Functions of GCN5 and P /CAF acetyltransferases in mammalian development

Histone acetyltransferases are important chromatin modifiers that function as transcriptional co-activators. The identification of the transcriptional regulator GCN5 as the first nuclear histone acetyltransferase in yeast directly linked chromatin remodeling to transcriptional regulation. Although emerging evidence suggests that acetyltransferases participate in multiple cellular processes, their roles in mammalian development remain undefined. In this study, I have cloned and characterized the mouse homolog of GCN5 and a closely related protein P/CAF that interacts with p300/CBP. In contrast to yeast GCN5, but similar to P/CAF, mouse GCN5 possesses an additional N-terminal domain that confers the ability to acetylate nucleosomal histones. GCN5 and P/CAF exhibit identical substrate specificity and both interact with p300/CBP. Interestingly, expression levels of GCN5 and P/CAF display a complementary pattern in mouse embryos and in adult tissues, suggesting that they have distinct tissue or developmental stage specific roles. To define the in vivo function of GCN5 and P/CAF, I have generated mice that are nullizygous for GCN5 or P/CAF. P/CAF null mice are viable and fertile with no gross morphological defects, indicating that P/CAF is dispensable for development and p300/CBP function in vivo. In contrast, mice lacking GCN5 die between 10.5–11 days of gestation. GCN5 null mice are severely retarded but have anterior ectopic outgrowth. Molecular marker analyses reveal that early mesoderm is formed in GCN5 null mice but further differentiation into distinct mesodermal lineages is perturbed. While presomitic mesoderm and chodamesoderm are missing in GCN5 mutant mice, extraembryonic tissues and lateral mesoderm are unaffected. This is consistent with our finding that GCN5 expression is absent in the heart and extraembryonic tissues but is uniform throughout the rest of the embryo. Remarkably, GCN5 mutant mice exhibit an unusually high incidence of apoptosis in the embryonic ectoderm and mesoderm. Finally, mice doubly null for GCN5 and P/CAF die much earlier than mice harboring the GCN5 mutation alone, suggesting that P/CAF and GCN5 share some overlapping function during embryogenesis. This work is the first study to show that specific acetyltransferase is important for cell survival as well as mesoderm differentiation or maintenance during early mammalian development. ^

Characterization of cellular and molecular functions of the p21 -activated kinase, Shk1, in the fission yeast, Schizosaccharomyces pombe

The p21-activated kinase, Shk1, is an essential serine/threonine kinase required for normal cell polarity, proper mating response, and hyperosmotic stress response, in the fission yeast, Schizosaccharomyces pombe. This study has established a novel role for Shk1 as a microtubule regulator in fission yeast and, in addition, characterized a potential biological substrate of Shk1. Cells defective in Shk1 function were found to exhibit malformed interphase and mitotic microtubules, are hypersensitive to the microtubule disrupting drug thiabendazole (TBZ), and are cold sensitive for growth. Microtubule disruption by TBZ results in a significant reduction of Shk1 kinase activity, which is restored after cells are released from the drug, thus providing a correlation between Shk1 kinase activity and active microtubule polymerization. Consistent with a role for Shk1 as a microtubule regulator, GFP-Shk1 fusion proteins localize to interphase microtubules and mitotic microtubule spindles. Furthermore, loss of Tea1, a presumptive microtubule regulator in fission yeast, exacerbates the growth and microtubule defects of cells deficient in Shk1 function, and results in illicit Shk1 localization. Moreover, loss of the Cdc2 inhibitory kinase Wee1, which has been implicated as a mediator of the Shk1 pathway, leads to significant microtubule defects. Intriguingly, Wee1 protein levels are markedly reduced both by partial loss of Shk1 function and by treatment with TBZ. These results suggest that Shk1 is required for proper regulation of microtubule dynamics in fission yeast and may interact with Tea1 and Wee1 in this regulatory process. ^ To further understand Shk1 function in fission yeast, a yeast two-hybrid screen for proteins that interact with the Shk1 catalytic domain was performed. This screen led to the identification of a novel protein, Skb10 (for S&barbelow;hk1 k&barbelow;inase b&barbelow;inding protein 10). Coprecipitation experiments demonstrated that Skb10 associates with Shk1 in S. pombe cells. (Abstract shortened by UMI.) ^

Developmental functions of XKaiso, a transcriptional repressor associating with the p120 catenin in Xenopus laevis

The Armadillo family catenin proteins function in multiple capacities including cadherin-mediated cell-cell adhesion and nuclear signaling. The newest catenin, p120 catenin, differs from the classical catenins and binds to the membrane-proximal domain of cadherins. Recently, a novel transcription factor Kaiso was found to interact with p120 catenin, suggesting that p120 catenin also possesses a nuclear function. We isolated the Xenopus homolog of Kaiso, XKaiso, from a Xenopus stage 17 cDNA library. XKaiso contains an amino-terminal BTB/POZ domain and three carboxyl-terminal zinc fingers. The XKaiso transcript was present maternally and expressed throughout early embryonic development. XKaiso's spatial expression was defined via in situ hybridization and was found localized to the brain, eye, ear, branchial arches, and spinal cord. Co-immunoprecipitation of Xenopus p120 catenin and XKaiso demonstrated their mutual association, while related experiments employing differentially epitope-tagged XKaiso constructs suggest that XKaiso also self-associates. On the functional level, reporter assays employing a chimera of XKaiso fused to the GAL4 DNA binding domain indicated that XKaiso is a transcriptional repressor. To better understand the significance of the Kaiso-p120 catenin complex in vertebrate development, Kaiso knock-down experiments were undertaken, and the modulatory role of p120 catenin in Kaiso function examined during Xenopus development. Using morpholino antisense oligonucleotides to block translation of XKaiso, XKaiso was found to be essential for Xenopus gastrulation, being required for correct morphogenetic movements in early embryogenesis. Molecular marker analyses indicated that one target gene of the Wnt/β-catenin pathway, Siamois, is significantly increased in embryos depleted for XKaiso, while other dorsal, ventral, and mesodermal cell fate markers were unaltered. In addition, the non-canonical Wnt-11, known to participate in planar cell polarity/convergent extension processes, was significantly upregulated following depletion of XKaiso. Such increased Wnt-11 expression likely contributed to the XKaiso depletion phenotype because a dominant negative form of Wnt-11 or of the downstream effector Dishevelled partially rescued the observed gastrulation defects. These results show that XKaiso is essential for proper gastrulation movements, resulting at least in part from its modulation of non-canonical Wnt signaling. The significance of the XKaiso-p120 catenin interaction has yet to be determined, but appears to include a role in modulating genes promoting canonical and non-canonical Wnt signals. ^

Tissue metal concentrations and metal transfer functions of Glaucous Gulls (Larus hyperboreus) from Bjørnøya and Jan Mayen

Heavy metals (Cd, Cu, Fe, Mn and Zn) concentrations were determined in different tissues (muscle, kidney, liver, brain, gonads, heart and feathers) of Glaucous Gulls (Larus hyperboreus) from Bjornoya and Jan Mayen. The age and spatial dependent variations in heavy metals were quantified and interpreted in view of the three chemometric techniques, i.e. non-parametric Mann-Whitney U test, redundancy gradient analysis and detrended correspondence analysis. The Glaucous Gulls from Bjornoya contained significantly higher (p < 0.05) levels of Cd, Cu and Zn than those inhabited Jan Mayen. Adult birds were characterized by greater (p < 0.01) concentration of muscle, hepatic and renal heavy metals in comparison to chicks. Insignificantly higher slope constant Zn/Cd for the liver than for the kidney may reflect insignificant Cd exposure. Estimate of transfer factor (TF) allows us to assess variations in heavy metal concentrations during the individual development of Glaucous Gulls. It may be stated that there is a distinct increase of bioaccumulation of all the studied metals during subsequent stages of the bird life.

Statistical Calculation of the Main Reliability Functions of GaAs Concentrator III-V Solar Cells

This paper presents some of the results of a method to determine the main reliability functions of concentrator solar cells. High concentrator GaAs single junction solar cells have been tested in an Accelerated Life Test. The method can be directly applied to multi-junction solar cells. The main conclusions of this test carried out show that these solar cells are robust devices with a very low probability of failure caused by degradation during their operation life (more than 30 years). The evaluation of the probability operation function (i.e. the reliability function R(t)) is obtained for two nominal operation conditions of these cells, namely simulated concentration ratios of 700 and 1050 suns. Preliminary determination of the Mean Time to Failure indicates a value much higher than the intended operation life time of the concentrator cells.

Boundary element approach to the dynamic stiffness functions of circular foundations

A boundary element approach for time harmonic axisymmetric problems using the complete space point load fundamental solution is presented. The fundamental solution is integrated numerically along the azimuthal co-ordinate of each axisymmetric element. To increase the accuracy of the numerical integration a simple co-ordinate transformation is proposed. The approach is applied to the computation of the dynamic stiffness functions of rigid circular foundations on layered viscoelastic soils. Three different sites are considered: a uniform half-space, a soil layer on a half-space, and a soil consisting of four horizontal layers and a compliant half-space. The numerical results obtained by the proposed approach for surface circular foundations are very close to corresponding published results obtained by different procedures.

Vector spaces of entire functions of unbounded type

Let E be an infinite dimensional complex Banach space. We prove the existence of an infinitely generated algebra, an infinite dimensional closed subspace and a dense subspace of entire functions on E whose non-zero elements are functions of unbounded type. We also show that the τδ topology on the space of all holomorphic functions cannot be obtained as a countable inductive limit of Fr´echet spaces. RESUMEN. Sea E un espacio de Banach complejo de dimensión infinita y sea H(E) el espacio de funciones holomorfas definidas en E. En el artículo se demuestra la existencia de un álgebra infinitamente generada en H(E), un subespacio vectorial en H(E) cerrado de dimensión infinita y un subespacio denso en H(E) cuyos elementos no nulos son funciones de tipo no acotado. También se demuestra que el espacio de funciones holomorfas con la topología ? no es un límite inductivo numberable de espacios de Fréchet.

On real analytic functions of unbounded type

In this paper we prove several results on the existence of analytic functions on an infinite dimensional real Banach space which are bounded on some given collection of open sets and unbounded on others. In addition, we also obtain results on the density of some subsets of the space of all analytic functions for natural locally convex topologies on this space. RESUMEN. Los autores demuestran varios resultados de existencia de funciones analíticas en espacios de Banach reales de dimensión infinita que están acotadas en un colección de subconjuntos abiertos y no acotadas en los conjuntos de otra colección. Además, se demuestra la densidad de ciertos subconjuntos de funciones analíticas para varias topologías localmente convexas.

Leader Rural Development Program in Spain: Profile and Functions of Workers in Local Action Groups.

Occupations in the labor market are linked with to a minimum basic training and other capacities. Hired workers should be able to accomplish required functions related to their specific job. Regarding the rural development labor market, local action groups? workers have defined performance areas?projects, strategy, organization and training & market?but specific functions within each of these areas are not as clearly defined. Neither both, basic training and capacities needed to perform each job profile within the local action group are defined. This communication analyses training and other capacities linked to each of the job profiles within the local action group. Functions within each of the performance areas previously defined are also analyzed regarding the job profiles.

Singularities and undefinitions in the calibration functions of sonic anemometers

A mathematical model of the process employed by a sonic anemometer to build up the measured wind vector in a steady flow is presented to illustrate the way the geometry of these sensors as well as the characteristics of aerodynamic disturbance on the acoustic path can lead to singularities in the transformation function that relates the measured (disturbed) wind vector with the real (corrected) wind vector, impeding the application of correction/calibration functions for some wind conditions. An implicit function theorem allows for the identification of those combinations of real wind conditions and design parameters that lead to undefined correction/ calibration functions. In general, orthogonal path sensors do not show problematic combination of parameters. However, some geometric sonic sensor designs, available in the market, with paths forming smaller angles could lead to undefined correction functions for some levels of aerodynamic disturbances and for certain wind directions. The parameters studied have a strong influence on the existence and number of singularities in the correction/ calibration function as well as on the number of singularities for some combination of parameters. Some conclusions concerning good design practices are included.

Segregating the functions of human hippocampus

It is now accepted that hippocampal lesions impair episodic memory. However, the precise functional role of the hippocampus in episodic memory remains elusive. Recent functional imaging data implicate the hippocampus in processing novelty, a finding supported by human in vivo recordings and event-related potential studies. Here we measure hippocampal responses to novelty, using functional MRI (fMRI), during an item-learning paradigm generated from an artificial grammar system. During learning, two distinct types of novelty were periodically introduced: perceptual novelty, pertaining to the physical characteristics of stimuli (in this case visual characteristics), and exemplar novelty, reflecting semantic characteristics of stimuli (in this case grammatical status within a rule system). We demonstrate a left anterior hippocampal response to both types of novelty and adaptation of these responses with stimulus familiarity. By contrast to these novelty effects, we also show bilateral posterior hippocampal responses with increasing exemplar familiarity. These results suggest a functional dissociation within the hippocampus with respect to the relative familiarity of study items. Neural responses in anterior hippocampus index generic novelty, whereas posterior hippocampal responses index familiarity to stimuli that have behavioral relevance (i.e., only exemplar familiarity). These findings add to recent evidence for functional segregation within the human hippocampus during learning.