Treatment with the HIV protease inhibitor Nelfinavir triggers the unfolded protein response and may overcome proteasome inhibitor resistance of multiple myeloma in combination with bortezomib: a phase I trial (SAKK 65/08).

Downregulation of the unfolded protein response mediates proteasome inhibitor resistance in Multiple Myeloma.The Human Immunodeficieny Virus protease inhibitor nelfinavir activates the unfolded protein response in vitro. We determined dose limiting toxicity and recommended dose for phase II of nelfinavir in combination with the proteasome inhibitor bortezomib. 12 patients with advanced hematological malignancies were treated with nelfinavir (2500 - 5000 mg/d p.o., d 1-14, 3+3 dose escalation) and bortezomib (1.3 mg/m2, d 1, 4, 8, 11; 21 day cycles). A run in phase with nelfinavir monotherapy allowed pharmakokinetic/pharmakodynamic assessment of nelfinavir in the presence or absence of concomittant bortezomib. Endpoints included dose limiting toxicity, activation of the unfolded protein response, proteasome activity, toxicity and response to trial treatment. Nelfinavir 2 x 2500 mg was the recommended phase II dose identified. Nelfinavir alone significantly upregulated expression of proteins related to the unfolded protein response in peripheral blood mononuclear cells and inhibited proteasome activity. Of 10 evaluable patients in the dose escalation cohort, 3 achieved a partial response, 4 stable disease for ≥ 2 cycles, while 3 had progressive disease as best response. In an exploratory extension cohort with 6 relapsed, bortezomib-refractory, lenalidomide-resistant myeloma patients treated at the recommended phase II dose, 3 reached a partial response, 2 a minor response and one progressive disease. The combination of nelfinavir with bortezomib is safe and shows promising signals for activity in advanced, bortezomib-refractory MM. Induction of the unfolded protein response by nelfinavir may overcome the biological features of proteasome inhibitor resistance. (Trial registration NCT01164709).

Patterns of landscape form in the upper Rhône basin, Central Swiss Alps, predominantly show lithologic controls despite multiple glaciations and variations in rock uplift rates

The development of topography depends mainly on the interplay between uplift and erosion. These processes are controlled by various factors including climate, glaciers, lithology, seismic activity and short-term variables, such as anthropogenic impact. Many studies in orogens all over the world have shown how these controlling variables may affect the landscape's topography. In particular, it has been hypothesized that lithology exerts a dominant control on erosion rates and landscape morphology. However, clear demonstrations of this influence are rare and difficult to disentangle from the overprint of other signals such as climate or tectonics. In this study we focus on the upper Rhône Basin situated in the Central Swiss Alps in order to explore the relation between topography, possible controlling variables and lithology in particular. The Rhône Basin has been affected by spatially variable uplift, high orographically driven rainfalls and multiple glaciations. Furthermore, lithology and erodibility vary substantially within the basin. Thanks to high-resolution geological, climatic and topographic data, the Rhône Basin is a suitable laboratory to explore these complexities. Elevation, relief, slope and hypsometric data as well as river profile information from digital elevation models are used to characterize the landscape's topography of around 50 tributary basins. Additionally, uplift over different timescales, glacial inheritance, precipitation patterns and erodibility of the underlying bedrock are quantified for each basin. Results show that the chosen topographic and controlling variables vary remarkably between different tributary basins. We investigate the link between observed topographic differences and the possible controlling variables through statistical analyses. Variations of elevation, slope and relief seem to be linked to differences in long-term uplift rate, whereas elevation distributions (hypsometry) and river profile shapes may be related to glacial imprint. This confirms that the landscape of the Rhône Basin has been highly preconditioned by (past) uplift and glaciation. Linear discriminant analyses (LDAs), however, suggest a stronger link between observed topographic variations and differences in erodibility. We therefore conclude that despite evident glacial and tectonic conditioning, a lithologic control is still preserved and measurable in the landscape of the Rhône tributary basins.

Assemblages and isotopic signals of modern ostracodes and host waters from Patagonia, Argentine

Ostracode species assemblages and stable oxygen and carbon isotope ratios of living and recent ostracodes, together with delta18O and delta13C_DIC values of host water samples, provide a first data set that characterizes a wide range of modern aquatic environments in the Laguna Cari-Laufquen (41°S, 68 - 69°W) and the Lago Cardiel area (48 - 49°S, 70 - 71°W) in Patagonia, Argentina. This data set will ultimately be used to interpret and calibrate data acquired from lake sediment cores with the goal of reconstructing past climate. Species assemblages and isotope values can be assigned to three groups; (1) springs, seeps and streams, (2) permanent ponds and lakes, and (3) ephemeral ponds and lakes. Springs, seeps and streams are characterized by Darwinula sp., Heterocypris incongruens, Eucypris fontana, Amphicypris nobilis and Ilyocypris ramirezi. Ostracode and water isotope values range between -13 and -5 per mil for oxygen, and between -15 and -3 per mil for carbon. They are the most negative of the entire sample set, reflecting ground water input with little or no evaporative enrichment. Limnocythere patagonica, Eucypris labyrinthica, Limnocythere sp. and Eucypris aff. fontana are typical species of permanent ponds and lakes. Isotope values indicate high degree of evaporation of lake waters relative to feeder springs and streams and range between -7 and +5 per mil for oxygen, and -5 and +4 per mil for carbon. Limnocythere rionegroensis is the dominant species in ephemeral ponds and lakes. These systems display the most enriched isotope values in both ostracodes and host waters, extending from -5 to +7 per mil for oxygen, and from -5 to +6 per mil for carbon. Living ostracodes show a positive offset from equilibrium values of up to 2 per mil for oxygen. Carbon-isotope values are up to 6? more negative than equilibrium values in highly productive pools. Comparison of ostracode and host water isotope signals permits assessment of the life span of the aquatic environments. Valves from dead ostracodes collected from ephemeral ponds and lakes show a wide scatter with each sample providing a snapshot of the seasonal history of the host water. The presence of the stream species Ilyocypris ramirezi and a wide range of ostracode isotope values suggest that ephemeral ponds and lakes are fed by streams during spring run-off and seasonally dry. A temporary character is also indicated by Heterocypris incongruens, a drought-resistant species that occupies most springs and seeps. In addition, Limnocythere rionegroensis has adjusted its reproduction strategies to its environment. Whereas only females were collected in fresh host waters, males were found in ephemeral ponds and lakes with higher solute content. Sexual reproduction seems to be the more successful reproduction strategy in high and variable salinities and seasonal droughts. The temporary character of the aquatic environments shows that the availability of meteoric water controls the life span of host waters and underlines the sensitivity of the area to changes in precipitation.

Signals of resilience to ocean change: high thermal tolerance of early stage Antarctic sea urchins (Sterechinus neumayeri) reared under present-day and future pCO2 and temperature

We tested the hypothesis that development of the Antarctic urchin Sterechinus neumayeri under future ocean conditions of warming and acidification would incur physiological costs, reducing the tolerance of a secondary stressor. The aim of this study is twofold: (1) quantify current austral spring temperature and pH near sea urchin habitat at Cape Evans in McMurdo Sound, Antarctica and (2) spawn S. neumayeri in the laboratory and raise early developmental stages (EDSs) under ambient (-0.7 °C; 400 µatm pCO2) and future (+2.6 °C; 650 and 1,000 µatm pCO2) ocean conditions and expose four EDSs (blastula, gastrula, prism, and 4-arm echinopluteus) to a one hour acute heat stress and assess survivorship. Results of field data from 2011 to 2012 show extremely stable inter-annual pH conditions ranging from 7.99 to 8.08, suggesting that future ocean acidification will drastically alter the pH-seascape for S. neumayeri. In the laboratory, S. neumayeri EDSs appear to be tolerant of temperatures and pCO2 levels above their current habitat conditions. EDSs survived acute heat exposures >20 °C above habitat temperatures of -1.9 °C. No pCO2 effect was observed for EDSs reared at -0.7 °C. When reared at +2.6 °C, small but significant pCO2 effects were observed at the blastula and prism stage, suggesting that multiple stressors are more detrimental than single stressors. While surprisingly tolerant overall, blastulae were the most sensitive stage to ocean warming and acidification. We conclude that S. neumayeri may be unexpectedly physiologically tolerant of future ocean conditions.

Large aperture antenna arrays with subband delay compensation for reception of wideband satellite signals

Con esta disertación se pretenden resolver algunos de los problemas encontrados actualmente en la recepción de señales de satélites bajo dos escenarios particularmente exigentes: comunicaciones de Espacio Profundo y en banda Ka. Las comunicaciones con sondas de Espacio Profundo necesitan grandes aperturas en tierra para poder incrementar la velocidad de datos. La opción de usar antennas con diámetro mayor de 35 metros tiene serios problemas, pues antenas tan grandes son caras de mantener, difíciles de apuntar, pueden tener largos tiempo de reparación y además tienen una efeciencia decreciente a medida que se utilizan bandas más altas. Soluciones basadas en agrupaciones de antenas de menor tamaño (12 ó 35 metros) son mas ecónomicas y factibles técnicamente. Las comunicaciones en banda Ka tambien pueden beneficiarse de la combinación de múltiples antennas. Las antenas de menor tamaño son más fáciles de apuntar y además tienen un campo de visión mayor. Además, las técnicas de diversidad espacial pueden ser reemplazadas por una combinación de antenas para así incrementar el margen del enlace. La combinación de antenas muy alejadas sobre grandes anchos de banda, bien por recibir una señal de banda ancha o múltiples de banda estrecha, es complicada técnicamente. En esta disertación se demostrará que el uso de conformador de haz en el dominio de la frecuencia puede ayudar a relajar los requisitos de calibración y, al mismo tiempo, proporcionar un mayor campo de visión y mayores capacidades de ecualización. Para llevar esto a cabo, el trabajo ha girado en torno a tres aspectos fundamentales. El primero es la investigación bibliográfica del trabajo existente en este campo. El segundo es el modelado matemático del proceso de combinación y el desarrollo de nuevos algoritmos de estimación de fase y retardo. Y el tercero es la propuesta de nuevas aplicaciones en las que usar estas técnicas. La investigación bibliográfica se centra principalmente en los capítulos 1, 2, 4 y 5. El capítulo 1 da una breve introducción a la teoría de combinación de antenas de gran apertura. En este capítulo, los principales campos de aplicación son descritos y además se establece la necesidad de compensar retardos en subbandas. La teoría de bancos de filtros se expone en el capítulo 2; se selecciona y simula un banco de filtros modulado uniformemente con fase lineal. Las propiedades de convergencia de varios filtros adaptativos se muestran en el capítulo 4. Y finalmente, las técnicas de estimación de retardo son estudiadas y resumidas en el capítulo 5. Desde el punto de vista matemático, las principales contribución de esta disertación han sido: • Sección 3.1.4. Cálculo de la desviación de haz de un conformador de haz con compensación de retardo en pasos discretos en frecuencia intermedia. • Sección 3.2. Modelo matemático de un conformador de haz en subbandas. • Sección 3.2.2. Cálculo de la desviación de haz de un conformador de haz en subbandas con un buffer de retardo grueso. • Sección 3.2.4. Análisis de la influencia de los alias internos en la compensación en subbandas de retardo y fase. • Sección 3.2.4.2. Cálculo de la desviación de haz de un conformador de haz con compensación de retardo en subbandas. • Sección 3.2.6. Cálculo de la ganancia de relación señal a ruido de la agrupación de antenas en cada una de las subbandas. • Sección 3.3.2. Modelado de la función de transferencia de la agrupación de antenas bajo errores de estimación de retardo. • Sección 3.3.3. Modelado de los efectos de derivas de fase y retardo entre actualizaciones de las estimaciones. • Sección 3.4. Cálculo de la directividad de la agrupación de antenas con y sin compensación de retardos en subbandas. • Sección 5.2.6. Desarrollo de un algorimo para estimar la fase y el retardo entre dos señales a partir de su descomposición de subbandas bajo entornos estacionarios. • Sección 5.5.1. Desarrollo de un algorimo para estimar la fase, el retardo y la deriva de retardo entre dos señales a partir de su descomposición de subbandas bajo entornos no estacionarios. Las aplicaciones que se pueden beneficiar de estas técnicas son descritas en el capítulo 7: • Sección 6.2. Agrupaciones de antenas para comunicaciones de Espacio Profundo con capacidad multihaz y sin requisitos de calibración geométrica o de retardo de grupo. • Sección 6.2.6. Combinación en banda ancha de antenas con separaciones de miles de kilómetros, para recepción de sondas de espacio profundo. • Secciones 6.4 and 6.3. Combinación de estaciones remotas en banda Ka en escenarios de diversidad espacial, para recepción de satélites LEO o GEO. • Sección 6.3. Recepción de satélites GEO colocados con arrays de antenas multihaz. Las publicaciones a las que ha dado lugar esta tesis son las siguientes • A. Torre. Wideband antenna arraying over long distances. Interplanetary Progress Report, 42-194:1–18, 2013. En esta pulicación se resumen los resultados de las secciones 3.2, 3.2.2, 3.3.2, los algoritmos en las secciones 5.2.6, 5.5.1 y la aplicación destacada en 6.2.6. • A. Torre. Reception of wideband signals from geostationary collocated satellites with antenna arrays. IET Communications, Vol. 8, Issue 13:2229–2237, September, 2014. En esta segunda se muestran los resultados de la sección 3.2.4, el algoritmo en la sección 5.2.6.1 , y la aplicación mostrada en 6.3. ABSTRACT This dissertation is an attempt to solve some of the problems found nowadays in the reception of satellite signals under two particular challenging scenarios: Deep Space and Ka-band communications. Deep Space communications require from larger apertures on ground in order to increase the data rate. The option of using single dishes with diameters larger than 35 meters has severe drawbacks. Such antennas are expensive to maintain, prone to long downtimes, difficult to point and have a degraded performance in high frequency bands. The array solution, either with 12 meter or 35 meter antennas is deemed to be the most economically and technically feasible solution. Ka-band communications can also benefit from antenna arraying technology. The smaller aperture antennas that make up the array are easier to point and have a wider field of view allowing multiple simultaneous beams. Besides, site diversity techniques can be replaced by pure combination in order to increase link margin. Combination of far away antennas over a large bandwidth, either because a wideband signal or multiple narrowband signals are received, is a demanding task. This dissertation will show that the use of frequency domain beamformers with subband delay compensation can help to ease calibration requirements and, at the same time, provide with a wider field of view and enhanced equalization capabilities. In order to do so, the work has been focused on three main aspects. The first one is the bibliographic research of previous work on this subject. The second one is the mathematical modeling of the array combination process and the development of new phase/delay estimation algorithms. And the third one is the proposal of new applications in which these techniques can be used. Bibliographic research is mainly done in chapters 1, 2, 4 and 5. Chapter 1 gives a brief introduction to previous work in the field of large aperture antenna arraying. In this chapter, the main fields of application are described and the need for subband delay compensation is established. Filter bank theory is shown in chapter 2; a linear phase uniform modulated filter bank is selected and simulated under diverse conditions. The convergence properties of several adaptive filters are shown in chapter 4. Finally, delay estimation techniques are studied and summarized in chapter 5. From a mathematical point of view, the main contributions of this dissertation have been: • Section 3.1.4. Calculation of beam squint of an IF beamformer with delay compensation at discrete time steps. • Section 3.2. Establishment of a mathematical model of a subband beamformer. • Section 3.2.2. Calculation of beam squint in a subband beamformer with a coarse delay buffer. • Section 3.2.4. Analysis of the influence of internal aliasing on phase and delay subband compensation. • Section 3.2.4.2. Calculation of beam squint of a beamformer with subband delay compensation. • Section 3.2.6. Calculation of the array SNR gain at each of the subbands. • Section 3.3.2. Modeling of the transfer function of an array subject to delay estimation errors. • Section 3.3.3. Modeling of the effects of phase and delay drifts between estimation updates. • Section 3.4. Calculation of array directivity with and without subband delay compensation. • Section 5.2.6. Development of an algorithm to estimate relative delay and phase between two signals from their subband decomposition in stationary environments. • Section 5.5.1. Development of an algorithm to estimate relative delay rate, delay and phase between two signals from their subband decomposition in non stationary environments. The applications that can benefit from these techniques are described in chapter 7: • Section 6.2. Arrays of antennas for Deep Space communications with multibeam capacity and without geometric or group delay calibration requirement. • Section 6.2.6. Wideband antenna arraying over long distances, in the range of thousands of kilometers, for reception of Deep Space probes. • Sections 6.4 y 6.3. Combination of remote stations in Ka-band site diversity scenarios for reception of LEO or GEO satellites. • Section 6.3. Reception of GEO collocated satellites with multibeam antenna arrays. The publications that have been made from the work in this dissertation are • A. Torre. Wideband antenna arraying over long distances. Interplanetary Progress Report, 42-194:1–18, 2013. This article shows the results in sections 3.2, 3.2.2, 3.3.2, the algorithms in sections 5.2.6, 5.5.1 and the application in section 6.2.6. • A. Torre. Reception of wideband signals from geostationary collocated satellites with antenna arrays. IET Communications, Vol. 8, Issue 13:2229–2237, September, 2014. This second article shows among others the results in section 3.2.4, the algorithm in section 5.2.6.1 , and the application in section 6.3.

In silico detection of control signals: mRNA 3′-end-processing sequences in diverse species

We have investigated mRNA 3′-end-processing signals in each of six eukaryotic species (yeast, rice, arabidopsis, fruitfly, mouse, and human) through the analysis of more than 20,000 3′-expressed sequence tags. The use and conservation of the canonical AAUAAA element vary widely among the six species and are especially weak in plants and yeast. Even in the animal species, the AAUAAA signal does not appear to be as universal as indicated by previous studies. The abundance of single-base variants of AAUAAA correlates with their measured processing efficiencies. As found previously, the plant polyadenylation signals are more similar to those of yeast than to those of animals, with both common content and arrangement of the signal elements. In all species examined, the complete polyadenylation signal appears to consist of an aggregate of multiple elements. In light of these and previous results, we present a broadened concept of 3′-end-processing signals in which no single exact sequence element is universally required for processing. Rather, the total efficiency is a function of all elements and, importantly, an inefficient word in one element can be compensated for by strong words in other elements. These complex patterns indicate that effective tools to identify 3′-end-processing signals will require more than consensus sequence identification.

Membrane-anchored Plakoglobins Have Multiple Mechanisms of Action in Wnt Signaling

In Wnt signaling, β-catenin and plakoglobin transduce signals to the nucleus through interactions with TCF-type transcription factors. However, when plakoglobin is artificially engineered to restrict it to the cytoplasm by fusion with the transmembrane domain of connexin (cnxPg), it efficiently induces a Wnt-like axis duplication phenotype in Xenopus. In Xenopus embryos, maternal XTCF3 normally represses ventral expression of the dorsalizing gene Siamois. Two models have been proposed to explain the Wnt-like activity of cnxPg: 1) that cnxPg inhibits the machinery involved in the turnover of cytosolic β-catenin, which then accumulates and inhibits maternal XTCF3, and 2) that cnxPg directly acts to inhibit XTCF3 activity. To distinguish between these models, we created a series of N-terminal deletion mutations of cnxPg and examined their ability to induce an ectopic axis in Xenopus, activate a TCF-responsive reporter (OT), stabilize β-catenin, and colocalize with components of the Wnt signaling pathway. cnxPg does not colocalize with the Wnt pathway component Dishevelled, but it does lead to the redistribution of APC and Axin, two proteins involved in the regulation of β-catenin turnover. Expression of cnxPg increases levels of cytosolic β-catenin; however, this effect does not completely explain its signaling activity. Although cnxPg and Wnt-1 stabilize β-catenin to similar extents, cnxPg activates OT to 10- to 20-fold higher levels than Wnt-1. Moreover, although LEF1 and TCF4 synergize with β-catenin and plakoglobin to activate OT, both suppress the signaling activity of cnxPg. In contrast, XTCF3 suppresses the signaling activity of both β-catenin and cnxPg. Both exogenous XLEF1 and XTCF3 are sequestered in the cytoplasm of Xenopus cells by cnxPg. Based on these data, we conclude that, in addition to its effects on β-catenin, cnxPg interacts with other components of the Wnt pathway, perhaps TCFs, and that these interactions contribute to its signaling activity.

The adaptor protein Crk connects multiple cellular stimuli to the JNK signaling pathway

c-Jun N-terminal kinases (JNKs) are potently activated by a number of cellular stimuli. Small GTPases, in particular Rac, are responsible for initiating the activation of the JNK pathways. So far, the signals leading from extracellular stimuli to the activation of Rac have remained elusive. Recent studies have demonstrated that the Src homology 2 (SH2)- and Src homology 3 (SH3)-containing adaptor protein Crk is capable of activating JNK when ectopically expressed. We found here that transient expression of Crk induces JNK activation, and this activation was dependent on both the SH2- and SH3-domains of Crk. Expression of p130Cas (Cas), a major binding protein for the Crk SH2-domain, also induced JNK activation, which was blocked by the SH2-mutant of Crk. JNK activation by Cas and Crk was effectively blocked by a dominant-negative form of Rac, suggesting for a linear pathway from the Cas-Crk-complex to the Rac-JNK activation. Many of the stimuli that activate the Rac-JNK pathway enhance engagement of the Crk SH2-domain. JNK activation by these stimuli, such as epidermal growth factor, integrin ligand binding and v-Src, was efficiently blocked by dominant-negative mutants of Crk. A dominant-negative form of Cas in turn blocked the integrin-, but not epidermal growth factor - nor v-Src-mediated JNK activation. Together, these results demonstrate an important role for Crk in connecting multiple cellular stimuli to the Rac-JNK pathway, and a role for the Cas-Crk complex in integrin-mediated JNK activation.

Altered thymic positive selection and intracellular signals in Cbl-deficient mice

Cbl is the product of the protooncogene c-cbl and is involved in T cell antigen receptor (TCR)-mediated signaling. To understand the role of Cbl for immune system development and function, we generated a Cbl-deficient mouse strain. In Cbl-deficient mice, positive selection of the thymocytes expressing major histocompatibility complex class II-restricted transgenic TCR was significantly enhanced. Two factors may have contributed to the altered thymic selection. First, Cbl deficiency markedly up-regulated the activity of ZAP-70 and mitogen-activated protein kinases. The mitogen-activated protein kinase pathway was shown previously to be involved in thymic positive selection. Second, Cbl-deficient thymocytes expressed CD3 and CD4 molecules at higher levels, which consequently may increase the avidity of TCR/major histocompatibility complex/coreceptor interaction. Thus, Cbl plays a novel role in modulating TCR-mediated multiple signaling pathways and fine-tunes the signaling threshold for thymic selection.

Refractoriness to melatonin occurs independently at multiple brain sites in Siberian hamsters

The mid-winter development of refractoriness to melatonin (Mel) triggers recrudescence of the atrophied reproductive apparatus of rodents. As a consequence, over-wintering animals become reproductively competent just before the onset of spring conditions favorable for breeding. The neural target tissues that cease to respond to winter Mel signals have not been identified. We now report that the suprachiasmatic nucleus of the hypothalamus, which contains the principal circadian clock, and the reuniens and paraventricular nuclei of the thalamus, each independently becomes refractory to melatonin. Small implants of Mel that were left in place for 40 wk and that act locally on these brain nuclei, induced testicular regression within 6 wk in male Siberian hamsters; 12 wk later Mel implants no longer suppressed reproduction and gonadal recrudescence ensued. Hamsters that were then given a systemic Mel infusion s.c. immediately initiated a second gonadal regression, implying that neurons at each site become refractory to Mel without compromising responsiveness of other Mel target tissues. Refractoriness occurs locally and independently at each neural target tissue, rather than in a separate “refractoriness” substrate. Restricted, target-specific actions of Mel are consistent with the independent regulation by day length of the several behavioral and physiological traits that vary seasonally in mammals.

Signals and signs in the nervous system: The dynamic anatomy of electrical activity is probably information-rich

The dichotomy between two groups of workers on neuroelectrical activity is retarding progress. To study the interrelations between neuronal unit spike activity and compound field potentials of cell populations is both unfashionable and technically challenging. Neither of the mutual disparagements is justified: that spikes are to higher functions as the alphabet is to Shakespeare and that slow field potentials are irrelevant epiphenomena. Spikes are not the basis of the neural code but of multiple codes that coexist with nonspike codes. Field potentials are mainly information-rich signs of underlying processes, but sometimes they are also signals for neighboring cells, that is, they exert influence. This paper concerns opportunities for new research with many channels of wide-band (spike and slow wave) recording. A wealth of structure in time and three-dimensional space is different at each scale—micro-, meso-, and macroactivity. The depth of our ignorance is emphasized to underline the opportunities for uncovering new principles. We cannot currently estimate the relative importance of spikes and synaptic communication vs. extrasynaptic graded signals. In spite of a preponderance of literature on the former, we must consider the latter as probably important. We are in a primitive stage of looking at the time series of wide-band voltages in the compound, local field, potentials and of choosing descriptors that discriminate appropriately among brain loci, states (functions), stages (ontogeny, senescence), and taxa (evolution). This is not surprising, since the brains in higher species are surely the most complex systems known. They must be the greatest reservoir of new discoveries in nature. The complexity should not deter us, but a dose of humility can stimulate the flow of imaginative juices.

Sonic hedgehog protein signals not as a hydrolytic enzyme but as an apparent ligand for Patched

The amino-terminal signaling domain of the Sonic hedgehog secreted protein (Shh-N), which derives from the Shh precursor through an autoprocessing reaction mediated by the carboxyl-terminal domain, executes multiple functions in embryonic tissue patterning, including induction of ventral and suppression of dorsal cell types in the developing neural tube. An apparent catalytic site within Shh-N is suggested by structural homology to a bacterial carboxypeptidase. We demonstrate here that alteration of residues presumed to be critical for a hydrolytic activity does not cause a loss of inductive activity, thus ruling out catalysis by Shh-N as a requirement for signaling. We favor the alternative, that Shh-N functions primarily as a ligand for the putative receptor Patched (Ptc). This possibility is supported by new evidence for direct binding of Shh-N to Ptc and by a strong correlation between the affinity of Ptc-binding and the signaling potency of Shh-N protein variants carrying alterations of conserved residues in a particular region of the protein surface. These results together suggest that direct Shh-N binding to Ptc is a critical event in transduction of the Shh-N signal.

Expression of an Arabidopsis Aspartate Kinase/Homoserine Dehydrogenase Gene Is Metabolically Regulated by Photosynthesis-Related Signals but Not by Nitrogenous Compounds1

Although the control of carbon fixation and nitrogen assimilation has been studied in detail, relatively little is known about the regulation of carbon and nitrogen flow into amino acids. In this paper we report our study of the metabolic regulation of expression of an Arabidopsis aspartate kinase/homoserine dehydrogenase (AK/HSD) gene, which encodes two linked key enzymes in the biosynthetic pathway of aspartate family amino acids. Northern blot analyses, as well as expression of chimeric AK/HSD-β-glucuronidase constructs, have shown that the expression of this gene is regulated by the photosynthesis-related metabolites sucrose and phosphate but not by nitrogenous compounds. In addition, analysis of AK/HSD promoter deletions suggested that a CTTGACTCTA sequence, resembling the binding site for the yeast GCN4 transcription factor, is likely to play a functional role in the expression of this gene. Nevertheless, longer promoter fragments, lacking the GCN4-like element, were still able to confer sugar inducibility, implying that the metabolic regulation of this gene is apparently obtained by multiple and redundant promoter sequences. The present and previous studies suggest that the conversion of aspartate into either the storage amino acid asparagine or aspartate family amino acids is subject to a coordinated, reciprocal metabolic control, and this biochemical branch point is a part of a larger, coordinated regulatory mechanism of nitrogen and carbon storage and utilization.

Three 1-Aminocyclopropane-1-Carboxylate Synthase Genes Regulated by Primary and Secondary Pollination Signals in Orchid Flowers1

The temporal and spatial expression patterns of three 1-aminocyclopropane-1-carboxylate (ACC) synthase genes were investigated in pollinated orchid (Phalaenopsis spp.) flowers. Pollination signals initiate a cascade of development events in multiple floral organs, including the induction of ethylene biosynthesis, which coordinates several postpollination developmental responses. The initiation and propagation of ethylene biosynthesis is regulated by the coordinated expression of three distinct ACC synthase genes in orchid flowers. One ACC synthase gene (Phal-ACS1) is regulated by ethylene and participates in amplification and interorgan transmission of the pollination signal, as we have previously described in a related orchid genus. Two additional ACC synthase genes (Phal-ACS2 and Phal-ACS3) are expressed primarily in the stigma and ovary of pollinated orchid flowers. Phal-ACS2 mRNA accumulated in the stigma within 1 h after pollination, whereas Phal-ACS1 mRNA was not detected until 6 h after pollination. Similar to the expression of Phal-ACS2, the Phal-ACS3 gene was expressed within 2 h after pollination in the ovary. Exogenous application of auxin, but not ACC, mimicked pollination by stimulating a rapid increase in ACC synthase activity in the stigma and ovary and inducing Phal-ACS2 and Phal-ACS3 mRNA accumulation in the stigma and ovary, respectively. These results provide the basis for an expanded model of interorgan regulation of three ACC synthase genes that respond to both primary (Phal-ACS2 and Phal-ACS3) and secondary (Phal-ACS1) pollination signals.

A potential pathogenetic mechanism for multiple endocrine neoplasia type 2 syndromes involves ret-induced impairment of terminal differentiation of neuroepithelial cells.

Germ-line missense mutations of the receptor-like tyrosine kinase ret are the causative genetic event of the multiple endocrine neoplasia (MEN) type 2A and type 2B syndromes and of the familial medullary thyroid carcinoma. We have used the rat pheochromocytoma cell line, PC12, as a model system to investigate the mechanism or mechanisms by which expression of activated ret alleles contributes to the neoplastic phenotype in neuroendocrine cells. Here we show that stable expression of ret mutants (MEN2A and MEN2B alleles) in PC12 cells causes a dramatic conversion from a round to a flat morphology, accompanied by the induction of genes belonging to the early as well as the delayed response to nerve growth factor. However, in the transfected PC12 cells, the continuous expression of neuronal specific genes is not associated with the suppression of cell proliferation. Furthermore, expression of ret mutants renders PC12 cells unresponsive to nerve growth factor-induced inhibition of proliferation. These results suggest that induction of an aberrant pattern of differentiation, accompanied by unresponsiveness to growth-inhibitory physiological signals, may be part of the mechanism of action of activated ret alleles in the pathogenesis of neuroendocrine tumors associated with MEN2 syndromes.