Functional analysis of the Galpha protein, GNA -3, in the development of Neurospora crassa using biochemical and genetic studies

Extracellular signals regulate fungal development and, to sense and respond to these cues, fungi evolved signal transduction pathways similar to those in mammalian systems. In fungi, heterotrimeric G proteins, composed of α, β, and γ subunits, transduce many signals, such as pheromones and nutrients, intracellularly to alter adenylyl cyclase and MAPK cascades activity. ^ Previously, the Gα proteins GNA-1 and GNA-2 were characterized in regulating development in the fungus Neurospora crassa. R. A. Baasiri isolated a third Gα, gna-3, and P. S. Rowley generated Δgna-3 mutants. GNA-3 belongs to a fungal Gα family that regulates cAMP metabolism and virulence. The Δ gna-3 sexual cycle is defective in homozygous crosses, producing inviable spores. Δgna-3 mutants have reduced aerial hyphae formation and derepressed asexual sporulation (conidiation), causing accumulation of asexual spores (conidia). These defects are similar to an adenylyl cyclase mutant, cr-1; cAMP supplementation suppressed Δ gna-3 and cr-1. Inappropriate conidiation and expression of a conidiation gene, con-10, were higher in Δ gna-3 than cr-1 submerged cultures; peptone suppressed conidiation. Adenylyl cyclase activity and expression demonstrated that GNA-3 regulates enzyme levels. ^ A Δgna-1 cr-1 was analyzed with F. D. Ivey to differentiate GNA-1 roles in cAMP-dependent and -independent pathways. Δ gna-1 cr-1 defects were worse than cr-1 and refractory to cAMP, suggesting that GNA-1 is necessary for sensing extracellular CAMP. Submerged culture conidiation was highest in Δgna-1 cr-1, and only high cell density Δgna-1 cultures conidiated, which correlated with con-10 levels. Transcription of a putative heat shock cognate protein was highest in Δgna-1 cr-1. ^ Functional relationships between the three Gαs was analyzed by constructing Δgna-1 Δgna-2 Δ gna-3, Δgna-1 Δgna-3, and Δgna-2 Δgna-3 strains. Δ gna-2 Δgna-3 strains exhibited intensified Δ gna-3 phenotypes; Δgna-1 Δgna-2 Δgna-3 and Δgna-1 Δ gna-3 strains were identical to Δgna-1 cr-1 on plates and were non-responsive to cAMP. The highest levels of conidiation and con-10 were detected in submerged cultures of Δ gna-1 Δgna-2 Δgna-3 and Δgna-1 Δgna-3 mutants, which was partially suppressed by peptone supplementation. Stimulation of adenylyl cyclase is completely deficient in Δgna-1 Δ gna-2 Δgna-3 and Δgna-1 Δ gna-3 strains. Δgna-3 and Δ gna-1 Δgna-3 aerial hyphae and conidiation defects were suppressed by mutation of a PKA regulatory subunit. ^

Structural and functional analysis of p84N5

Normal development and tissue homeostasis requires the carefully orchestrated balance between cell proliferation and cell death. Cell cycle checkpoints control the extent of cell proliferation. Cell death is coordinated through the activation of a cell suicide pathway that results in the morphologically recognizable form of death, apoptosis. Tumorigenesis requires that the balance between these two pathways be disrupted. The tumor suppressor protein Rb has not only been shown to be involved in the enforcement of cell cycle checkpoints, but has also been implicated in playing a role in the regulation of apoptosis. The manner in which Rb enforces cell cycle checkpoints has been well studied; however, its involvement in the regulation of apoptosis is still very unclear. p84N5 is a novel nuclear death domain containing protein that has been shown to interact with the N-terminus of Rb. The fact that it contains a death domain and the fact that it is nuclear localized possibly provides the first known mechanism for apoptotic signaling from the nucleus. The following study tested the hypothesis that the novel exclusively nuclear death domain containing protein p84N5 is an important mediator of programmed cell death and that its apoptotic function is reliant upon its nuclear localization and is regulated by unique functional domains within the p84N5 protein. We identified the p84N5 nuclear localization signal (NLS), eliminated it, and tested the functional significance of nuclear localization by using wild type and mutant sequences fused to EGFP-C1 (Clontech) to create wild type GFPN5 and subsequent mutants. The results of these assays demonstrated exclusive nuclear localization of GFPN5 is required for normal p84N5 induced apoptosis. We further conducted large-scale mutagenesis of the GFPN5 construct to identify a minimal region within p84N5 capable of interacting with Rb. We were able to identify a minimal sequence containing p84N5 amino acids 318 to 464 that was capable of interacting with Rb in co-immunoprecipitation assays. We continued by conducting a structural and functional analysis to identify the region or regions within p84N5 responsible for inducing apoptosis. Point mutations and small-scale deletions within the death domain of p84N5 lessened the effect but did not eliminate p84N5-induced cytotoxicity. Further analysis revealed that the minimal sequence of 318 to 464 of p84N5 was capable of inducing apoptosis to a similar degree as wild-type GFPN5 protein. Since amino acids 318 to 464 of p84N5 are capable of inducing apoptosis and interacting with Rb, we propose possible mechanisms whereby p84N5 may function in a Rb regulated manner. These results demonstrate that p84N5 induced apoptosis is reliant upon its nuclear localization and is regulated by unique functional domains within the p84N5 protein. ^

Sedimentological and clay mineralogical analysis of ODP sites in the central Fram Strait

This study presents the results of high-resolution sedimentological and clay mineralogical investigations on sediments from ODP Sites 908A and 909AlC located in the central Fram Strait. The objective was to reconstruct the paleoclimate and paleoceanography of the high northern latitudes since the middle Miocene. The sediments are characterised in particular by a distinctive input of ice-rafted material, which most probably occurs since 6 Ma and very likely since 15 Ma. A change in the source area at 1 1.2 Ma is clearly marked by variations within clay mineral composition and increasing accumulation rates. This is interpreted as a result of an increase in water mass exchange through the Fram Strait. A further period of increasing exchange between 4-3 Ma is identified by granulometric investigations and points to a synchronous intensification of deep water production in the North Atlantic during this time interval. A comparison of the components of coarse and clay fraction clearly shows that both are not delivered by the Same transport process. The input of the clay fraction can be related to transport mechanisms through sea ice and glaciers and very likely also through oceanic currents. A reconstruction of source areas for clay minerals is possible only with some restrictions. High smectite contents in middle and late Miocene sediments indicate a background signal produced by soil formation together with sediment input, possibly originating from the Greenland- Scotland Ridge. The applicability of clay mineral distribution as a climate proxy for the high northern latitudes can be confirmed. Based on a comparison of sediments from Site 909C, characterised by the smectite/illite and chlorite ratio, with regional and global climatic records (oxygen isotopes), a middle Miocene cooling phase between 14.8-14.6 Ma can be proposed. A further cooling phase between 10-9 Ma clearly shows similarities in its Progress toward drastic decrease in carbonate sedimentation and preservation in the eastern equatorial Pacific. The modification in sea water and atmosphere chemistry may represent a possible link due to the built-up of equatorial carbonate platforms. Between 4.8-4.6 Ma clay mineral distribution indicates a distinct cooling trend in the Fram Strait region. This is not accompanied by relevant glaciation, which would otherwise be indicated by the coarse fraction. The intensification of glaciation in the northern hemisphere is distinctly documented by a rapid increase of illite and chlorite starting from 3.3 Ma, which corresponds to oxygen isotope data trends from North Atlantic.

Deuterium analysis of ice core GISP2

The Greenland Ice Sheet Project 2 (GISP2) core can enhance our understanding of the relationship between parameters measured in the ice in central Greenland and variability in the ocean, atmosphere, and cryosphere of the North Atlantic Ocean and adjacent land masses. Seasonal (summer, winter) to annual responses of dD and deuterium excess isotopic signals in the GISP2 core to the seesaw in winter temperatures between West Greenland and northern Europe from A.D. 1840 to 1970 are investigated. This seesaw represents extreme modes of the North Atlantic Oscillation, which also influences sea surface temperatures (SSTs), atmospheric pressures, geostrophic wind strength, and sea ice extents beyond the winter season. Temperature excursions inferred from the dD record during seesaw/extreme NAO mode years move in the same direction as the West Greenland side of the seesaw. Symmetry with the West Greenland side of the seesaw suggests a possible mechanism for damping in the ice core record of the lowest decadal temperatures experienced in Europe from A.D. 1500 to 1700. Seasonal and annual deuterium excess excursions during seesaw years show negative correlation with dD. This suggests an isotopic response to a SST/ land temperature seesaw. The isotopic record from GISP2 may therefore give information on both ice sheet and sea surface temperature variability. Cross-plots of dD and d show a tendency for data to be grouped according to the prevailing mode of the seesaw, but do not provide unambiguous identification of individual seesaw years. A combination of ice core and tree ring data sets may allow more confident identification of GA and GB (extreme NAO mode) years prior to 1840.

Stable isotope analysis of Porites coral core EILAT-1

The environmental interpretation of the 13C/12C variations in the skeletons of massive corals is still a matter of debate. A 19-year seasonal skeletal 13C/12C record of a shallow-water Pontes coral from the northern Red Sea (Gulf of Aqaba) documents interannual events of extraordinarily large plankton blooms, indicated by anomalous 13C depletions in the coral skeleton. These blooms are caused by deep vertical water mass mixing, convectively driven in colder winters, which results in increased supplies of nutrients to the surface waters. The deep vertical mixings can sometimes be driven by the cooling occurring throughout the Middle East after large tropical volcanic eruptions. We therefore have evidence in our coral skeletal 13C/12C record for an indirect volcanic signal of the eruptions of El Chichón (1982) and Mount Pinatubo (1991). Deep mixing induced 13C/12C variations of the dissolved inorganic carbon in the surface waters can be neglected at this location. We therefore suggest that the 13C skeletal depletions can be best explained by changes in the coral's autotrophy-heterotrophy diet, through increased heterotrophic feeding on Zooplankton during the blooms. Increased feeding on 13C-depleted Zooplankton or increased heterotrophy at the expense of autotrophy can both result in a 13C-depleted coral skeleton. However, this suggestion requires more testing. If our conclusions are substantiated, seasonal skeletal 13C/12C records of corals which change from autotrophy under normal conditions to increased heterotrophy during bloom events may be used as indicators of ocean paleoproductivity at interannual resolution, available from no other source.

Grain size analysis of sediment core GeoB1510-2

Detailed information about the sediment properties and microstructure can be provided through the analysis of digital ultrasonic P wave seismograms recorded automatically during full waveform core logging. The physical parameter which predominantly affects the elastic wave propagation in water-saturated sediments is the P wave attenuation coefficient. The related sedimentological parameter is the grain size distribution. A set of high-resolution ultrasonic transmission seismograms (-50-500 kHz), which indicate downcore variations in the grain size by their signal shape and frequency content, are presented. Layers of coarse-grained foraminiferal ooze can be identified by highly attenuated P waves, whereas almost unattenuated waves are recorded in fine-grained areas of nannofossil ooze. Color -encoded pixel graphics of the seismograms and instantaneous frequencies present full waveform images of the lithology and attenuation. A modified spectral difference method is introduced to determine the attenuation coefficient and its power law a = kF. Applied to synthetic seismograms derived using a "constant Q" model, even low attenuation coefficients can be quantified. A downcore analysis gives an attenuation log which ranges from -700 dB/m at 400 kHz and a power of n=1-2 in coarse-grained sands to few decibels per meter and n :s; 0.5 in fine-grained clays. A least squares fit of a second degree polynomial describes the mutual relationship between the mean grain size and the attenuation coefficient. When it is used to predict the mean grain size, an almost perfect coincidence with the values derived from sedimentological measurements is achieved.

Analysis of frame-compatible subsampling structures for efficient 3DTV broadcast

The evolution of the television market is led by 3DTV technology, and this tendency can accelerate during the next years according to expert forecasts. However, 3DTV delivery by broadcast networks is not currently developed enough, and acts as a bottleneck for the complete deployment of the technology. Thus, increasing interest is dedicated to ste-reo 3DTV formats compatible with current HDTV video equipment and infrastructure, as they may greatly encourage 3D acceptance. In this paper, different subsampling schemes for HDTV compatible transmission of both progressive and interlaced stereo 3DTV are studied and compared. The frequency characteristics and preserved frequency content of each scheme are analyzed, and a simple interpolation filter is specially designed. Finally, the advantages and disadvantages of the different schemes and filters are evaluated through quality testing on several progressive and interlaced video sequences.

Molecular analysis of endo-β-mannanase genes upon seed imbibition suggest a cross-talk between radicle and micropylar endosperm during germination of Arabidopsis thaliana.

The endo-β-mannanase (MAN) family is represented in the Arabidopsis genome by eight members, all with canonical signal peptides and only half of them being expressed in germinating seeds. The transcripts of these genes were localized in the radicle and micropylar endosperm (ME) before radicle protrusion and this expression disappears as soon as the endosperm is broken by the emerging radicle tip. However, only three of these MAN genes, AtMAN5, AtMAN7 and especially AtMAN6 influence the germination time (t50) as assessed by the analysis of the corresponding knock-out lines. The data suggest a possible interaction between embryo and ME regarding the role of MAN during the Arabidopsis germination process.

Analysis of jitter influence in fast frequency measurements

This paper presents a theoretical analysis of possible jitter impact in application of numeric criterion for fastmeasurement of frequency by coincidence principle. The primary goal is the generation of a signal containing a known amount of each jitter components. This signal was used for testing signals with regular pulse trains. Initially, jitter components are analyzed and modeled individually. Next, sequences for combining different kinds of jitter are modeled, simulated and evaluated. Jitter model simulation in Matlab is utilized to show the independence of frequencymeasurement results on the total jitter present in the reference and desired pulse trains independently. A good agreement between previously introduced theory of fastmeasurement of frequency and simulation in jitter presence is verified; these results allows to engineers use the numeric criterion for fastmeasurement of frequency in spite to interactions among jitter components in various applications for frequency domain sensors.

Analysis of neuronal functions based on feynman diagrams

A method to study some neuronal functions, based on the use of the Feynman diagrams, employed in many-body theory, is reported. An equation obtained from the neuron cable theory is the basis for the method. The Green's function for this equation is obtained under some simple boundary conditions. An excitatory signal, with different conditions concerning high and pulse duration, is employed as input signal. Different responses have been obtained

Analysis of digital chaotic optical signals

The main objective of this paper is to present some tools to analyze a digital chaotic signal. We have proposed some of them previously, as a new type of phase diagrams with binary signals converted to hexadecimal. Moreover, the main emphasis will be given in this paper to an analysis of the chaotic signal based on the Lempel and Ziv method. This technique has been employed partly by us to a very short stream of data. In this paper we will extend this method to long trains of data (larger than 2000 bit units). The main characteristics of the chaotic signal are obtained with this method being possible to present numerical values to indicate the properties of the chaos.

Sparse spectral analysis of atrial fibrillation electrograms

Atrial fibrillation (AF) is a common heart disorder. One of the most prominent hypothesis about its initiation and maintenance considers multiple uncoordinated activation foci inside the atrium. However, the implicit assumption behind all the signal processing techniques used for AF, such as dominant frequency and organization analysis, is the existence of a single regular component in the observed signals. In this paper we take into account the existence of multiple foci, performing a spectral analysis to detect their number and frequencies. In order to obtain a cleaner signal on which the spectral analysis can be performed, we introduce sparsity-aware learning techniques to infer the spike trains corresponding to the activations. The good performance of the proposed algorithm is demonstrated both on synthetic and real data. RESUMEN. Algoritmo basado en técnicas de regresión dispersa para la extracción de las señales cardiacas en pacientes con fibrilación atrial (AF).

Hybrid analysis of nonlinear circuits: DAE models with indices zero and one

We extend in this paper some previous results concerning the differential-algebraic index of hybrid models of electrical and electronic circuits. Specifically, we present a comprehensive index characterization which holds without passivity requirements, in contrast to previous approaches, and which applies to nonlinear circuits composed of uncoupled, one-port devices. The index conditions, which are stated in terms of the forest structure of certain digraph minors, do not depend on the specific tree chosen in the formulation of the hybrid equations. Additionally, we show how to include memristors in hybrid circuit models; in this direction, we extend the index analysis to circuits including active memristors, which have been recently used in the design of nonlinear oscillators and chaotic circuits. We also discuss the extension of these results to circuits with controlled sources, making our framework of interest in the analysis of circuits with transistors, amplifiers, and other multiterminal devices.

Experimental investigation of radio signal propagation in scientific facilities for telerobotic applications

Understanding the radio signal transmission characteristics in the environment where the telerobotic application is sought is a key part of achieving a reliable wireless communication link between a telerobot and a control station. In this paper, wireless communication requirements and a case study of a typical telerobotic application in an underground facility at CERN are presented. Then, the theoretical and experimental characteristics of radio propagation are investigated with respect to time, distance, location and surrounding objects. Based on analysis of the experimental findings, we show how a commercial wireless system, such as Wi-Fi, can be made suitable for a case study application at CERN.