Fenología y capacidad de dispersión de Monochamus galloprovincialis (Olivier 1795) en la Península Ibérica = Phenology and dispersal ability of Monochamus galloprovincialis (Olivier 1795) at Iberian Peninsula

La aparición y avance de la enfermedad del marchitamiento del pino (Pine Wilt Desease, PWD), causada por Bursaphelenchus xylophilus (Nematoda; Aphelenchoididae), el nematodo de la madera del pino (NMP), en el suroeste de Europa, ha puesto de manifiesto la necesidad de estudiar la fenología y la dispersión de su único vector conocido en Europa, Monochamus galloprovincialis (Col., Cerambycidae). El análisis de 12 series de emergencias entre 2010 y 2014, registradas en Palencia, València y Teruel, con material procedente de diversos puntos de la península ibérica, demostró una alta variabilidad en la fenología de M. galloprovincialis y la divergencia térmica respecto de las poblaciones portuguesas. Para éstas, el establecimiento de los umbrales térmicos de desarrollo de las larvas post-dormantes del vector (12,2 y 33,5ºC) permitió la predicción de la emergencia mediana para la fecha en la que se acumulaban de 822 grados-día. Ninguna de las series analizadas en este trabajo necesitó de dichos grados-día estimados para la emergencia mediana. Asimismo, la emergencia se adelantó en las regiones más calurosas, mientras que se retrasó en las zonas más templadas. Más allá de la posible variabilidad entre poblaciones locales peninsulares, se detectaron indicios de que la diferencia en la acumulación de calor durante el otoño puede afectar el grado de maduración de las larvas invernantes, y su posterior patrón temporal de emergencia. Por último, también fueron observados comportamientos de protandria en las emergencias. Respecto a la fenología de su vuelo, entre los años 2010 y 2015, fueron ejecutados un total de 8 experimentos de captura de M. galloprovincialis mediante trampas cebadas con atrayentes en diferentes regiones (Castellón, Teruel, Segovia y Alicante) permitiendo el seguimiento del periodo de vuelo. Su análisis permitió constatar la disminución de las capturas y el acortamiento del periodo de vuelo con la altitud, el inicio del vuelo en el mes de mayo/junio a partir de los 14ºC de temperatura media diaria, la influencia de las altas temperaturas en la disminución de las capturas estivales (potencial causante de perfiles bimodales en las curvas de vuelo en las zonas menos frías), la evolución de la proporción de sexos a lo largo del periodo de vuelo (que muestra una mayor captura de hembras al inicio y de machos al final) y el comportamiento diurno y ligado a las altas temperaturas del vuelo circadiano del insecto. Dos redes de muestreo sistemático de insectos saproxílicos instaladas en la Comunitat Valencia (Red MUFFET, 15 parcelas, año 2013) y en Murcia (Red ESFP, 20 parcelas, años 2008-2010) permitieron el estudio de la comunidad de insectos relacionada con M. galloprovincialis. Cada una de las parcelas contaba con una trampa cebada con atrayentes y una estación meteorológica. El registro de más de 250 especies de coleópteros saproxílicos demostró el potencial que tiene el empleo de redes de trampas vigía para la detección temprana de organismos exóticos, además de permitir la caracterización y evaluación de las comunidades de entomofauna útil, representando una de las mejores herramientas de la gestión integrada de plagas. En este caso, la comunidad de saproxílicos estudiada mostró ser muy homogénea respecto a la variación ambiental de las zonas de muestreo, y que pese a las pequeñas variaciones entre las comunidades de los diferentes ecosistemas, el rol que M. galloprovincialis desempeña en ellas a lo largo de todo el gradiente estudiado es el mismo. Con todo, el análisis mediante redes de interacción mostró su relevancia ecológica al actuar de conector entre los diferentes niveles tróficos. Por último, un total de 12 experimentos de marcaje-liberación-recaptura desarrollados entre 2009 y 2012 en Castellón, Teruel, Valencia y Murcia permitieron evaluar el comportamiento dispersivo de M. galloprovincialis. Las detecciones mediante trampas cebadas de los insectos liberados se dieron por lo menos 8 días después de la emergencia. La abundancia de población pareció relacionada con la continuidad, la naturalización de la masa, y con la afección previa de incendios. La dispersión no estuvo influida por la dirección ni la intensidad de los vientos dominantes. La abundancia de material hospedante (en lo referente a las variables de masa y a los índices de competencia) influyó en la captura del insecto en paisajes fragmentados, aunque la ubicación de las trampas optimizó el número de capturas cuando se ubicaron en el límite de la masa y en zonas visibles. Por último también se constató que M. galloprovincialis posee suficiente capacidad de dispersión como para recorrer hasta 1500 m/día, llegando a alcanzar distancias máximas de 13600m o de 22100 m. ABSTRACT The detection and expansion of the Pine Wilt Desease (PWD), caused by Bursaphelenchus xylophilus (Nematoda; Aphelenchoididae), Pine Wood Nematode (PWN), in southwestern Europe since 1999, has triggered off the study of the phenology and the dispersion of its unique vector in the continent, Monochamus galloprovincialis (Coleoptera, Cerambycidae). The analysis of 12 emergence series between 2010 and 2014 registered in Palencia, Teruel and Valencia (Spain), registered from field colonized material collected at several locations of the Iberian Peninsula, showed a high variability in the emergence phenology of M. galloprovincialis. In addition, these patterns showed a very acute thermal divergence regarding a development model fitted earlier in Portugal. Such model forecasted the emergence of 50% of M. galloprovincialis individuals in the Setúbal Peninsula (Portugal) when an average of 822 degree-days (DD) were reached, based on the accumulation of heat from the 1st of March until emergence and lower and upper thresholds of 12.2 ºC and 33,5 °C respectively. In our results, all analyzed series needed less than 822 DD to complete the 50% of the emergence. Also, emergency occurred earlier in the hottest regions, while it was delayed in more temperate areas. Beyond the possible variability between local populations, the difference in the heat accumulation during the fall season may have affected the degree of maturation of overwintering larvae, and subsequently, the temporal pattern of M. galloprovincialis emergences. Therefore these results suggest the need to differentiate local management strategies for the PWN vector, depending on the location, and the climatic variables of each region. Finally, protandrous emergence patterns were observed for M. galloprovincialis in most of the studied data-sets. Regarding the flight phenology of M. galloprovincialis, a total of 8 trapping experiments were carried out in different regions of the Iberian Peninsula (Castellón, Teruel, Segovia and Alicante) between 2010 and 2015. The use of commercial lures and traps allowed monitoring of the flight period of M. galloprovincialis. The analyses of such curves, helped confirming different aspects. First, a decline in the number of catches and a shortening of the flight period was observed as the altitude increased. Flight period was recorded to start in May / June when the daily average temperature went over 14 ° C. A significant influence of high temperatures on the decrease of catches in the summer was found in many occasions, which frequently lead to a bimodal profile of the flight curves in warm areas. The evolution of sex ratio along the flight period shows a greater capture of females at the beginning of the period, and of males at the end. In addition, the circadian response of M. galloprovincialis to lured traps was described for the first time, concluding that the insect is diurnal and that such response is linked to high temperatures. Two networks of systematic sampling of saproxylic insects were installed in the Region of Valencia (Red MUFFET, 15 plots, 2013) and Murcia (Red ICPF, 20 plots, 2008-2010). These networks, intended to serve the double purpose of early-detection and long term monitoring of the saproxylic beetle assemblies, allowed the study of insect communities related to M. galloprovincialis. Each of the plots had a trap baited with attractants and a weather station. The registration of almost 300 species of saproxylic beetles demonstrated the potential use of such trapping networks for the early detection of exotic organisms, while at the same time allows the characterization and evaluation of useful entomological fauna communities, representing one of the best tools for the integrated pest management. In this particular case, the studied community of saproxylic beetles was very homogeneous with respect to environmental variation of the sampling areas, and despite small variations between communities of different ecosystems, the role that M. galloprovincialis apparently plays in them across the studied gradient seems to be the same. However, the analysis through food-webs showed the ecological significance of M. galloprovincialis as a connector between different trophic levels. Finally, 12 mark-release-recapture experiments were carried out between 2009 and 2012 in Castellón, Teruel, Valencia and Murcia (Spain) with the aim to describe the dispersive behavior of M. galloprovincialis as well as the stand and landscape characteristics that could influence its abundance and dispersal. No insects younger than 8 days were caught in lured traps. Population abundance estimates from mark-release-recapture data, seemed related to forest continuity, naturalization, and to prior presence of forest fires. On the other hand, M. galloprovincialis dispersal was not found to be significantly influenced by the direction and intensity of prevailing winds. The abundance of host material, very related to stand characteristics and spacing indexes, influenced the insect abundance in fragmented landscapes. In addition, the location of the traps optimized the number of catches when they were placed in the edge of the forest stands and in visible positions. Finally it was also found that M. galloprovincialis is able to fly up to 1500 m / day, reaching maximum distances of up to 13600 m or 22100 m.

Arquitectura modal o ensamblaje de repertorios, procesos y disposiciones

En muchas ocasiones se ha establecido una crítica sobre la excesiva disposicionalidad estética del arquitecto: el abuso subjetivista, un innatismo desligado del entorno socioambiental o su desafección hacia las necesidades y los requerimientos pertinentes del usuario final, buscando, principalmente, la objetivación cultural como expresión personal e individualista. Criticas que responden a un malestar persistente, que subyace a la práctica arquitectónica. En otras ocasiones se han ideado y propuesto tentativas o aproximaciones para la corrección y regulación de dicho “exceso”. En busca de la “pretendida objetividad”, éstas han recurrido a un pasado prístino, a una tradición inconsciente, colectiva e impersonal, a la ausencia de autoría, a lo artesanal y a lo vernáculo. También hacia un futuro tecnocrático: al mesianismo basado en la máquina, la ciencia, los métodos y el progreso tecnológico. La búsqueda de un lenguaje compartido, de un diseño sistemático y de una adecuación funcional han establecido nuevos excesos –repertoriales-, relegando la creatividad y la concepción estética a un plano casi inexistente, subordinándolo a variables de diseño externas a la autoría y la expresión subjetiva del diseñador. Esta tensión se ha saldado en una pugna de binomios categoriales que reducen la realidad a un dualismo aporético y contrapuesto, difícilmente superable. El propósito articulatorio de dichas posturas, en modelos ontológicos integrales, permite la incorporación de dichas categorías bajo estructuras y ensamblajes que despliegan los modos de relación, mediación y producción arquitectónicas. Estos ensamblajes combinan los distintos agentes –disposicionalidades- y sus repertorios en cada una de las fases del proceso arquitectónico, construyendo modos de acción y relación en base a los acoplamientos diferenciales, que devienen en técnicas y dinámicas de producción arquitectónica. Cada modo integrará diversas variables en base a los modelos propuestos -estratos, períodos y polos- y a la elección intencional –“situada”- de cada práctica: la Autonomía Modal Arquitectónica. El producto final de cada modo –o ensamblaje- será inconmensurable, estableciendo sistemas y métodos diversos para la práctica estética, sin que por ello esté garantizado el valor o éxito del resultado. ABSTRACT On many occasions an aesthetic criticism of the excessive disposition of the architect´s subjectivist abuse, innate posture detached from socioenvironmental issues or disaffection regarding the needs and requirements of who -or what- will finally use it has been brought up mainly in pursuit of a cultural objectification as a personal expression and an individualistic one. It is a criticism which goes hand in hand with a persistent uneasiness which is interwoven in the architectural practice. On other occasions we have seen ideas, attempts and approaches developed and proposed in order to correct, improve or adjust that so-called “excess”. In the search for this " expected objectivity" theorists have resorted to delving into a pristine past, an unconscious tradition both collective and personal, to the absence of authorship, regarding craft and vernacular. The pursuit of a common language, a systematic design and a functional adequacy has established new excesses -repertoires-, leaving behind creativity and aesthetic conception at an almost nonexistent level and subordinating it to variables outside the authoring design and subjective expression of the designer. This tension has resulted in a conflict between categorical binomial, which reduces reality to an aporetic and opposed dualism which is not easily overcome. The purpose of such articulatory positions in comprehensive ontological modals allows for the incorporation of those categories under structures and assemblies which display and unfold different models of relationship, mediation and production of architecture. These assemblies combine different agents - disposition - and their repertoires in each of the phases of the architectural process, building actions and relational modes based on the differential couplings, which become techniques and dynamics in architectural production. Each mode will integrate several variables within the proposed ontological models - strata, periods and poles - based on an intentional choice –“situated knowledge”- of each practice: Architectural Modal Autonomy. The final product of each mode -or assembly- will be immeasurable, establishing different systems and methods for aesthetic practice without the value or success of the outcome ever being guaranteed.

ANÁLISE SISTÊMICA DAS RELAÇÕES DE PODER EM GRUPOS DE LIDERANÇA OFICIAL DE IGREJAS DE GOVERNO CONGREGACIONAL: Estudo de Caso em Igrejas Menonitas de Curitiba

A pesquisa a seguir tem o poder como objeto de estudos, especificamente na forma de relações de poder em conselhos diretores (pastoral e administrativo) de igrejas menonitas em Curitiba. A forma presumida de governo dessas igrejas é historicamente congregacional, isto é, as assembléias das respectivas igrejas são supremas e últimas nas deliberações. A metodologia de pesquisa é sociológica, indutiva, funcionalista, com tendências fenomenológicas, tendo como perspectiva principal a teoria sistêmica e seus recursos de pesquisa. A teoria geral dos sistemas orienta os fundamentos da pesquisa, a teoria dos sistemas sociais é o arcabouço da pesquisa. O método de pesquisa é a observação-participante com registros em vídeo e áudio, transcrição, análise e elaboração de conclusões. A técnica de pesquisa citada demonstrou-se eficaz e útil para o levantamento de dados em pesquisas de campo que tratam de grupos e suas funções diferenciadas em agremiações eclesiásticas. Os objetivos buscam identificar a dinâmica sistêmica nas reuniões dos conselhos observados, bem como identificar o processo das relações de poder nos mesmos. As hipóteses lançadas como ponto de partida afirmam que a teoria sistêmica em qualquer perspectiva é observável, bem como os elementos (conselheiros) das relações de poder se demonstram inconscientes de seus atos de poder. As hipóteses principais e secundárias foram confirmadas pela observação-participante, a saber: a pesquisa verifica a hipótese geral de que mesmo igrejas e seus sistemas sociais evidenciam princípios sistêmicos segundo a teoria geral dos sistemas e a teoria dos sistemas sociais de Niklas Luhmann. As hipóteses específicas verificam se os conselheiros dos grupos de liderança evidenciam consciência do poder exercido e do poder implícito em suas funções e papéis; se os grupos formais, eleitos pela assembléia exercem poder de fato ou se existe influência do poder informal; se existe uma correlação, entre o poder formal e as fronteiras rígidas , o poder informal e a fronteira difusa ; se é possível trabalhar preventivamente e interventivamente através do conceito de relações de poder e os princípios sociais sistêmicos. As igrejas menonitas de Curitiba de maneira geral preservam traços de governo congregacional, mas, a transição para estilos de governo pastorcêntricos e autocráticos a partir dos conselhos observados é um fato e parece irreversível. Aparentemente a causalidade desse movimento surge no contexto social metropolitano e suas implicações, mais do que numa mudança estrategicamente planejada pelas lideranças. Portanto, a transição parece ser cultural. Aliás, as transformações sociais das tradições menonitas são diretamente proporcionais à sua inclusão e inculturação no contexto social em que se situam, confirmando assim os princípios sistêmicos da sociedade em geral.(AU)

ANÁLISE SISTÊMICA DAS RELAÇÕES DE PODER EM GRUPOS DE LIDERANÇA OFICIAL DE IGREJAS DE GOVERNO CONGREGACIONAL: Estudo de Caso em Igrejas Menonitas de Curitiba

A pesquisa a seguir tem o poder como objeto de estudos, especificamente na forma de relações de poder em conselhos diretores (pastoral e administrativo) de igrejas menonitas em Curitiba. A forma presumida de governo dessas igrejas é historicamente congregacional, isto é, as assembléias das respectivas igrejas são supremas e últimas nas deliberações. A metodologia de pesquisa é sociológica, indutiva, funcionalista, com tendências fenomenológicas, tendo como perspectiva principal a teoria sistêmica e seus recursos de pesquisa. A teoria geral dos sistemas orienta os fundamentos da pesquisa, a teoria dos sistemas sociais é o arcabouço da pesquisa. O método de pesquisa é a observação-participante com registros em vídeo e áudio, transcrição, análise e elaboração de conclusões. A técnica de pesquisa citada demonstrou-se eficaz e útil para o levantamento de dados em pesquisas de campo que tratam de grupos e suas funções diferenciadas em agremiações eclesiásticas. Os objetivos buscam identificar a dinâmica sistêmica nas reuniões dos conselhos observados, bem como identificar o processo das relações de poder nos mesmos. As hipóteses lançadas como ponto de partida afirmam que a teoria sistêmica em qualquer perspectiva é observável, bem como os elementos (conselheiros) das relações de poder se demonstram inconscientes de seus atos de poder. As hipóteses principais e secundárias foram confirmadas pela observação-participante, a saber: a pesquisa verifica a hipótese geral de que mesmo igrejas e seus sistemas sociais evidenciam princípios sistêmicos segundo a teoria geral dos sistemas e a teoria dos sistemas sociais de Niklas Luhmann. As hipóteses específicas verificam se os conselheiros dos grupos de liderança evidenciam consciência do poder exercido e do poder implícito em suas funções e papéis; se os grupos formais, eleitos pela assembléia exercem poder de fato ou se existe influência do poder informal; se existe uma correlação, entre o poder formal e as fronteiras rígidas , o poder informal e a fronteira difusa ; se é possível trabalhar preventivamente e interventivamente através do conceito de relações de poder e os princípios sociais sistêmicos. As igrejas menonitas de Curitiba de maneira geral preservam traços de governo congregacional, mas, a transição para estilos de governo pastorcêntricos e autocráticos a partir dos conselhos observados é um fato e parece irreversível. Aparentemente a causalidade desse movimento surge no contexto social metropolitano e suas implicações, mais do que numa mudança estrategicamente planejada pelas lideranças. Portanto, a transição parece ser cultural. Aliás, as transformações sociais das tradições menonitas são diretamente proporcionais à sua inclusão e inculturação no contexto social em que se situam, confirmando assim os princípios sistêmicos da sociedade em geral.(AU)

(In)VERSÕES POLÍTICO-ESCATOLÓGICAS NO PENTECOSTALISMO BRASILEIRO Uma análise da posição e ação política das Assembleias de Deus de 1930-1945 e 1978-1988 a partir do jornal Mensageiro da Paz

A presente pesquisa analisou a posição e ação política nas Assembleias de Deus do Brasil nos períodos 1930-1945 e 1978-1988. Defendemos a tese de que desde 1930 há no interior do pentecostalismo brasileiro posições e intervenções no mundo da política. Tanto no período de 1930-1945 como o de 1978-1988 nossas análises serão realizadas a partir das temporalidades discutidas por Giorgio Agamben: chronos, aiôn e kairos. No que diz respeito ao primeiro período 1930-1945, as pesquisas quase sempre vinculam o discurso escatológico do pentecostalismo a processos de alienação e não envolvimento com a política partidária. Entretanto, acredita-se que as narrativas escatológicas não foram causa de certo afastamento da esfera pública brasileira, mas sim efeito de processos de exclusão aos quais homens e mulheres de pertença pentecostal estiveram circunscritos. Doutrinas como a escatologia e a pneumatologia foram potencializadoras de processos que aqui denominamos de biopotência. Já no segundo período, de 1978-1988, a posição e a ação política que predominaram no pentecostalismo estiveram relacionadas com a biopolítica. Chamamos de capítulo intermedário ou de transição o período correspondente às datas 1946-1977. Nele descreveremos e analisaremos personalidades pentecostais de destaque no campo da política brasileira. Metodologicamente, fizemos nossa análise a partir de artigos publicados no órgão oficial de comunicação da denominação religiosa em questão, o jornal Mensageiro da Paz. Esse periódico circula desde 1930. Além dos artigos, destacamos também as autoras e os autores, todas elas e todos eles figuras de destaque no assembleianismo. Ao longo da pesquisa questionamos a ideia do apoliticismo pentecostal. Defendemos a tese de que desde 1930, que é o início de nossa pesquisa, há posição e ação política nas Assembleias de Deus. Como resultado disso, questionamos a ideia do apoliticismo pentecostal. Nossa hipótese é de que no período 1930-1945 o pentecostalismo foi um polo de biopotência. Se a biopolítica é o poder sobre a vida, a biopotência é o poder da vida. Doutrinas como a escatologia e pneumatologia contribuíram para que nos espaços marginais onde se reuniam os pentecostais fossem criados novos modelos de sociabilidade e de cooperação; eram também espaços de criação de outras narrativas e de crítica a modelos hegemônicos e excludentes. O pentecostalismo foi um movimento que promoveu a dignidade humana de sujeitos subalternos.

In situ atomic force microscopy study of Alzheimer’s β-amyloid peptide on different substrates: New insights into mechanism of β-sheet formation

We have applied in situ atomic force microscopy to directly observe the aggregation of Alzheimer’s β-amyloid peptide (Aβ) in contact with two model solid surfaces: hydrophilic mica and hydrophobic graphite. The time course of aggregation was followed by continuous imaging of surfaces remaining in contact with 10–500 μM solutions of Aβ in PBS (pH 7.4). Visualization of fragile nanoscale aggregates of Aβ was made possible by the application of a tapping mode of imaging, which minimizes the lateral forces between the probe tip and the sample. The size and the shape of Aβ aggregates, as well as the kinetics of their formation, exhibited pronounced dependence on the physicochemical nature of the surface. On hydrophilic mica, Aβ formed particulate, pseudomicellar aggregates, which at higher Aβ concentration had the tendency to form linear assemblies, reminiscent of protofibrillar species described recently in the literature. In contrast, on hydrophobic graphite Aβ formed uniform, elongated sheets. The dimensions of those sheets were consistent with the dimensions of β-sheets with extended peptide chains perpendicular to the long axis of the aggregate. The sheets of Aβ were oriented along three directions at 120° to each other, resembling the crystallographic symmetry of a graphite surface. Such substrate-templated self-assembly may be the distinguishing feature of β-sheets in comparison with α-helices. These studies show that in situ atomic force microscopy enables direct assessment of amyloid aggregation in physiological fluids and suggest that Aβ fibril formation may be driven by interactions at the interface of aqueous solutions and hydrophobic substrates, as occurs in membranes and lipoprotein particles in vivo.

Mass spectrometry of ribosomes and ribosomal subunits

Nanoflow electrospray ionization has been used to introduce intact Escherichia coli ribosomes into the ion source of a mass spectrometer. Mass spectra of remarkable quality result from a partial, but selective, dissociation of the particles within the mass spectrometer. Peaks in the spectra have been assigned to individual ribosomal proteins and to noncovalent complexes of up to five component proteins. The pattern of dissociation correlates strongly with predicted features of ribosomal protein–protein and protein–RNA interactions. The spectra allow the dynamics and state of folding of specific proteins to be investigated in the context of the intact ribosome. This study demonstrates a potentially general strategy to probe interactions within complex biological assemblies.

Recurrent excitatory postsynaptic potentials induced by synchronized fast cortical oscillations

Gamma frequency (about 20–70 Hz) oscillations occur during novel sensory stimulation, with tight synchrony over distances of at least 7 mm. Synchronization in the visual system has been proposed to reflect coactivation of different parts of the visual field by a single spatially extended object. We have shown that intracortical mechanisms, including spike doublet firing by interneurons, can account for tight long-range synchrony. Here we show that synchronous gamma oscillations in two sites also can cause long-lasting (>1 hr) potentiation of recurrent excitatory synapses. Synchronous oscillations lasting >400 ms in hippocampal area CA1 are associated with an increase in both excitatory postsynaptic potential (EPSP) amplitude and action potential afterhyperpolarization size. The resulting EPSPs stabilize and synchronize a prolonged beta frequency (about 10–25 Hz) oscillation. The changes in EPSP size are not expressed during non-oscillatory behavior but reappear during subsequent gamma-oscillatory events. We propose that oscillation-induced EPSPs serve as a substrate for memory, whose expression either enhances or blocks synchronization of spatially separated sites. This phenomenon thus provides a dynamical mechanism for storage and retrieval of stimulus-specific neuronal assemblies.

Polycaps: Reversibly formed polymeric capsules

Described are assemblies consisting of polymeric capsules, “polycaps,” formed from two calix[4]arene tetraureas covalently connected at their lower rims. In these structures self-assembly leads to reversibly formed capsule sites along a chain, reminiscent of beads on a string. Their dynamic behavior is characterized by 1H NMR spectroscopy through encapsulation of guest species, reversible polymerization, and the formation of sharply defined hybrid capsules.

A hierarchical neuronal network for planning behavior

Planning a goal-directed sequence of behavior is a higher function of the human brain that relies on the integrity of prefrontal cortical areas. In the Tower of London test, a puzzle in which beads sliding on pegs must be moved to match a designated goal configuration, patients with lesioned prefrontal cortex show deficits in planning a goal-directed sequence of moves. We propose a neuronal network model of sequence planning that passes this test and, when lesioned, fails in a way that mimics prefrontal patients’ behavior. Our model comprises a descending planning system with hierarchically organized plan, operation, and gesture levels, and an ascending evaluative system that analyzes the problem and computes internal reward signals that index the correct/erroneous status of the plan. Multiple parallel pathways connecting the evaluative and planning systems amend the plan and adapt it to the current problem. The model illustrates how specialized hierarchically organized neuronal assemblies may collectively emulate central executive or supervisory functions of the human brain.

Tissue phenotype depends on reciprocal interactions between the extracellular matrix and the structural organization of the nucleus

What determines the nuclear organization within a cell and whether this organization itself can impose cellular function within a tissue remains unknown. To explore the relationship between nuclear organization and tissue architecture and function, we used a model of human mammary epithelial cell acinar morphogenesis. When cultured within a reconstituted basement membrane (rBM), HMT-3522 cells form polarized and growth-arrested tissue-like acini with a central lumen and deposit an endogenous BM. We show that rBM-induced morphogenesis is accompanied by relocalization of the nuclear matrix proteins NuMA, splicing factor SRm160, and cell cycle regulator Rb. These proteins had distinct distribution patterns specific for proliferation, growth arrest, and acini formation, whereas the distribution of the nuclear lamina protein, lamin B, remained unchanged. NuMA relocalized to foci, which coalesced into larger assemblies as morphogenesis progressed. Perturbation of histone acetylation in the acini by trichostatin A treatment altered chromatin structure, disrupted NuMA foci, and induced cell proliferation. Moreover, treatment of transiently permeabilized acini with a NuMA antibody led to the disruption of NuMA foci, alteration of histone acetylation, activation of metalloproteases, and breakdown of the endogenous BM. These results experimentally demonstrate a dynamic interaction between the extracellular matrix, nuclear organization, and tissue phenotype. They further show that rather than passively reflecting changes in gene expression, nuclear organization itself can modulate the cellular and tissue phenotype.

Multiple-complete-digest restriction fragment mapping: Generating sequence-ready maps for large-scale DNA sequencing

Multiple-complete-digest mapping is a DNA mapping technique based on complete-restriction-digest fingerprints of a set of clones that provides highly redundant coverage of the mapping target. The maps assembled from these fingerprints order both the clones and the restriction fragments. Maps are coordinated across three enzymes in the examples presented. Starting with yeast artificial chromosome contigs from the 7q31.3 and 7p14 regions of the human genome, we have produced cosmid-based maps spanning more than one million base pairs. Each yeast artificial chromosome is first subcloned into cosmids at a redundancy of ×15–30. Complete-digest fragments are electrophoresed on agarose gels, poststained, and imaged on a fluorescent scanner. Aberrant clones that are not representative of the underlying genome are rejected in the map construction process. Almost every restriction fragment is ordered, allowing selection of minimal tiling paths with clone-to-clone overlaps of only a few thousand base pairs. These maps demonstrate the practicality of applying the experimental and software-based steps in multiple-complete-digest mapping to a target of significant size and complexity. We present evidence that the maps are sufficiently accurate to validate both the clones selected for sequencing and the sequence assemblies obtained once these clones have been sequenced by a “shotgun” method.

Electrostatic interaction between helical macromolecules in dense aggregates: An impetus for DNA poly- and meso-morphism

DNA exhibits a surprising multiplicity of structures when it is packed into dense aggregates. It undergoes various polymorphous transitions (e.g., from the B to A form) and mesomorphous transformations (from hexagonal to orthorhombic or monoclinic packing, changes in the mutual alignment of nearest neighbors, etc). In this report we show that such phenomena may have their origin in the specific helical symmetry of the charge distribution on DNA surface. Electrostatic interaction between neighboring DNA molecules exhibits strong dependence on the patterns of molecular surface groups and adsorbed counter-ions. As a result, it is affected by such structural parameters as the helical pitch, groove width, the number of base pairs per helical turn, etc. We derive expressions which relate the energy of electrostatic interaction with these parameters and with the packing variables characterizing the axial and azimuthal alignment between neighboring macromolecules. We show, in particular, that the structural changes upon the B-to-A transition reduce the electrostatic energy by ≈kcal/mol per base pair, at a random adsorption of counter ions. Ion binding into the narrow groove weakens or inverts this effect, stabilizing B-DNA, as it is presumably the case in Li+-DNA assemblies. The packing symmetry and molecular alignment in DNA aggregates are shown to be affected by the patterns of ion binding.

Exogenous Administration of Gangliosides Displaces GPI-anchored Proteins from Lipid Microdomains in Living Cells

Exogenous application of gangliosides to cells affects many cellular functions. We asked whether these effects could be attributed to the influence of gangliosides on the properties of sphingolipid–cholesterol microdomains on the plasma membrane, also termed rafts. The latter are envisaged as lateral assemblies of sphingolipids (including gangliosides), cholesterol, and a specific set of proteins. Rafts have been implicated in processes such as membrane trafficking, signal transduction, and cell adhesion. Recently, using a chemical cross-linking approach with Madin-Darby canine kidney (MDCK) cells permanently expressing a GPI-anchored form of growth hormone decay accelerating factor (GH-DAF) as a model system, we could show that GPI-anchored proteins are clustered in rafts in living cells. Moreover, this clustering was dependent on the level of cholesterol in the cell. Here we show that incubation of MDCK cells with gangliosides abolished subsequent chemical cross-linking of GH-DAF. Furthermore, insertion of gangliosides into the plasma membrane of MDCK GH-DAF cells renders GH-DAF soluble when subjected to extraction with Triton X-114 at 4°C. Our data suggest that exogenous application of gangliosides displaces GPI-anchored proteins from sphingolipid–cholesterol microdomains in living cells.

Modulation of Endocytic Traffic in Polarized Madin-Darby Canine Kidney Cells by the Small GTPase RhoA

Efficient postendocytic membrane traffic in polarized epithelial cells is thought to be regulated in part by the actin cytoskeleton. RhoA modulates assemblies of actin in the cell, and it has been shown to regulate pinocytosis and phagocytosis; however, its effects on postendocytic traffic are largely unexplored. To this end, we expressed wild-type RhoA (RhoAWT), dominant active RhoA (RhoAV14), and dominant inactive RhoA (RhoAN19) in Madin-Darby canine kidney (MDCK) cells expressing the polymeric immunoglobulin receptor. RhoAV14 expression stimulated the rate of apical and basolateral endocytosis, whereas RhoAN19 expression decreased the rate from both membrane domains. Polarized basolateral recycling of transferrin was disrupted in RhoAV14-expressing cells as a result of increased ligand release at the apical pole of the cell. Degradation of basolaterally internalized epidermal growth factor was slowed in RhoAV14-expressing cells. Although apical recycling of immunoglobulin A (IgA) was largely unaffected in cells expressing RhoAV14, transcytosis of basolaterally internalized IgA was severely impaired. Morphological and biochemical analyses demonstrated that a large proportion of IgA internalized from the basolateral pole of RhoAV14-expressing cells remained within basolateral early endosomes and was slow to exit these compartments. RhoAN19 and RhoAWT expression had little effect on these postendocytic pathways. These results indicate that in polarized MDCK cells activated RhoA may modulate endocytosis from both membrane domains and postendocytic traffic at the basolateral pole of the cell.