bluuebunny/crossref_metadata_2025 huggingface dataset:
From Lost Property to Explorer' s Relics: The Rediscovery of the Personal Library of Ludwig Leichhardt
Authors: Stephens Matthew
Published: November 2007

In late 1853, a small number of unclaimed boxes containing the worldly possessions of the missing explorer Ludwig Leichhardt were deposited at the Australian Museum, Sydney. An estimated 137 volumes of Leichhardt's books and pamphlets were stored alongside his manuscripts, field notes, seed specimens and scientific instruments. While the manuscripts have proved invaluable to those researching the life and work of Leichhardt, his books have lain forgotten and virtually irretrievable in the collections of the State Library of New South Wales and the Australian Museum Research Library. A significant proportion of the library has now been identified and its contents listed and described for the first time, providing new insight into Leichhard's intellectual background and interests.
Not Being Ernest: Uncovering Competitors in the Foundation of Australian Wireless
Authors: Given Jock
Published: November 2007

Ernest Fisk was the dominant figure in early wireless in Australia. He headed Amalgamated Wireless (Australasia), AWA, 1917–44, and was managing director of Electric and Musical Industries (EMI) in London, 1945–51. Arriving in Australia in 1911 at a critical moment in wireless development, Fisk became the main local representative for an industry that was born global. He was not, however, the first, as is often claimed. This article examines his predecessors, whose failures tell a good deal about Fisk's strengths, the good fortune of his timing and the business strategies of early multinational wireless companies.
The micro-Petri dish, a million-well growth chip for the culture and high-throughput screening of microorganisms
Authors: Ingham Colin J. , Sprenkels Ad , Bomer Johan , Molenaar Douwe , van den Berg Albert , van Hylckama Vlieg Johan E. T. , de Vos Willem M.
Published: November 2007

A miniaturized, disposable microbial culture chip has been fabricated by microengineering a highly porous ceramic sheet with up to one million growth compartments. This versatile culture format, with discrete compartments as small as 7 × 7 μm, allowed the growth of segregated microbial samples at an unprecedented density. The chip has been used for four complementary applications in microbiology. (i) As a fast viable counting system that showed a dynamic range of over 10,000, a low degree of bias, and a high culturing efficiency. (ii) In high-throughput screening, with the recovery of 1 fluorescent microcolony in 10,000. (iii) In screening for an enzyme-based, nondominant phenotype by the targeted recovery ofEscherichia colitransformed with the plasmid pUC18, based on expression of thelacZreporter gene without antibiotic-resistance selection. The ease of rapid, successive changes in the environment of the organisms on the chip, needed for detection of β-galactosidase activity, highlights an advantageous feature that was also used to screen a metagenomic library for the same activity. (iv) In high-throughput screening of >200,000 isolates from Rhine water based on metabolism of a fluorogenic organophosphate compound, resulting in the recovery of 22 microcolonies with the desired phenotype. These isolates were predicted, on the basis of rRNA sequence, to include six new species. These four applications suggest that the potential for such simple, readily manufactured chips to impact microbial culture is extensive and may facilitate the full automation and multiplexing of microbial culturing, screening, counting, and selection.
Epicardium-derived progenitor cells require β-catenin for coronary artery formation
Authors: Zamora Mónica , Männer Jörg , Ruiz-Lozano Pilar
Published: November 2007

We have previously identified several members of the Wnt/β-catenin pathway that are differentially expressed in a mouse model with deficient coronary vessel formation. Systemic ablation of β-catenin expression affects mouse development at gastrulation with failure of both mesoderm development and axis formation. To circumvent this early embryonic lethality and study the specific role of β-catenin in coronary arteriogenesis, we have generated conditional β-catenin-deletion mutant animals in the proepicardium by interbreeding with a Cre-expressing mouse that targets coronary progenitor cells in the proepicardium and its derivatives. Ablation of β-catenin in the proepicardium results in lethality between embryonic day 15 and birth. Mutant mice display impaired coronary artery formation, whereas the venous system and microvasculature are normal. Analysis of proepicardial β-catenin mutant cells in the context of an epicardial tracer mouse reveals that the formation of the proepicardium, the migration of proepicardial cells to the heart, and the formation of the primitive epicardium are unaffected. However, subsequent processes of epicardial development are dramatically impaired in epicardial-β-catenin mutant mice, including failed expansion of the subepicardial space, blunted invasion of the myocardium, and impaired differentiation of epicardium-derived mesenchymal cells into coronary smooth muscle cells. Our data demonstrate a functional role of the epicardial β-catenin pathway in coronary arteriogenesis.
Dimerization of the thyrotropin-releasing hormone receptor potentiates hormone-dependent receptor phosphorylation
Authors: Song Gyun Jee , Jones Brian W. , Hinkle Patricia M.
Published: November 2007

The G protein-coupled thyrotropin (TSH)-releasing hormone (TRH) receptor forms homodimers. Regulated receptor dimerization increases TRH-induced receptor endocytosis. These studies test whether dimerization increases receptor phosphorylation, which could potentiate internalization. Phosphorylation at residues 355–365, which is critical for internalization, was measured with a highly selective phospho-site-specific antibody. Two strategies were used to drive receptor dimerization. Dimerization of a TRH receptor-FK506-binding protein (FKBP) fusion protein was stimulated by a dimeric FKBP ligand. The chemical dimerizer caused a large increase in TRH-dependent phosphorylation within 1 min, whereas a monomeric FKBP ligand had no effect. The dimerizer did not alter phoshorylation of receptors lacking the FKBP domain. Dimerization of receptors containing an N-terminal HA epitope also was induced with anti-HA antibody. Anti-HA IgG strongly increased TRH-induced phosphorylation, whereas monomeric Fab fragments had no effect. Anti-HA antibody did not alter phosphorylation in receptors lacking an HA tag. Furthermore, two phosphorylation-defective TRH receptors functionally complemented one another and permitted phosphorylation. Receptors with a D71A mutation in the second transmembrane domain do not signal, whereas receptors with four Ala mutations in the 355–365 region signal normally but lack phosphorylation sites. When D71A- and 4Ala-TRH receptors were expressed alone, neither underwent TRH-dependent phosphorylation. When they were expressed together, D71A receptor was phosphorylated by G protein-coupled receptor kinases in response to TRH. These results suggest that the TRH receptor is phosphorylated preferentially when it is in dimers or when preexisting receptor dimers are driven into microaggregates. Increased receptor phosphorylation may amplify desensitization.
Temporal isolation of neural processes underlying face preference decisions
Authors: Kim Hackjin , Adolphs Ralph , O'Doherty John P. , Shimojo Shinsuke
Published: November 2007

Decisions about whether we like someone are often made so rapidly from first impressions that it is difficult to examine the engagement of neural structures at specific points in time. Here, we used a temporally extended decision-making paradigm to examine brain activation with functional MRI (fMRI) at sequential stages of the decision-making process. Activity in reward-related brain structures—the nucleus accumbens (NAC) and orbitofrontal cortex (OFC)—was found to occur at temporally dissociable phases while subjects decided which of two unfamiliar faces they preferred. Increases in activation in the OFC occurred late in the trial, consistent with a role for this area in computing the decision of which face to choose. Signal increases in the NAC occurred early in the trial, consistent with a role for this area in initial preference formation. Moreover, early signal increases in the NAC also occurred while subjects performed a control task (judging face roundness) when these data were analyzed on the basis of which of those faces were subsequently chosen as preferred in a later task. The findings support a model in which rapid, automatic engagement of the NAC conveys a preference signal to the OFC, which in turn is used to guide choice.
The conformation of acetylcholine at its target site in the membrane-embedded nicotinic acetylcholine receptor
Authors: Williamson P. T. F. , Verhoeven A. , Miller K. W. , Meier B. H. , Watts A.
Published: November 2007

The conformation of the neurotransmitter acetylcholine bound to the fully functional nicotinic acetylcholine receptor embedded in its native membrane environment has been characterized by using frequency-selective recoupling solid-state NMR. Six dipolar couplings among five resolved
13
C-labeled atoms of acetylcholine were measured. Bound acetylcholine adopts a bent conformation characterized with a quaternary ammonium-to-carbonyl distance of 5.1 Å. In this conformation, and with its orientation constrained to that previously determined by us, the acetylcholine could be docked satisfactorily in the agonist pocket of the agonist-bound, but not the agonist-free, crystal structure of a soluble acetylcholine-binding protein from
Lymnaea stagnali
. The quaternary ammonium group of the acetylcholine was determined to be within 3.9 Å of five aromatic residues and its acetyl group close to residues C187/188 of the principle and residue L112 of the complementary subunit. The observed >C

O chemical shift is consistent with H bonding to the nicotinic acetylcholine receptor residues γY116 and δT119 that are homologous to L112 in the soluble acetylcholine-binding protein.
Distribution of node characteristics in complex networks
Authors: Park Juyong , Barabási Albert-László
Published: November 2007

Our enhanced ability to map the structure of various complex networks is increasingly accompanied by the possibility of independently identifying the functional characteristics of each node. Although this led to the observation that nodes with similar characteristics have a tendency to link to each other, in general we lack the tools to quantify the interplay between node properties and the structure of the underlying network. Here we show that when nodes in a network belong to two distinct classes, two independent parameters are needed to capture the detailed interplay between the network structure and node properties. We find that the network structure significantly limits the values of these parameters, requiring a phase diagram to uniquely characterize the configurations available to the system. The phase diagram shows a remarkable independence from the network size, a finding that, together with a proposed heuristic algorithm, allows us to determine its shape even for large networks. To test the usefulness of the developed methods, we apply them to biological and socioeconomic systems, finding that protein functions and mobile phone usage occupy distinct regions of the phase diagram, indicating that the proposed parameters have a strong discriminating power.
α-Catenin is essential in intestinal adenoma formation
Authors: Shibata Hiroyuki , Takano Hiroshi , Ito Masaki , Shioya Hisashi , Hirota Morihisa , Matsumoto Hiroshi , Kakudo Yuichi , Ishioka Chikashi , Akiyama Tetsu , Kanegae Yumi , Saito Izumu , Noda Tetsuo
Published: November 2007

A loss-of-function mutation in the
APC
gene initiates colorectal carcinogenesis. Although the molecular mechanism of tumor initiation is complex, several modifier genes have been identified using mouse models, including the
ApcMin
mouse. Among the familial adenomatous polyposis mouse lines carrying a truncation mutation at codon 580 in
Apc
(
Apc580D
), one line (line19-
Apc

580D
/+

) showed a remarkably reduced incidence of intestinal adenomas (<5% compared with other lines). Extensive genetic analysis identified a deletion in the α-
catenin
(
Ctnna1
) gene as the cause of this suppression. Notably, the suppression only occurred when the
Ctnna1
deletion was in
cis
-configuration with the
Apc580D
mutation. In all adenomas generated in line19-
Apc

580D
/+

, somatic recombination between the
Apc
and
Ctnna1
loci retained the wild-type
Ctnna1
allele. These data strongly indicate that simultaneous inactivation of α-
catenin
and
Apc
during tumor initiation suppresses adenoma formation in line19-
Apc

580D
/+

, suggesting that α-
catenin
plays an essential role in the initiation of intestinal adenomas. Although accumulating evidence obtained from human colon tumors with invasive or metastatic potential has established a tumor-suppressive role for α-catenin in late-stage tumorigenesis, the role of α-catenin in the initiation of intestinal tumorigenesis is not well documented, especially compared with that of β-catenin. A mouse model used in this study focused on the early stage of tumor initiation and clearly indicated an essential role for α-catenin. Thus, α-catenin has dual roles in intestinal tumorigenesis, a supporting role in tumor initiation, and a suppressive role in tumor progression.
Temporal and spatial activation of caspase-like enzymes induced by self-incompatibility in
Papaver
pollen
Authors: Bosch Maurice , Franklin-Tong Vernonica E.
Published: November 2007

Caspase-like proteases are universal mediators of programmed cell death (PCD). Because plants have no caspase homologs, establishing the nature of their caspase-like activities is of considerable importance to our understanding of PCD in plants. Caspase-3, displaying DEVD specificity, is a key executioner caspase in animal cells. Self-incompatibility (SI) is an important mechanism to prevent self-fertilization and inbreeding in higher plants by inhibiting incompatible pollen. In
Papaver rhoeas
, SI activates a caspase-3-like/DEVDase activity in incompatible pollen that plays a pivotal role in regulating PCD. Here we characterize the SI-induced caspase-like activities in detail; our work provides insights into the temporal and spatial activation of plant caspase-like enzymes. We show that SI also activates a VEIDase and a LEVDase and that the VEIDase plays a role in SI-induced PCD. The DEVDase and VEIDase are activated remarkably rapidly: detectable within 1–2 h after SI induction; the LEVDase activity peaks later. Importantly, we show live-cell imaging of a DEVDase activity in a higher plant cell; the SI-activated DEVDase has a cytosolic and nuclear localization. We also demonstrate that SI induces a rapid and substantial cytosolic acidification that matches the
in vitro
pH optima for the SI-induced caspase activities. Because both cytosolic acidification and nuclear caspase localization are observed during apoptosis in animal cells, our data provide striking parallels between SI-induced PCD and apoptosis in animal cells.
Tyrosine-sulfated glycopeptide involved in cellular proliferation and expansion in
Arabidopsis
Authors: Amano Yukari , Tsubouchi Hiroko , Shinohara Hidefumi , Ogawa Mari , Matsubayashi Yoshikatsu
Published: November 2007

Posttranslational modification can confer special functions to peptides. Based on exhaustive liquid chromatography mass spectrometry analysis targeting tyrosine-sulfated peptides, we identified an 18-aa tyrosine-sulfated glycopeptide in
Arabidopsis
cell suspension culture medium. This peptide, which we named PSY1, significantly promotes cellular proliferation and expansion at nanomolar concentrations.
PSY1
is widely expressed in various
Arabidopsis
tissues, including shoot apical meristem, and is highly up-regulated by wounding. Perception of PSY1 depends on At1g72300, which is a leucine-rich repeat receptor kinase (LRR-RK) whose two paralogs are involved in the perception of phytosulfokine (PSK), which is a 5-aa tyrosine-sulfated peptide that primarily promotes cellular proliferation. Multiple loss-of-function mutations in these three paralogous LRR-RKs significantly enhanced phenotypes, compared with single disruptants, suggesting that these LRR-RKs have overlapping functions. Triple mutations in these LRR-RKs resulted in dwarfism because of decreases in cell number and cell size and caused insufficiency in tissue repair after wounding. The present results suggest that this paralogous LRR-RK family integrates growth-promoting signals mediated by two structurally distinct sulfated peptides: PSY1 and PSK.
Functional screening identifies miR-315 as a potent activator of Wingless signaling
Authors: Silver Serena J. , Hagen Joshua W. , Okamura Katsutomo , Perrimon Norbert , Lai Eric C.
Published: November 2007

The existence of vast regulatory networks mediated by microRNAs (miRNAs) suggests broad potential for miRNA dysfunction to contribute to disease. However, relatively few miRNA–target interactions are likely to make detectable contributions to phenotype, and effective strategies to identify these few interactions are currently wanting. We hypothesized that signaling cascades represent critical points of susceptibility to miRNA dysfunction, and we developed a strategy to test this theory by using quantitative cell-based screens. Here we report a screen for miRNAs that affect the Wingless (Wg) pathway, a conserved pathway that regulates growth and tissue specification. This process identified ectopic miR-315 as a potent and specific activator of Wg signaling, an activity that we corroborated in transgenic animals. This miR-315 activity was mediated by direct inhibition of
Axin
and
Notum
, which encode essential, negatively acting components of the Wg pathway. Genetic interaction tests substantiated both of these genes as key functional targets of miR-315. The ability of ectopic miR-315 to activate Wg signaling was not a trivial consequence of predicted miRNA–target relationships because other miRNAs with conserved sites in the
Axin
3′ UTR neither activated Wg outputs nor inhibited an
Axin
sensor. In summary, activity-based screening can selectively identify miRNAs whose deregulation can lead to interpretable phenotypic consequences.
Mutational activation of niche-specific genes provides insight into regulatory networks and bacterial function in a complex environment
Authors: Giddens Stephen R. , Jackson Robert W. , Moon Christina D. , Jacobs Michael A. , Zhang Xue-Xian , Gehrig Stefanie M. , Rainey Paul B.
Published: November 2007

The genome of the plant-colonizing bacterium
Pseudomonas fluorescens
SBW25 harbors a subset of genes that are expressed specifically on plant surfaces. The function of these genes is central to the ecological success of SBW25, but their study poses significant challenges because no phenotype is discernable
in vitro
. Here, we describe a genetic strategy with general utility that combines suppressor analysis with IVET (SPyVET) and provides a means of identifying regulators of niche-specific genes. Central to this strategy are strains carrying operon fusions between plant environment-induced loci (EIL) and promoterless ′
dapB
. These strains are prototrophic in the plant environment but auxotrophic on laboratory minimal medium. Regulatory elements were identified by transposon mutagenesis and selection for prototrophs on minimal medium. Approximately 10
6
mutants were screened for each of 27 strains carrying ′
dapB
fusions to plant EIL and the insertion point for the transposon determined in approximately 2,000 putative regulator mutants. Regulators were functionally characterized and used to provide insight into EIL phenotypes. For one strain carrying a fusion to the cellulose-encoding
wss
operon, five different regulators were identified including a diguanylate cyclase, the flagella activator, FleQ, and alginate activator, AmrZ (AlgZ). Further rounds of suppressor analysis, possible by virtue of the SPyVET strategy, revealed an additional two regulators including the activator AlgR, and allowed the regulatory connections to be determined.
Melanocortin 2 receptor is required for adrenal gland development, steroidogenesis, and neonatal gluconeogenesis
Authors: Chida Dai , Nakagawa Shinichi , Nagai So , Sagara Hiroshi , Katsumata Harumi , Imaki Toshihiro , Suzuki Harumi , Mitani Fumiko , Ogishima Tadashi , Shimizu Chikara , Kotaki Hayato , Kakuta Shigeru , Sudo Katsuko , Koike Takao , Kubo Mitsumasa , Iwakura Yoichiro
Published: November 2007

ACTH (i.e., corticotropin) is the principal regulator of the hypothalamus–pituitary–adrenal axis and stimulates steroidogenesis in the adrenal gland via the specific cell-surface melanocortin 2 receptor (MC2R). Here, we generated mice with an inactivation mutation of the MC2R gene to elucidate the roles of MC2R in adrenal development, steroidogenesis, and carbohydrate metabolism. These mice, the last of the knockout (KO) mice to be generated for melanocortin family receptors, provide the opportunity to compare the phenotype of proopiomelanocortin KO mice with that of MC1R–MC5R KO mice. We found that the MC2R KO mutation led to neonatal lethality in three-quarters of the mice, possibly as a result of hypoglycemia. Those surviving to adulthood exhibited macroscopically detectable adrenal glands with markedly atrophied zona fasciculata, whereas the zona glomerulosa and the medulla remained fairly intact. Mutations of MC2R have been reported to be responsible for 25% of familial glucocorticoid deficiency (FGD) cases. Adult MC2R KO mice resembled FGD patients in several aspects, such as undetectable levels of corticosterone despite high levels of ACTH, unresponsiveness to ACTH, and hypoglycemia after prolonged (36 h) fasting. However, MC2R KO mice differ from patients with MC2R-null mutations in several aspects, such as low aldosterone levels and unaltered body length. These results indicate that MC2R is required for postnatal adrenal development and adrenal steroidogenesis and that MC2R KO mice provide a useful animal model by which to study FGD.
Escherichia coli
succinate dehydrogenase variant lacking the heme
b
Authors: Tran Quang M. , Rothery Richard A. , Maklashina Elena , Cecchini Gary , Weiner Joel H.
Published: November 2007

The
Escherichia coli
enzyme succinate:ubiquinone oxidoreductase [(succinate dehydrogenase (SdhCDAB)] couples succinate oxidation to ubiquinone reduction and is structurally and functionally equivalent to mitochondrial complex II, an essential component of the aerobic respiratory chain and tricarboxylic acid cycle. All such enzymes contain a heme within their membrane anchor domain with a highly contentious, but as-yet-undetermined, function. Here, we report the generation of a complex II that lacks heme, which is confirmed by both optical and EPR spectroscopy. Despite the absence of heme, this mutant still assembles properly and retains physiological activity. However, the mutants lacking heme are highly sensitive to the presence of detergent. In addition, the heme does not appear to be involved in reactive oxygen species suppression. Our results indicate that redox cycling of the heme in complex II is not essential for the enzyme's ubiquinol reductase activity.
Cell division modulates prion accumulation in cultured cells
Authors: Ghaemmaghami Sina , Phuan Puay-Wah , Perkins Beth , Ullman Julie , May Barnaby C. H. , Cohen Fred E. , Prusiner Stanley B.
Published: November 2007

The phenotypic effect of prions on host cells is influenced by the physical properties of the prion strain and its level of accumulation. In mammalian cell cultures, prion accumulation is determined by the interplay between
de novo
prion formation, catabolism, cell division, and horizontal cell-to-cell transmission. Understanding this dynamic enables the analytical modeling of protein-based heritability and infectivity. Here, we quantitatively measured these competing effects in a subline of neuroblastoma (N2a) cells and propose a concordant reaction mechanism to explain the kinetics of prion propagation. Our results show that cell division leads to a predictable reduction in steady-state prion levels but not to complete clearance. Scrapie-infected N2a cells were capable of accumulating different steady-state levels of prions, dictated partly by the rate of cell division. We also show that prions in this subline of N2a cells are transmitted primarily from mother to daughter cells, rather than horizontal cell-to-cell transmission. We quantitatively modeled our kinetic results based on a mechanism that assumes a subpopulation of prions is capable of self-catalysis, and the levels of this subpopulation reach saturation in fully infected cells. Our results suggest that the apparent effectiveness of antiprion compounds in culture may be strongly influenced by the growth phase of the target cells.
Illuminating the mechanistic roles of enzyme conformational dynamics
Authors: Hanson Jeffrey A. , Duderstadt Karl , Watkins Lucas P. , Bhattacharyya Sucharita , Brokaw Jason , Chu Jhih-Wei , Yang Haw
Published: November 2007

Many enzymes mold their structures to enclose substrates in their active sites such that conformational remodeling may be required during each catalytic cycle. In adenylate kinase (AK), this involves a large-amplitude rearrangement of the enzyme's lid domain. Using our method of high-resolution single-molecule FRET, we directly followed AK's domain movements on its catalytic time scale. To quantitatively measure the enzyme's entire conformational distribution, we have applied maximum entropy-based methods to remove photon-counting noise from single-molecule data. This analysis shows unambiguously that AK is capable of dynamically sampling two distinct states, which correlate well with those observed by x-ray crystallography. Unexpectedly, the equilibrium favors the closed, active-site-forming configurations even in the absence of substrates. Our experiments further showed that interaction with substrates, rather than locking the enzyme into a compact state, restricts the spatial extent of conformational fluctuations and shifts the enzyme's conformational equilibrium toward the closed form by increasing the closing rate of the lid. Integrating these microscopic dynamics into macroscopic kinetics allows us to model lid opening-coupled product release as the enzyme's rate-limiting step.
Processing of tactile information by the hippocampus
Authors: Pereira Antonio , Ribeiro Sidarta , Wiest Michael , Moore Leonardo C. , Pantoja Janaina , Lin Shih-Chieh , Nicolelis Miguel A. L.
Published: November 2007

The ability to detect unusual events occurring in the environment is essential for survival. Several studies have pointed to the hippocampus as a key brain structure in novelty detection, a claim substantiated by its wide access to sensory information through the entorhinal cortex and also distinct aspects of its intrinsic circuitry. Novelty detection is implemented by an associative match–mismatch algorithm involving the CA1 and CA3 hippocampal subfields that compares the stream of sensory inputs received by CA1 to the stored representation of spatiotemporal sequences in CA3. In some rodents, including the rat, the highly sensitive facial whiskers are responsible for providing accurate tactile information about nearby objects. Surprisingly, however, not much is known about how inputs from the whiskers reach CA1 and how they are processed therein. Using concurrent multielectrode neuronal recordings and chemical inactivation in behaving rats, we show that trigeminal inputs from the whiskers reach the CA1 region through thalamic and cortical relays associated with discriminative touch. Ensembles of hippocampal neurons also carry precise information about stimulus identity when recorded during performance in an aperture-discrimination task using the whiskers. We also found broad similarities between tactile responses of trigeminal stations and the hippocampus during different vigilance states (wake and sleep). Taken together, our results show that tactile information associated with fine whisker discrimination is readily available to the hippocampus for dynamic updating of spatial maps.
Mutations in ionotropic AMPA receptor 3 alter channel properties and are associated with moderate cognitive impairment in humans
Authors: Wu Ye , Arai Amy C. , Rumbaugh Gavin , Srivastava Anand K. , Turner Gillian , Hayashi Takashi , Suzuki Erika , Jiang Yuwu , Zhang Lilei , Rodriguez Jayson , Boyle Jackie , Tarpey Patrick , Raymond F. Lucy , Nevelsteen Joke , Froyen Guy , Stratton Mike , Futreal Andy , Gecz Jozef , Stevenson Roger , Schwartz Charles E. , Valle David , Huganir Richard L. , Wang Tao
Published: November 2007

Ionotropic α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors (iGluRs) mediate the majority of excitatory synaptic transmission in the CNS and are essential for the induction and maintenance of long-term potentiation and long-term depression, two cellular models of learning and memory. We identified a genomic deletion (0.4 Mb) involving the entire
GRIA3
(encoding iGluR3) by using an X-array comparative genomic hybridization (CGH) and four missense variants (G833R, M706T, R631S, and R450Q) in functional domains of iGluR3 by sequencing 400 males with X-linked mental retardation (XLMR). Three variants were found in males with moderate MR and were absent in 500 control males. Expression studies in HEK293 cells showed that G833R resulted in a 78% reduction of iGluR3 due to protein misfolding. Whole-cell recording studies of iGluR3 homomers in HEK293 cells revealed that neither iGluR3-M706T (S2 domain) nor iGluR3-R631S (near channel core) had substantial channel function, whereas R450Q (S1 domain) was associated with accelerated receptor desensitization. When forming heteromeric receptors with iGluR2 in HEK293 cells, all four iGluR3 variants had altered desensitization kinetics. Our study provides the genetic and functional evidence that mutant iGluR3 with altered kinetic properties is associated with moderate cognitive impairment in humans.
Identification of yeast proteins necessary for cell-surface function of a potassium channel
Authors: Haass Friederike A. , Jonikas Martin , Walter Peter , Weissman Jonathan S. , Jan Yuh-Nung , Jan Lily Y. , Schuldiner Maya
Published: November 2007

Inwardly rectifying potassium (Kir) channels form gates in the cell membrane that regulate the flow of K
+
ions into and out of the cell, thereby influencing the membrane potential and electrical signaling of many cell types, including neurons and cardiomyocytes. Kir-channel function depends on other cellular proteins that aid in the folding of channel subunits, assembly into tetrameric complexes, trafficking of quality-controlled channels to the plasma membrane, and regulation of channel activity at the cell surface. We used the yeast
Saccharomyces cerevisiae
as a model system to identify proteins necessary for the functional expression of a mammalian Kir channel at the cell surface. A screen of 376 yeast strains, each lacking one nonessential protein localized to the early secretory pathway, identified seven deletion strains in which functional expression of the Kir channel at the plasma membrane was impaired. Six deletions were of genes with known functions in trafficking and lipid biosynthesis (
sur4
Δ,
csg2
Δ,
erv14
Δ,
emp24
Δ,
erv25
Δ, and
bst1
Δ), and one deletion was of an uncharacterized gene (
yil039w
Δ). We provide genetic and functional evidence that Yil039wp, a conserved, phosphoesterase domain-containing protein, which we named “trafficking of Emp24p/Erv25p-dependent cargo disrupted 1” (Ted1p), acts together with Emp24p/Erv25p in cargo exit from the endoplasmic reticulum (ER). The seven yeast proteins identified in our screen likely impact Kir-channel functional expression at the level of vesicle budding from the ER and/or the local lipid environment at the plasma membrane.
Activating mutation in a mucolipin transient receptor potential channel leads to melanocyte loss in varitint–waddler mice
Authors: Xu Haoxing , Delling Markus , Li Linyu , Dong Xianping , Clapham David E.
Published: November 2007

Transient receptor potential (TRP) genes of the mucolipin subfamily (TRPML1–3 and MCOLN1–3) are presumed to encode ion channel proteins of intracellular endosomes and lysosomes. Mutations in human
TRPML1
(mucolipin 1/MCOLN1) result in mucolipidosis type IV, a severe inherited neurodegenerative disease associated with defective lysosomal biogenesis and trafficking. A mutation in mouse TRPML3 (A419P; TRPML3

Va

) results in the varitint–waddler (
Va
) phenotype.
Va
mice are deaf, exhibit circling behavior due to vestibular defects, and have variegated/dilute coat color as a result of pigmentation defects. Prior electrophysiological studies of presumed TRPML plasma membrane channels are contradictory and inconsistent with known TRP channel properties. Here, we report that the
Va
mutation produces a gain-of-function that allows TRPML1 and TRPML3 to be measured and identified as inwardly rectifying, proton-impermeant, Ca
2+
-permeant cation channels. TRPML3 is highly expressed in normal melanocytes. Melanocyte markers are lost in the
Va
mouse, suggesting that their variegated and hypopigmented fur is caused by severe alteration of melanocyte function or cell death. TRPML3

Va

expression in melanocyte cell lines results in high resting Ca
2+
levels, rounded, poorly adherent cells, and loss of membrane integrity. We conclude that the
Va
phenotype is caused by mutation-induced TRPML3 gain-of-function, resulting in cell death.
RNA sequence analysis defines Dicer's role in mouse embryonic stem cells
Authors: Calabrese J. Mauro , Seila Amy C. , Yeo Gene W. , Sharp Phillip A.
Published: November 2007

Short RNA expression was analyzed from
Dicer
-positive and
Dicer
-knockout mouse embroyonic stem (ES) cells, using high-throughput pyrosequencing. A correlation of miRNA quantification with sequencing frequency estimates that there are 110,000 miRNAs per ES cell, the majority of which can be accounted for by six distinct miRNA loci. Four of these miRNA loci or their human homologues have demonstrated roles in cell cycle regulation or oncogenesis, suggesting that a major function of the miRNA pathway in ES cells may be to shape their distinct cell cycle. Forty-six previously uncharacterized miRNAs were identified, most of which are expressed at low levels and are less conserved than the set of known miRNAs. Low-abundance short RNAs matching all classes of repetitive elements were present in cells lacking
Dicer
, although the production of some SINE- and simple repeat-associated short RNAs appeared to be Dicer-dependent. These and other Dicer-dependent sequences resembled miRNAs. At a depth of sequencing that approaches the total number of 5′ phosphorylated short RNAs per cell, miRNAs appeared to be Dicer's only substrate. The results presented suggest a model in which repeat-associated miRNAs serve as host defenses against repetitive elements, a function canonically ascribed to other classes of short RNA.
Identification of an Axonal Kinesin-3 Motor for Fast Anterograde Vesicle Transport that Facilitates Retrograde Transport of Neuropeptides
Authors: Barkus Rosemarie V. , Klyachko Olga , Horiuchi Dai , Dickson Barry J. , Saxton William M.
Published: November 2007

A screen for genes required in Drosophila eye development identified an UNC-104/Kif1 related kinesin-3 microtubule motor. Analysis of mutants suggested that Drosophila Unc-104 has neuronal functions that are distinct from those of the classic anterograde axonal motor, kinesin-1. In particular, unc-104 mutations did not cause the distal paralysis and focal axonal swellings characteristic of kinesin-1 (Khc) mutations. However, like Khc mutations, unc-104 mutations caused motoneuron terminal atrophy. The distributions and transport behaviors of green fluorescent protein-tagged organelles in motor axons indicate that Unc-104 is a major contributor to the anterograde fast transport of neuropeptide-filled vesicles, that it also contributes to anterograde transport of synaptotagmin-bearing vesicles, and that it contributes little or nothing to anterograde transport of mitochondria, which are transported primarily by Khc. Remarkably, unc-104 mutations inhibited retrograde runs by neurosecretory vesicles but not by the other two organelles. This suggests that Unc-104, a member of an anterograde kinesin subfamily, contributes to an organelle-specific dynein-driven retrograde transport mechanism.
The SPARC-related Factor SMOC-2 Promotes Growth Factor-induced Cyclin D1 Expression and DNA Synthesis via Integrin-linked Kinase
Authors: Liu Peijun , Lu Jining , Cardoso Wellington V. , Vaziri Cyrus
Published: November 2007

Secreted modular calcium-binding protein-2 (SMOC-2) is a recently-identified SPARC-related protein of unknown function. In mRNA profiling experiments we, found that SMOC-2 expression was elevated in quiescent (G0) mouse fibroblasts and repressed after mitogenic stimulation with serum. The G0-specific expression of SMOC-2 was similar to that of platelet-derived growth factor-β receptor (PDGFβR), a major mitogenic receptor. Therefore, we tested a possible role for SMOC-2 in growth factor-induced cell cycle progression. SMOC-2 overexpression augmented DNA synthesis induced by serum and fibroblast mitogens (including PDGF-BB and basic fibroblast growth factor). Conversely, SMOC-2 ablation by using small interfering RNA attenuated DNA synthesis in response to PDGF-BB and other growth factors. Mitogen-induced expression of cyclin D1 was attenuated in SMOC-2–ablated cells, and cyclin D1-overexpressing cells were resistant to inhibition of mitogenesis after SMOC-2 ablation. Therefore, cyclin D1 is limiting for G1 progression in SMOC-2–deficient cells. SMOC-2 ablation did not inhibit PDGF-induced PDGFβR autophosphorylation or PDGF-BB–dependent activation of mitogen-activated protein kinase and Akt kinases, suggesting that SMOC-2 is dispensable for growth factor receptor activation. However, integrin-linked kinase (ILK) activity was reduced in SMOC-2–ablated cells. Ectopic expression of hyperactive ILK corrected the defective mitogenic response of SMOC-2–deficient cells. Therefore, SMOC-2 contributes to cell cycle progression by maintaining ILK activity during G1. These results identify a novel role for SMOC-2 in cell cycle control.
Large-Scale Analysis of Yeast Filamentous Growth by Systematic Gene Disruption and Overexpression
Authors: Jin Rui , Dobry Craig J. , McCown Phillip J. , Kumar Anuj
Published: November 2007

Under certain conditions of nutrient stress, the budding yeast Saccharomyces cerevisiae initiates a striking developmental transition to a filamentous form of growth, resembling developmental transitions required for virulence in closely related pathogenic fungi. In yeast, filamentous growth involves known mitogen-activated protein kinase and protein kinase A signaling modules, but the full scope of this extensive filamentous response has not been delineated. Accordingly, we have undertaken the first systematic gene disruption and overexpression analysis of yeast filamentous growth. Standard laboratory strains of yeast are nonfilamentous; thus, we constructed a unique set of reagents in the filamentous Σ1278b strain, encompassing 3627 integrated transposon insertion alleles and 2043 overexpression constructs. Collectively, we analyzed 4528 yeast genes with these reagents and identified 487 genes conferring mutant filamentous phenotypes upon transposon insertion and/or gene overexpression. Using a fluorescent protein reporter integrated at the MUC1 locus, we further assayed each filamentous growth mutant for aberrant protein levels of the key flocculence factor Muc1p. Our results indicate a variety of genes and pathways affecting filamentous growth. In total, this filamentous growth gene set represents a wealth of yeast biology, highlighting 84 genes of uncharacterized function and an underappreciated role for the mitochondrial retrograde signaling pathway as an inhibitor of filamentous growth.
Rintact: enabling computational analysis of molecular interaction data from the IntAct repository
Authors: Chiang Tony , Li Nianhua , Orchard Sandra , Kerrien Samuel , Hermjakob Henning , Gentleman Robert , Huber Wolfgang
Published: November 2007

Motivation: The IntAct repository is one of the largest and most widely used databases for the curation and storage of molecular interaction data. These datasets need to be analyzed by computational methods. Software packages in the statistical environment R provide powerful tools for conducting such analyses.

Results: We introduce Rintact, a Bioconductor package that allows users to transform PSI-MI XML2.5 interaction data files from IntAct into R graph objects. On these, they can use methods from R and Bioconductor for a variety of tasks: determining cohesive subgraphs, computing summary statistics, fitting mathematical models to the data or rendering graphical layouts. Rintact provides a programmatic interface to the IntAct repository and allows the use of the analytic methods provided by R and Bioconductor.

Availability:  Rintact is freely available at http://bioconductor.org

Contact:  huber@ebi.ac.uk
Statistical methods to infer cooperative binding among transcription factors inSaccharomyces cerevisiae
Authors: Datta Debayan , Zhao Hongyu
Published: November 2007

Motivation: Transcription factors regulate transcription in prokaryotes and eukaryotes by binding to specific DNA sequences in the regulatory regions of the genes. This regulation usually occurs in a coordinated manner involving multiple transcription factors. Genome-wide location data, also called ChIP-chip data, have enabled researchers to infer the binding sites for individual regulatory proteins. However, current methods to infer binding sites, such as simple thresholding based on p-values, are not optimal for a number of study objectives like combinatorial regulation, leading to potential loss of information. Hence, there is a need to develop more efficient statistical methods for analyzing such data.

Results: We propose to use log-linear models to study cooperative binding among transcription factors and have developed an Expectation-Maximization algorithm for statistical inferences. Our method is advantageous over simple thresholding methods both based on simulation and real data studies. We apply our method to infer the cooperative network of 204 regulators in Rich Medium and a subset of them in four different environmental conditions. Our results indicate that the cooperative network is condition specific; for a set of regulators, the network structure changes under different environmental conditions.

Availability: Our program is available at http://bioinformatics.med.yale.edu/TFcooperativity

Contact:  hongyu.zhao@yale.edu

Supplementary information: Supplementary information is available at Bioinformatics online.
Hierarchical tree snipping: clustering guided by prior knowledge
Authors: Dotan-Cohen Dikla , Melkman Avraham A. , Kasif Simon
Published: November 2007

Motivation: Hierarchical clustering is widely used to cluster genes into groups based on their expression similarity. This method first constructs a tree. Next this tree is partitioned into subtrees by cutting all edges at some level, thereby inducing a clustering. Unfortunately, the resulting clusters often do not exhibit significant functional coherence.

Results: To improve the biological significance of the clustering, we develop a new framework of partitioning by snipping—cutting selected edges at variable levels. The snipped edges are selected to induce clusters that are maximally consistent with partially available background knowledge such as functional classifications. Algorithms for two key applications are presented: functional prediction of genes, and discovery of functionally enriched clusters of co-expressed genes. Simulation results and cross-validation tests indicate that the algorithms perform well even when the actual number of clusters differs considerably from the requested number. Performance is improved compared with a previously proposed algorithm.

Availability: A java package is available at http://www.cs.bgu.ac.il/~dotna/ TreeSnipping

Contact:  dotna@cs.bgu.ac.il

Supplementary information: Supplementary data are available at Bioinformatics online.
Support Vector Machine-based classification of protein folds using the structural properties of amino acid residues and amino acid residue pairs
Authors: Shamim Mohammad Tabrez Anwar , Anwaruddin Mohammad , Nagarajaram H.A.
Published: November 2007

Motivation: Fold recognition is a key step in the protein structure discovery process, especially when traditional sequence comparison methods fail to yield convincing structural homologies. Although many methods have been developed for protein fold recognition, their accuracies remain low. This can be attributed to insufficient exploitation of fold discriminatory features.

Results: We have developed a new method for protein fold recognition using structural information of amino acid residues and amino acid residue pairs. Since protein fold recognition can be treated as a protein fold classification problem, we have developed a Support Vector Machine (SVM) based classifier approach that uses secondary structural state and solvent accessibility state frequencies of amino acids and amino acid pairs as feature vectors. Among the individual properties examined secondary structural state frequencies of amino acids gave an overall accuracy of 65.2% for fold discrimination, which is better than the accuracy by any method reported so far in the literature. Combination of secondary structural state frequencies with solvent accessibility state frequencies of amino acids and amino acid pairs further improved the fold discrimination accuracy to more than 70%, which is ∼8% higher than the best available method. In this study we have also tested, for the first time, an all-together multi-class method known as Crammer and Singer method for protein fold classification. Our studies reveal that the three multi-class classification methods, namely one versus all, one versus one and Crammer and Singer method, yield similar predictions.

Availability: Dataset and stand-alone program are available upon request.

Contact:  han@cdfd.org.in

Supplementary information: Supplementary data are available at Bioinformatics online.
Unsupervised feature selection under perturbations: meeting the challenges of biological data
Authors: Varshavsky Roy , Gottlieb Assaf , Horn David , Linial Michal
Published: November 2007

Motivation: Feature selection methods aim to reduce the complexity of data and to uncover the most relevant biological variables. In reality, information in biological datasets is often incomplete as a result of untrustworthy samples and missing values. The reliability of selection methods may therefore be questioned.

Method: Information loss is incorporated into a perturbation scheme, testing which features are stable under it. This method is applied to data analysis by unsupervised feature filtering (UFF). The latter has been shown to be a very successful method in analysis of gene-expression data.

Results: We find that the UFF quality degrades smoothly with information loss. It remains successful even under substantial damage. Our method allows for selection of a best imputation method on a dataset treated by UFF. More importantly, scoring features according to their stability under information loss is shown to be correlated with biological importance in cancer studies. This scoring may lead to novel biological insights.

Contact:  royke@cs.huji.ac.il

Supplementary information and code availability: Supplementary data are available at Bioinformatics online.