG neurons including putative nociceptive neurons coding for calcitonin gene-related peptide, substance P, or vanilloid receptor 1. Taken together, we provide evidence that non-neuronally synthesized TNFα may directly act on primary afferent neurons via TNFR1 but not TNFR2. This is likely to be relevant under conditions of inflammatory pain and infections accompanied by widespread TNFα synthesis and release and may drive sickness behavior.
Visual Experience Is Necessary for the Development of Multisensory Integration
Authors: Wallace Mark T. , Perrault Thomas J. , Hairston W. David , Stein Barry E.
Published: October 2004

Multisensory neurons and their ability to integrate multisensory cues develop gradually in the midbrain [i.e., superior colliculus (SC)]. To examine the possibility that early sensory experiences might play a critical role in these maturational processes, animals were raised in the absence of visual cues. As adults, the SC of these animals were found to contain many multisensory neurons, the large majority of which were visually responsive. Although these neurons responded robustly to each of their cross-modal inputs when presented individually, they were incapable of synthesizing this information. These observations suggest that visual experiences are critical for the SC to develop the ability to integrate multisensory information and lead to the prediction that, in the absence of such experience, animals will be compromised in their sensitivity to cross-modal events.
Classical Photoreceptors RegulateMelanopsinmRNA Levels in the Rat Retina
Authors: Sakamoto Katsuhiko , Liu Cuimei , Tosini Gianluca
Published: October 2004

Recent studies have demonstrated that melanopsin is a key photopigment in the mammalian circadian system. This novel opsin is exclusively expressed in retinal ganglion cells that are intrinsically sensitive to light, perhaps responding via a melanopsin-based signaling pathway. Previous investigations using transgenic mice have also demonstrated that ablation of the classical photoreceptors and of melanopsin prevents entrainment of several circadian rhythms, thus demonstrating that these photoreceptors are necessary and sufficient for circadian photoreception. In this study, we investigated the effect of photoreceptor degeneration onmelanopsinmRNA regulation in RCS/N-rdyrats (Royal College of Surgeons rats with a defect in the retinal dystrophy gene). We used animals at postnatal day 21 (P21), P33, P45, and P60. At P60 degeneration of the retina in RCS/N-rdyhas advanced to the point where the majority of the photoreceptors have degenerated. Our data indicate thatmelanopsinmRNA levels were rhythmic in light/dark cycle and in constant darkness in congenic controls (RCS/N-rdy+) and in RCS/N-rdyat P21 (i.e., before the degeneration of the photoreceptors). On the other hand, in RCS/N-rdyat P60,melanopsinmRNA levels were greatly reduced (<90%) and not rhythmic. Photoreceptor degeneration did not affect the expression ofpituitary adenylate cyclase-activating polypeptidemRNA (a marker for melanopsin-containing ganglion cells). Our results suggest that classical photoreceptors (rods and cones) regulate the expression ofmelanopsinmRNA in the rat. Because RCS/N-rdyrats are a model for studies on retinitis pigmentosa in human, our data may provide an important insight on melanopsin function in patients affected by retinitis pigmentosa.
Na-K-Cl Cotransporter-Mediated Intracellular Na+Accumulation Affects Ca2+Signaling in Astrocytes in anIn VitroIschemic Model
Authors: Lenart Brett , Kintner Douglas B. , Shull Gary E. , Sun Dandan
Published: October 2004

Na-K-Cl cotransporter isoform 1 (NKCC1) plays an important role in maintenance of intracellular Na+, K+, and Cl-levels in astrocytes. We propose that NKCC1 may contribute to perturbations of ionic homeostasis in astrocytes under ischemic conditions. After 3-8 hr of oxygen and glucose deprivation (OGD), NKCC1-mediated86Rb influx was significantly increased in astrocytes from NKCC1 wild-type (NKCC1+/+) and heterozygous mutant (NKCC1+/-) mice. Phosphorylated NKCC1 protein was increased in NKCC1+/+astrocytes at 2 hr of OGD. Two hours of OGD and 1 hr of reoxygenation (OGD/REOX) triggered an ∼3.6-fold increase in intracellular Na+concentration ([Na+]i) in NKCC1+/+astrocytes. Inhibition of NKCC1 activity by bumetanide or ablation of the NKCC1 gene significantly attenuated the rise in [Na+]i. Moreover, NKCC1+/+astrocytes swelled by 10-30% during 20-60 min of OGD. Either genetic ablation of NKCC1 or inhibition of NKCC1 by bumetanide-attenuated OGD-mediated swelling. An NKCC1-mediated increase in [Na+]imay subsequently affect Ca2+signaling through the Na+/Ca2+exchanger (NCX). A rise in [Ca2+]iwas detected after OGD/REOX in the presence of a sarcoplasmic-endoplasmic reticulum (ER) Ca2+-ATPase inhibitor thapsigargin. Moreover, OGD/REOX led to a significant increase in Ca2+release from ER Ca2+stores. Furthermore, KB-R7943 (2-[2-[4(4-nitrobenzyloxy)phenyl]ethyl]isothiourea mesylate), an inhibitor of reverse-mode operation of NCX, abolished the OGD/REOX-induced enhancement in filling of ER Ca2+stores. OGD/REOX-mediated Ca2+accumulation in ER Ca2+stores was absent when NKCC1 activity was ablated or pharmacologically inhibited. These findings imply that stimulation of NKCC1 activity leads to Na+accumulation after OGD/REOX and that subsequent reverse-mode operation of NCX contributes to increased Ca2+accumulation by intracellular Ca2+stores.
Corticotropin-Releasing Hormone Directly Activates Noradrenergic Neurons of the Locus Ceruleus RecordedIn Vitro
Authors: Jedema Hank P. , Grace Anthony A.
Published: October 2004

The neuropeptide corticotropin-releasing hormone (CRH) activates locus ceruleus (LC) neurons, thereby increasing norepinephrine levels throughout the CNS. Despite anatomical and physiological evidence for CRH innervation of the LC, the mechanism of CRH-evoked activation of LC neurons is unknown. Moreover, given the apparent absence of mRNA for CRH receptors in LC neurons, the exact location of action of CRH within the cerulear region is debated. Usingin vitrointracellular recordings from rat brainstem, we examined whether CRH exerts a direct effect on LC neurons and which ionic currents are likely affected by CRH. We demonstrate that CRH dose-dependently increases the firing rate of LC neurons through a direct (TTX- and cadmium-insensitive) mechanism by decreasing a potassium conductance. The CRH-evoked activation of LC neurons is, at least in part, mediated by CRH1receptors and a cAMP-dependent second messenger system. These data provide additional support that CRH functions as an excitatory neurotransmitter in the LC and the hypothesis that dysfunction of the CRH peptidergic and noradrenergic systems observed in patients with mood and anxiety disorders are functionally related.
Distinct Roles for the Kainate Receptor Subunits GluR5 and GluR6 in Kainate-Induced Hippocampal Gamma Oscillations
Authors: Fisahn André , Contractor Anis , Traub Roger D. , Buhl Eberhard H. , Heinemann Stephen F. , McBain Chris J.
Published: October 2004

Kainate receptors (KARs) play an important role in synaptic physiology, plasticity, and pathological phenomena such as epilepsy. However, the physiological implications for neuronal networks of the distinct expression patterns of KAR subunits are unknown. Using KAR knock-out mice, we show that subunits glutamate receptor (GluR) 5 and GluR6 play distinct roles in kainate-induced gamma oscillations and epileptiform burst activity. Ablation of GluR5 leads to a higher susceptibility of the network to the oscillogenic and epileptogenic effects of kainate, whereas lack of GluR6 prevents kainate-induced gamma oscillations or epileptiform bursts. Based on experimental and simulated neuronal network data as well as the consequences of GluR5 and GluR6 expression for cellular and synaptic physiology, we propose that the functional interplay of GluR5-containing KARs on axons of interneurons and GluR6-containing KARs in the somatodendritic region of both interneurons and pyramidal cells underlie the oscillogenic and epileptogenic effects of kainate.
Temporally Specific Burst in Cell Proliferation Increases Hippocampal Neurogenesis in Protracted Abstinence from Alcohol
Authors: Nixon Kimberly , Crews Fulton T.
Published: October 2004

Adult neurogenesis is a newly considered form of plasticity that could contribute to brain dysfunction in psychiatric disease. Chronic alcoholism, a disease affecting over 8% of the adult population, produces cognitive impairments and decreased brain volumes, both of which are partially reversed during abstinence. Clinical data and animal models implicate the hippocampus, a region important in learning and memory. In a model of alcohol dependence (chronic binge exposure for 4 d), we show that adult neurogenesis is inhibited during dependence with a pronounced increase in new hippocampal neuron formation after weeks of abstinence. This increase is attributable to a temporally and regionally specific fourfold increase in cell proliferation at day 7 of abstinence, with a majority of those cells surviving and differentiating at percentages similar to controls, effects that doubled the formation of new neurons. Although increases in cell proliferation correlated with alcohol withdrawal severity, proliferation remained increased when diazepam (10 mg/kg) was used to reduce withdrawal severity. Indeed, those animals with little withdrawal activity still show a twofold burst in cell proliferation at day 7 of abstinence. Thus, alcohol dependence and recovery from dependence continues to alter hippocampal plasticity during abstinence. Because neurogenesis may contribute to hippocampal function and/or learning, memory, and mood, compensatory neurogenesis and the return of normal neurogenesis may also have an impact on hippocampal structure and function. For the first time, these data provide a neurobiological mechanism that may underlie the return of human cognitive function and brain volume associated with recovery from addiction.
Asymmetrical Modes of Visual Bottom-Up and Top-Down Integration in the Thalamic Nucleus Rotundus of Pigeons
Authors: Folta Kristian , Diekamp Bettina , Güntürkün Onur
Published: October 2004

The aim of this study was to separate bottom-up and top-down influences within cerebral asymmetries. This was studied in the lateralized visual system of pigeons by recording from single units of the left and right diencephalic nucleus rotundus of the tectofugal pathway while visually stimulating the ipsilateral and/or contralateral eye. Analyses of response latencies revealed rotundal neurons with short and/or late response components. Cells with short latencies very likely represent bottom-up neurons participating in the ascending retinotectorotundal system. Because lidocaine injections into the visual Wulst produced a significant reduction of late response components only, neurons with long latencies were probably activated via a top-down telencephalotectorotundal system. The distribution and response characteristics of bottom-up and top-down neurons provided insight into several asymmetries of ascending and descending pathways. Asymmetries of the ascending retinotectorotundal system (bottom-up) were characterized by longer periods of tonic activation in the left and shorter response latencies in the right rotundus. Left-right differences in these responses probably facilitate faster access to visual input to the right hemisphere and a prolonged processing of this input in the left. The descending telencephalotectorotundal system (top-down) revealed a completely different lateralized organization. This system was characterized by long latency responses that exclusively derived from the left hemisphere, regardless of whether recordings took place in the left or the right rotundus. We assume that asymmetrical modes of visual processing within both hemispheres of the ascending tectofugal system are ultimately directed to left hemispheric forebrain mechanisms that subsequently generate executive control over sensory and motor structures.
Allelic Diversification at the C (OsC1) Locus of Wild and Cultivated Rice
Authors: Saitoh Kumi , Onishi Kazumitsu , Mikami Ichiho , Thidar Khin , Sano Yoshio
Published: October 2004

Divergent phenotypes are often detected in domesticated plants despite the existence of invariant phenotypes in their wild forms. One such example in rice is the occurrence of varying degrees of apiculus coloration due to anthocyanin pigmentation, which was previously reported to be caused by a series of alleles at the C locus. The present study reveals, on the basis of comparison of its maps, that the C gene appears to be the rice homolog (OsC1) of maize C1, which belongs to the group of R2R3-Myb factors. Two different types of deletions causing a frameshift were detected in the third exon, and both of the deleted nucleotides corresponded to the positions of putative base-contacting residues, suggesting that the Indica and Japonica types carry loss-of-function mutations with independent origins. In addition, replacement substitutions were frequently detected in OsC1 of strains carrying the previously defined C alleles. Molecular population analysis revealed that 17 haplotypes were found in 39 wild and cultivated rices, and the haplotypes of most cultivated forms could be classified into one of three distinct groups, with few shared haplotypes among taxa, including Indica and Japonica types. The genealogy of the OsC1 gene suggests that allelic diversification causing phenotypic change might have resulted from mutations in the coding region rather than from recombination between preexisting alleles. The McDonald and Kreitman test revealed that the changes in amino acids might be associated with selective forces acting on the lineage of group A whose haplotypes were carried by most Asian cultivated forms. The results regarding a significant implication for genetic diversity in landraces of rice are also discussed.
Effect of Two- and Three-Locus Linkage Disequilibrium on the Power to Detect Marker/Phenotype Associations
Authors: Nielsen Dahlia M , Ehm Margaret G , Zaykin Dmitri V , Weir Bruce S
Published: October 2004

There has been much recent interest in describing the patterns of linkage disequilibrium (LD) along a chromosome. Most empirical studies that have examined this issue have concentrated on LD between collections of pairs of markers and have not considered the joint effect of a group of markers beyond these pairwise connections. Here, we examine many different patterns of LD defined by both pairwise and joint multilocus LD terms. The LD patterns we considered were chosen in part by examining those seen in real data. We examine how changes in these patterns affect the power to detect association when performing single-marker and haplotype-based case-control tests, including a novel haplotype test based on contrasting LD between affected and unaffected individuals. Through our studies we find that differences in power between single-marker tests and haplotype-based tests in general do not appear to be large. Where moderate to high levels of multilocus LD exist, haplotype tests tend to be more powerful. Single-marker tests tend to prevail when pairwise LD is high. For moderate pairwise values and weak multilocus LD, either testing strategy may come out ahead, although it is also quite likely that neither has much power.
Drosophila Damaged DNA-Binding Protein 1 Is an Essential Factor for Development
Authors: Takata Kei-ichi , Yoshida Hideki , Yamaguchi Masamitsu , Sakaguchi Kengo
Published: October 2004

The damaged DNA-binding protein (DDB) complex, thought to recognize (6-4) photoproducts and other lesions in DNA, has been implicated to have a role in global genomic nucleotide excision repair (NER) and E2F-1-mediated transcription. The complex consists of a heterodimer of p127 (DDB1) and p48 (DDB2), the latter also being known as XPE. We reported previously that in Drosophila expression of the DDB1 (D-DDB1) gene is controlled by the DRE/DREF system, and external injury to DNA is not essential for D-DDB1 function. In the present study of the function of D-DDB1 in a multicellular system, we prepared transgenic flies, which were knocked down for the D-DDB1 gene due to RNA interference (RNAi), and performed immunocytochemistry to ascertain the distribution of D-DDB1 in the eye imaginal disc. It was found to be abundant in the anterior of the morphogenetic furrow (MF). Whole-body overexpression of dsRNA of D-DDB1 in Drosophila using a GAL4-UAS targeted expression system induced melanotic tumors and caused complete lethality. When limited to the eye imaginal disc, a severe rough eye phenotype resulted. Correspondingly, all of the D-DDB1 gene knocked-out flies also died. D-DDB1 therefore appears to be an essential development-associated factor in a multicellular organism.
The HKA Test Revisited
Authors: Wright Stephen I , Charlesworth Brian
Published: October 2004

We present a maximum-likelihood-ratio test of the standard neutral model, using multilocus data on polymorphism within species and divergence between species. The model is based on the Hudson-Kreitman-Aguadé (HKA) test, but allows for an explicit test of selection at individual loci in a multilocus framework. We use coalescent simulations to show that the likelihood-ratio test statistic is conservative, particularly when the assumption of no recombination is violated. Application of the method to polymorphism data from 18 loci from a population of Arabidopsis lyrata provides significant evidence for a balanced polymorphism at a candidate locus thought to be linked to the centromere. The method is also applied to polymorphism data in maize, providing support for the hypothesis of directional selection on genes in the starch pathway.
Connection of Propionyl-CoA Metabolism to Polyketide Biosynthesis in Aspergillus nidulans
Authors: Zhang Yong-Qiang , Brock Matthias , Keller Nancy P
Published: October 2004

Propionyl-CoA is an intermediate metabolite produced through a variety of pathways including thioesterification of propionate and catabolism of odd chain fatty acids and select amino acids. Previously, we found that disruption of the methylcitrate synthase gene, mcsA, which blocks propionyl-CoA utilization, as well as growth on propionate impaired production of several polyketides—molecules typically derived from acetyl-CoA and malonyl-CoA—including sterigmatocystin (ST), a potent carcinogen, and the conidiospore pigment. Here we describe three lines of evidence that demonstrate that excessive propionyl-CoA levels in the cell can inhibit polyketide synthesis. First, inactivation of a putative propionyl-CoA synthase, PcsA, which converts propionate to propionyl-CoA, restored polyketide production and reduced cellular propionyl-CoA content in a ΔmcsA background. Second, inactivation of the acetyl-CoA synthase, FacA, which is also involved in propionate utilization, restored polyketide production in the ΔmcsA background. Third, fungal growth on several compounds (e.g., heptadecanoic acid, isoleucine, and methionine) whose catabolism includes the formation of propionyl-CoA, were found to inhibit ST and conidiospore pigment production. These results demonstrate that excessive propionyl-CoA levels in the cell can inhibit polyketide synthesis.
Analysis of the Estimators of the Average Coefficient of Dominance of Deleterious Mutations
Authors: Fernández B , García-Dorado A , Caballero A
Published: October 2004

We investigate the sources of bias that affect the most commonly used methods of estimation of the average degree of dominance (h) of deleterious mutations, focusing on estimates from segregating populations. The main emphasis is on the effect of the finite size of the populations, but other sources of bias are also considered. Using diffusion approximations to the distribution of gene frequencies in finite populations as well as stochastic simulations, we assess the behavior of the estimators obtained from populations at mutation-selection-drift balance under different mutational scenarios and compare averages of h for newly arisen and segregating mutations. Because of genetic drift, the inferences concerning newly arisen mutations based on the mutation-selection balance theory can have substantial upward bias depending upon the distribution of h. In addition, estimates usually refer to h weighted by the homozygous deleterious effect in different ways, so that inferences are complicated when these two variables are negatively correlated. Due to both sources of bias, the widely used regression of heterozygous on homozygous means underestimates the arithmetic mean of h for segregating mutations, in contrast to their repeatedly assumed equality in the literature. We conclude that none of the estimators from segregating populations provides, under general conditions, a useful tool to ascertain the properties of the degree of dominance, either for segregating or for newly arisen deleterious mutations. Direct estimates of the average h from mutation-accumulation experiments are shown to suffer some bias caused by purging selection but, because they do not require assumptions on the causes maintaining segregating variation, they appear to give a more reliable average dominance for newly arisen mutations.
A Functional Analysis Reveals Dependence on the Anaphase-Promoting Complex for Prolonged Life Span in Yeast
Authors: Harkness Troy A A , Shea Kyla A , Legrand Charmaine , Brahmania Mayur , Davies Gerald F
Published: October 2004

Defects in anaphase-promoting complex (APC) activity, which regulates mitotic progression and chromatin assembly, results in genomic instability, a hallmark of premature aging and cancer. We investigated whether APC-dependent genomic stability affects aging and life span in yeast. Utilizing replicative and chronological aging assays, the APC was shown to promote longevity. Multicopy expression of genes encoding Snf1p (MIG1) and PKA (PDE2) aging-pathway components suppressed apc5CA phenotypes, suggesting their involvement in APC-dependent longevity. While it is known that PKA inhibits APC activity and reduces life span, a link between the Snf1p-inhibited Mig1p transcriptional modulator and the APC is novel. Our mutant analysis supports a model in which Snf1p promotes extended life span by inhibiting the negative influence of Mig1p on the APC. Consistent with this, we found that increased MIG1 expression reduced replicative life span, whereas mig1Δ mutations suppressed the apc5CA chronological aging defect. Furthermore, Mig1p and Mig2p activate APC gene transcription, particularly on glycerol, and mig2Δ, but not mig1Δ, confers a prolonged replicative life span in both APC5 and acp5CA cells. However, glucose repression of APC genes was Mig1p and Mig2p independent, indicating the presence of an uncharacterized factor. Therefore, we propose that APC-dependent genomic stability is linked to prolonged longevity by the antagonistic regulation of the PKA and Snf1p pathways.
A Novel Solution for the Time-Dependent Probability of Gene Fixation or Loss Under Natural Selection
Authors: Wang Ying , Rannala Bruce
Published: October 2004

Kimura (1955b) proposed a solution for the time-dependent probability of fixation, or loss, of a gene under selection. Example calculations suggest the formulas are prone to numerical inaccuracies. An alternative solution is proposed; the correctness of the solution is confirmed by numerically solving the Kolmogorov backward equation and by simulation.
Fixation Probability Favors Increased Fecundity Over Reduced Generation Time
Authors: Wahl L M , DeHaan C S
Published: October 2004

The cornerstone of population genetics is a probabilistic understanding of the ultimate fate—survival or extinction—of rare mutations. If a mutation is beneficial, it enables its carrier to reproduce faster than native wild-type individuals. In classic derivations and in the considerable body of research that has followed, “faster” has been defined mathematically to mean “able to produce more surviving offspring per generation.” Many organisms, however, may increase their reproductive rate by producing the same number of offspring in a shorter generation time: a mutant bacterium, for example, may complete the cell cycle and produce two offspring more quickly than the wild type. We find that the ultimate fixation probability of a mutation conferring a shorter generation time differs from that of a mutation conferring more offspring by a factor of 2 ln(2)—nearly 40%. This predicts a reduction in the overall substitution rate for any mutation that decreases the generation time: fixation probability is biased toward increased offspring number.
Genes Encoding Vitamin-K Epoxide Reductase Are Present in Drosophila and Trypanosomatid Protists
Authors: Robertson Hugh M
Published: October 2004

Vitamin-K epoxide reductase is encoded by the VKORC1 gene in mammals and other vertebrates, which also have a paralog, VKORC1L1. Single homologs are present in basal deuterostome and insect genomes, including Drosophila, and three trypanosomatid protists. VKOR is therefore an ancient gene/protein that can be studied in the Drosophila model system.
Mutation Rate and Predicted Phenotypic Target Sizes in Ethylnitrosourea-Treated Mice
Authors: Concepcion Dorothy , Seburn Kevin L , Wen Gen , Frankel Wayne N , Hamilton Bruce A
Published: October 2004

Chemical mutagenesis of the mouse is ongoing in several centers around the world, with varying estimates of mutation rate and number of sites mutable to phenotype. To address these questions, we sequenced ∼9.6 Mb of DNA from G1 progeny of ethylnitrosourea-treated mice in a large, broad-spectrum screen. We identified 10 mutations at eight unique sites, including six nonsynonymous coding substitutions. This calibrates the nucleotide mutation rate for two mutagenesis centers, implies significance criteria for positional cloning efforts, and provides working estimates of effective genetic target sizes for selected phenotypes.
CREB Binding Protein Functions During Successive Stages of Eye Development in Drosophila
Authors: Kumar Justin P , Jamal Tazeen , Doetsch Alex , Turner F Rudolf , Duffy Joseph B
Published: October 2004

During the development of the compound eye of Drosophila several signaling pathways exert both positive and inhibitory influences upon an array of nuclear transcription factors to produce a near-perfect lattice of unit eyes or ommatidia. Individual cells within the eye are exposed to many extracellular signals, express multiple surface receptors, and make use of a large complement of cell-subtype-specific DNA-binding transcription factors. Despite this enormous complexity, each cell will make the correct developmental choice and adopt the appropriate cell fate. How this process is managed remains a poorly understood paradigm. Members of the CREB binding protein (CBP)/p300 family have been shown to influence development by (1) acting as bridging molecules between the basal transcriptional machinery and specific DNA-binding transcription factors, (2) physically interacting with terminal members of signaling cascades, (3) acting as transcriptional coactivators of downstream target genes, and (4) playing a key role in chromatin remodeling. In a screen for new genes involved in eye development we have identified the Drosophila homolog of CBP as a key player in both eye specification and cell fate determination. We have used a variety of approaches to define the role of CBP in eye development on a cell-by-cell basis.
Interval Mapping of Quantitative Trait Loci for Resistance to Late Blight [Phytophthora infestans (Mont.) de Bary], Height and Maturity in a Tetraploid Population of Potato (Solanum tuberosum subsp. tuberosum)
Authors: Bradshaw John E , Pande Barnaly , Bryan Glenn J , Hackett Christine A , McLean Karen , Stewart Helen E , Waugh Robbie
Published: October 2004

Interval mapping of quantitative trait loci (QTL) for resistance to late blight, height, and maturity was performed on a tetraploid full-sib family of potato comprising 227 clones from a cross between a susceptible parent, 12601ab1, and a resistant cultivar, Stirling, which were of similar height and main crop maturity. Thirty-eight AFLP primer combinations provided 585 informative markers, and 23 SSRs proved useful for identifying linkage groups (LGs). A simplex QTL allele was found on LGV of Stirling close to marker STM3179, which was associated with early maturity, short plants, and susceptibility to blight and explained 54.7, 26.5, 26.3, and 17.5% of the variation for maturity, height, tuber blight, and foliage blight. When the residuals from the regressions of foliage and tuber blight on maturity were analyzed, there was no significant effect of a QTL on LGV, but a duplex QTL allele for resistance was found on LGIV of Stirling, which explained 30.7 and 13.6% of the variation for foliage and tuber blight on an additive model. Partial dominance for resistance explained even more of the variation, up to 37.2% for foliage blight. A major gene for blight resistance in Stirling was also mapped to LGXI.
Mutations in the Drosophila Condensin Subunit dCAP-G
Authors: Dej Kimberley J , Ahn Caroline , Orr-Weaver Terry L
Published: October 2004

Chromosomes are dynamic structures that are reorganized during the cell cycle to optimize them for distinct functions. SMC and non-SMC condensin proteins associate into complexes that have been implicated in the process of chromosome condensation. The roles of the individual non-SMC subunits of the complex are poorly understood, and mutations in the CAP-G subunit have not been described in metazoans. Here we elucidate a role for dCAP-G in chromosome condensation and cohesion in Drosophila. We illustrate the requirement of dCAP-G for condensation during prophase and prometaphase; however, we find that alternate mechanisms ensure that replicated chromosomes are condensed prior to metaphase. In contrast, dCAP-G is essential for chromosome condensation in metaphase of single, unreplicated sister chromatids, suggesting that there is an interplay between replicated chromatids and the condensin complex. In the dcap-g mutants, defects in sister-chromatid separation are also observed. Chromatid arms fail to resolve in prophase and are unable to separate at anaphase, whereas sister centromeres show aberrant separation in metaphase and successfully move to spindle poles at anaphase. We also identified a role for dCAP-G during interphase in regulating heterochromatic gene expression.
Construction and Evaluation of cDNA Libraries for Large-Scale Expressed Sequence Tag Sequencing in Wheat (Triticum aestivum L.)
Authors: Zhang D , Choi D W , Wanamaker S , Fenton R D , Chin A , Malatrasi M , Turuspekov Y , Walia H , Akhunov E D , Kianian P , Otto C , Simons K , Deal K R , Echenique V , Stamova B , Ross K , Butler G E , Strader L , Verhey S D , Johnson R , Altenbach S , Kothari K , Tanaka C , Shah M M , Laudencia-Chingcuanco D , Han P , Miller R E , Crossman C C , Chao S , Lazo G R , Klueva N , Gustafson J P , Kianian S F , Dubcovsky J , Walker-Simmons M K , Gill K S , Dvořák J , Anderson O D , Sorrells M E , McGuire P E , Qualset C O , Nguyen H T , Close T J
Published: October 2004

A total of 37 original cDNA libraries and 9 derivative libraries enriched for rare sequences were produced from Chinese Spring wheat (Triticum aestivum L.), five other hexaploid wheat genotypes (Cheyenne, Brevor, TAM W101, BH1146, Butte 86), tetraploid durum wheat (T. turgidum L.), diploid wheat (T. monococcum L.), and two other diploid members of the grass tribe Triticeae (Aegilops speltoides Tausch and Secale cereale L.). The emphasis in the choice of plant materials for library construction was reproductive development subjected to environmental factors that ultimately affect grain quality and yield, but roots and other tissues were also included. Partial cDNA expressed sequence tags (ESTs) were examined by various measures to assess the quality of these libraries. All ESTs were processed to remove cloning system sequences and contaminants and then assembled using CAP3. Following these processing steps, this assembly yielded 101,107 sequences derived from 89,043 clones, which defined 16,740 contigs and 33,213 singletons, a total of 49,953 “unigenes.” Analysis of the distribution of these unigenes among the libraries led to the conclusion that the enrichment methods were effective in reducing the most abundant unigenes and to the observation that the most diverse libraries were from tissues exposed to environmental stresses including heat, drought, salinity, or low temperature.
Chromosome Bin Map of Expressed Sequence Tags in Homoeologous Group 1 of Hexaploid Wheat and Homoeology With Rice and Arabidopsis
Authors: Peng J H , Zadeh H , Lazo G R , Gustafson J P , Chao S , Anderson O D , Qi L L , Echalier B , Gill B S , Dilbirligi M , Sandhu D , Gill K S , Greene R A , Sorrells M E , Akhunov E D , Dvořák J , Linkiewicz A M , Dubcovsky J , Hossain K G , Kalavacharla V , Kianian S F , Mahmoud A A ,   Miftahudin, Conley E J , Anderson J A , Pathan M S , Nguyen H T , McGuire P E , Qualset C O , Lapitan N L V
Published: October 2004

A total of 944 expressed sequence tags (ESTs) generated 2212 EST loci mapped to homoeologous group 1 chromosomes in hexaploid wheat (Triticum aestivum L.). EST deletion maps and the consensus map of group 1 chromosomes were constructed to show EST distribution. EST loci were unevenly distributed among chromosomes 1A, 1B, and 1D with 660, 826, and 726, respectively. The number of EST loci was greater on the long arms than on the short arms for all three chromosomes. The distribution of ESTs along chromosome arms was nonrandom with EST clusters occurring in the distal regions of short arms and middle regions of long arms. Duplications of group 1 ESTs in other homoeologous groups occurred at a rate of 35.5%. Seventy-five percent of wheat chromosome 1 ESTs had significant matches with rice sequences (E ≤ e−10), where large regions of conservation occurred between wheat consensus chromosome 1 and rice chromosome 5 and between the proximal portion of the long arm of wheat consensus chromosome 1 and rice chromosome 10. Only 9.5% of group 1 ESTs showed significant matches to Arabidopsis genome sequences. The results presented are useful for gene mapping and evolutionary and comparative genomics of grasses.