The structure of the acto-myosin subfragment 1 complex: Results of searches using data from electron microscopy and x-ray crystallography

Surmises of how myosin subfragment 1 (S1) interacts with actin filaments in muscle contraction rest upon knowing the relative arrangement of the two proteins. Although there exist crystallographic structures for both S1 and actin, as well as electron microscopy data for the acto–S1 complex (AS1), modeling of this arrangement has so far only been done “by eye.” Here we report fitted AS1 structures obtained using a quantitative method that is both more objective and makes more complete use of the data. Using undistorted crystallographic results, the best-fit AS1 structure shows significant differences from that obtained by visual fitting. The best fit is produced using the F-actin model of Holmes et al. [Holmes, K. C., Popp, D., Gebhard, W. & Kabsch, W. (1990) Nature (London) 347, 44–49]. S1 residues at the AS1 interface are now found at a higher radius as well as being translated axially and rotated azimuthally. Fits using S1 plus loops missing from the crystal structure were achieved using a homology search method to predict loop structures. These improved fits favor an arrangement in which the loop at the 50- to 20-kDa domain junction of S1 is located near the N terminus of actin. Rigid-body movements of the lower 50-kDa domain, which further improve the fit, produce closure of the large 50-kDa domain cleft and bring conserved residues in the lower 50-kDa domain into an apparently appropriate orientation for close interaction with actin. This finding supports the idea that binding of ATP to AS1 at the end of the ATPase cycle disrupts the actin binding site by changing the conformation of the 50-kDa cleft of S1.

Bicarbonate is an essential constituent of the water-oxidizing complex of photosystem II

It is shown that restoration of photoinduced electron flow and O2 evolution with Mn2+ in Mn-depleted photosystem II (PSII) membrane fragments isolated from spinach chloroplasts is considerably increased with bicarbonate in the region pH 5.0–8.0 in bicarbonate-depleted medium. In buffered solutions equilibrated with the atmosphere (nondepleted of bicarbonate), the bicarbonate effect is observed only at pH lower than the pK of H2CO3 dissociation (6.4), which indicates that HCO3− is the essential species for the restoration effect. The addition of just 2 Mn2+ atoms per one PSII reaction center is enough for the maximal reactivation when bicarbonate is present in the medium. Analysis of bicarbonate concentration dependence of the restoration effect reveals two binding sites for bicarbonate with apparent dissociation constant (Kd) of ≈2.5 μM and 20–34 μM when 2,6-dichloro-p-benzoquinone is used as electron acceptor, while in the presence of silicomolybdate only the latter one remains. Similar bicarbonate concentration dependence of O2 evolution was obtained in untreated Mn-containing PSII membrane fragments. It is suggested that the Kd of 20–34 μM is associated with the donor side of PSII while the location of the lower Kd binding site is not quite clear. The conclusion is made that bicarbonate is an essential constituent of the water-oxidizing complex of PSII, important for its assembly and maintenance in the functionally active state.

The neuroprotective effect of pituitary adenylate cyclase-activating polypeptide on cerebellar granule cells is mediated through inhibition of the CED3-related cysteine protease caspase-3/CPP32

Caspase-3 knockout mice exhibit thickening of the internal granule cell layer of the cerebellum. Concurrently, it has been shown that intracerebral injection of pituitary adenylate cyclase-activating polypeptide (PACAP) induces a transient increase of the thickness of the cerebellar cortex. In the present study, we have investigated the possible effect of PACAP on caspase activity in cultured cerebellar granule cells from 8-day-old rat. Incubation of granule neurons with PACAP for 24 h promoted cell survival and prevented DNA fragmentation. Exposure of cerebellar granule cells to the specific caspase-3 inhibitor N-benzyloxycarbonyl-Asp-Glu-Val-Asp fluoromethylketone (Z-DEVD-FMK) for 24 h markedly enhanced cell survival and inhibited apoptotic cell death. Time-course studies revealed that PACAP causes a prolonged inhibition of caspase-3 activity without affecting caspase-1. Administration of graded concentrations of PACAP for 3 h induced a dose-dependent inhibition of caspase-3 activity. Incubation of granule cells with both dibutyryl-cAMP (dbcAMP) and phorbol 12-myristate 13-acetate (PMA) mimicked the inhibitory effect of PACAP on caspase-3. Cotreatment of cultured neurons with the protein kinase A inhibitor H89 and the protein kinase C inhibitor chelerythrine abrogated the effect of PACAP on caspase-3 activity. In contrast, the ERK kinase inhibitor U0126 did not affect the action of PACAP on caspase-3 activity. These data demonstrate that PACAP prevents cerebellar granule neurons from apoptotic cell death through a protein kinase A- and protein kinase C-dependent inhibition of caspase-3 activity.

Postembryonic segregation of the germ line in sea urchins in relation to indirect development.

The four small micromeres of the sea urchin embryo contribute only to the coelomic sacs, which produce major components of the adult body plan during postembryonic development. To test the proposition that the small micromeres are the definitive primordial germ cell lineage of the sea urchin, we deleted their 4th cleavage parents, and raised the deleted embryos through larval life and metamorphosis to sexual maturity. Almost all of the experimental animals produced functional gametes, excluding the possibility that the germ cell lineage arises exclusively and obligatorily from descendants of the small micromeres; rather, the germ cell lineage arises during the postembryonic development of the rudiment. A survey of the literature indicates that there is no known case of an embryonic primordial germ cell lineage in a bilaterian species that displays maximal indirect development.

Association of the "major histocompatibility complex subregion" I-J determinant with bioactive glycosylation-inhibiting factor.

Murine suppressor T-cell hybridoma cells (231F1) secrete not only bioactive glycosylation-inhibiting factor (GIF) but also an inactive peptide comparable to bioactive GIF peptide in its molecular size and reactivity with anti-GIF; the amino acid sequence of the inactive peptide is identical to that of the bioactive homologue. The inactive GIF peptide in culture supernatant of both the 231F1 cells and a stable transfectant of human GIF cDNA in the murine suppressor T hybridoma selectively bound to Affi-Gel 10, whereas bioactive GIF peptides from the same sources failed to bind to the gel. The inactive cytosolic human GIF from the stable transfectant and Escherichia coli-derived recombinant human GIF also had affinity for Affi-Gel 10. Both the bioactive murine GIF peptide from the suppressor T hybridoma and bioactive recombinant human GIF from the stable transfectant bound to the anti-I-J monoclonal antibody H6 coupled to Affi-Gel. However, bioactive hGIF produced by a stable transfectant of human GIF cDNA in BMT10 cells failed to be retained in H6-coupled Affi-Gel. These results indicate that the I-J specificity is determined by the cell source of the GIF peptide and that the I-J determinant recognized by monoclonal antibody H6 does not represent a part of the primary amino acid sequence of GIF. It appears that the epitope is generated by a posttranslational modification of the peptide.

Critical role of the interleukin 2 (IL-2) receptor gamma-chain-associated Jak3 in the IL-2-induced c-fos and c-myc, but not bcl-2, gene induction.

The interleukin 2 receptor (IL-2R) consists of three subunits, the IL-2R alpha, IL-2R beta c, and IL-2R gamma c chains. Two Janus family protein tyrosine kinases (PTKs), Jak1 and Jak3, were shown to associate with IL-2R beta c and IL-2R gamma c, respectively, and their PTK activities are increased after IL-2 stimulation. A Jak3 mutant with truncation of the C-terminal PTK domain lacks its intrinsic kinase activity but can still associate with IL-2R gamma c. In a hematopoietic cell line, F7, that responds to either IL-2 or IL-3, overexpression of this Jak3 mutant results in selective inhibition of the IL-2-induced activation of Jak1/Jak3 PTKs and of cell proliferation. Of the three target nuclear protooncogenes of the IL-2 signaling, c-fos and c-myc genes, but not the bcl-2 gene, were found to be impaired. On the other hand, overexpression of the dominant negative form of the IL-2R gamma c chain, which lacks most of its cytoplasmic domain, in F7 cells resulted in the inhibition of all three protooncogenes. These results provide a further molecular basis for the critical role of Jak3 in IL-2 signaling and also suggest a Jak PTK-independent signaling pathway(s) for the bcl-2 gene induction by IL-2R.

Estudo retrospectivo das afecções orais em 754 felinos domésticos (Felis catus) atendidos no Laboratório de Odontologia Comparada da Universidade de São Paulo

Objetivou-se fazer um estudo retrospectivo avaliando quais as afecções da cavidade oral foram mais frequentes nos gatos domésticos atendidos no Laboratório de Odontologia Comparada da Faculdade de Medicina Veterinária e Zootecnia da Universidade de São Paulo, relatando estatisticamente a prevalência das afecções da cavidade oral de gatos, enfatizando se há correlação entre elas e com características como raça, sexo, faixa etária e estado reprodutivo. Os dados analisados dos 754 prontuários foram raça, idade, sexo, estado reprodutivo, diagnóstico, tratamento e, no caso de neoplasia, sua localização e diagnóstico histopatológico. As principais doenças diagnosticadas foram doença periodontal, fratura dentária, gengivoestomatite crônica felina, lesão de reabsorção dentária felina, neoplasia oral e traumatismo do sistema estomatognático (luxação de articulação temporomandibular, fenda palatina, fratura de processo coronoide, fratura de zigomático, disjunção de sínfise, fratura de maxila e mandíbula). A idade dos animais variou de menos de um ano a 20 anos, sendo que, os animais tinham, em média 7,2 anos (desvio padrão = 4,9) e a faixa etária mais frequente foi de um a cinco anos. Os gatos sem raça definida (66,5%), siameses (19,0%) e persas (10,2%) totalizaram 95,7% de todos os felinos atendidos no LOC. A doença periodontal foi a afecção mais frequente e esteve presente em 38,3% da população estudada. A fratura dentária, segunda mais frequente, esteve presente em 27,2% dos animais. Houve associação estatisticamente significativa (p=0,026) entre fratura dentária e faixa etária, já que a proporção de animais entre um e cinco anos de idade com fratura foi maior do que a das outras faixas etárias. A lesão de reabsorção dentária felina (LRDF) esteve presente em 19,6% dos gatos estudados, sendo a terceira afecção mais prevalente dentre as pesquisadas. Esta lesão foi mais frequente em gatos com idade entre 11 e 15 anos e houve associação estatisticamente significativa entre a LRDF e a doença periodontal e entre LRDF e gengivite. A prevalência de gengivoestomatite crônica felina foi de 15,7% entre os felinos pesquisados e a proporção de animais com idades entre seis e dez anos com esta doença foi maior do que em outras faixas etárias. As neoplasias estavam presentes em 9,8% dos gatos, sendo que em 46 dos 72 animais que apresentaram alguma neoplasia tinham mais de dez anos de idade. O carcinoma de células escamosas foi o neoplasma mais comum, correspondendo a 63,2% das neoformações que foram submetidas ao exame histopatológico. As fraturas ósseas do sistema estomatognático corresponderam a 19,3% dos atendimentos, sendo a sínfise mentoniana e o corpo da mandíbula os locais mais comuns de fraturas. Concluiu-se que: existe grande variedade de afecções que acometem a cavidade oral de gatos, sendo a doença periodontal, fratura dentária, lesão de reabsorção dentária, gengivite, gengivoestomatite crônica, neoplasias orais e fraturas dos ossos do sistema estomatognático as mais prevalentes delas; é de extrema importância que as anotações nas fichas de atendimento sejam feitas da maneira mais completa possível, para que informações não sejam perdidas

Critical properties of the four-state commutative random permutation glassy Potts model in three and four dimensions

We investigate the critical properties of the four-state commutative random permutation glassy Potts model in three and four dimensions by means of Monte Carlo simulations and a finite-size scaling analysis. By using a field programmable gate array, we have been able to thermalize a large number of samples of systems with large volume. This has allowed us to observe a spin-glass ordered phase in d=4 and to study the critical properties of the transition. In d=3, our results are consistent with the presence of a Kosterlitz-Thouless transition, but also with different scenarios: transient effects due to a value of the lower critical dimension slightly below 3 could be very important.

Preparação, caracterização e propriedades de lipossomas contendo o ácido -ciano-4-hidroxicinâmico e o agente fotossenbilizador AICIPc: um novo sistema carreador específico com ação sinérgica aplicado a terapia fotodinâmica

Ftalocianina de alumínio-cloro (AlClPc) é um fotossensibilizador de segunda geração em terapia fotodinâmica (TFD) caracterizado por seu caráter anfifílico e tendência de auto-agregação em meio aquoso, o que prejudica seu potencial de aplicação. O aCHC é um substrato de transportadores de monocarboxilato (MCT) superexpresso em células de MCF-7. Objetivando a solubilização da AlClPc e aumento de internalização em tecidos neoplásicos nos propomos aqui o uso de DSPC e DOPC em diferentes proporções para formar vesículas lipidicas mistas (LV) na presença de aCHC como sistemas veiculadores de fármaco. Lv foi preparado pelo método de injeção etanólica e formou vesículas de dimensões nanométricas (aproximadamente 100 nm) com bom índice de polidispersão, valores negativos de potencial zeta e estáveis em meio aquoso por mais de 50 dias. AlClPc se complexou com o fosfato das LV o que conferiu uma localização interfacial às moléculas de AlClPc como demonstrado pelos resultados de supressão de fluorescência. Medidas de anisotropia, fluorescência estática e resolvida no tempo corroboram com estes resultados e demonstram que a auto-agregação da AlClPc ocorre mesmo em lipossomas. Entretanto, a veiculação da AlClPc por LV em carcinoma de células escamosas oral (OSCC) levou a um processo de desagregação demonstrado por (FLIM). Este incrível comportamento é novo e aumenta o conhecimento científico sobre o mecanismo intracelular de ação de fotossensibilizadores em TFD. Em TFD, ambos os sistemas LVIII+AlClPc e LVIII+AlClPc+aCHC não apresentaram toxicidade no escuro no período de incubação de 3 h com as concentrações de lipídios, AlClPc e aCHC iguais a 0,15 mmol/L, 0,5 umol/L e 10,0 umol/L, respectivamente. De maneira inesperada, o sistema LVIII+AlClPc foi mais eficiente em TFD que o sistema LVIII+AlClPc+aCHC, devido ao caráter antioxidante do aCHC. Estes resultados abrem uma nova perspectiva do potencial uso de LV-AlClPc para o tratamento fotodinâmico.

The Pliocene Mediterranean infilling of the Messinian Erosional Surface: New biostratigraphic data based on calcareous nannofossils (Bajo Segura Basin, SE Spain)

The Bajo Segura Basin (eastern Betic Cordillera) is a Mediterranean marginal basin where the Messinian Erosional Surface (MES), formed during the Messinian Salinity Crisis sea-level fall, is well developed. Overlying this major discontinuity the lower Pliocene transgressive sediments record the reflooding of the Mediterranean and the return to an open marine environment, the continental shelf being rebuilt after the Messinian erosion. The stratigraphic and biostratigraphic study of six sections allows two transgressive-regressive sequences filling the MES to be distinguished, correlated with the previously distinguished Mediterranean offshore seismic units. Ten calcareous nannofossil bioevents have been identified. The lower sequence can be dated according to nannofossil biozones NN12 to NN14 and the upper sequence by NN15 to NN16. The boundary between both lower Pliocene sedimentary sequences occur after the first common occurrence (FCO) of Discoaster asymmetricus found in the uppermost sediments of the lower sequence and before the first occurrence (FO) of Discoaster tamalis in the lowermost part of the upper sequence. Thus this sequence boundary can be estimated at between 4.1 and 4.0Ma ago.

Tab. 1: Rates of advance of the frontal push-moraines of 19 lobes in front of an approx. 4.5 km long margin of the Kötlujökull

In the last years masses of ice, about 5 km long, have been protruding from the lowest part of an advancing glacier margin of the Kötlujökull in Southern Iceland. In the summer of 1983, they appeared as sediment-covered lobes, 10-60 m long, bordering the glacier rnargin like agarland. 1 to 3 push-rnoraines without ice core, rnostly sickle-shaped, occured first in the frontal parts of the lobes: behind thern came several ice-cored moraines with heights of up to several metres. The active ice in front of the precipice of the glacier is called the "glacier-foot" in this paper. The digging out of 9 lobes and the measuring of the advance of 19 lobes showed that in most cases this glacierfoot had split up at its distal end into several plate- or stem-shaped pieces of ice which were situated one upon the other, separated by moraine deposits and proceeding irregularly into the foreland at the rate of several mm/h, The sometimes different rate of advance in the same lobe and different rates of advanee in adjoining lobes (some being entirely inactive) point to a type of rnovement which is independent of the general advance of the glacier. Research in the winter of 1983/84 showed less activity in 3 examined lobes, but the activity had not ceased. The advancement of the lower parts of the glacier-foot into and across the sands of the foreland implies the following genesis of pushmoraines: Shoving off a plate of sand, folding it and pushing it over the foreland at average rates of up to 7,2 mm/h, according to the investigations in thc summer of 1983. At a certain stage of the folding process, new folds begin to develop in front of the old, and the old folds are shifted onto the backslope of thc folds in front of them until they are completely unired. In this way, "püe-moraines" arise, which become higher and higher. They include two or more folds declining towards the glacier. Systems of small moraines presumably of the same genesis occur on old moraine areas in front of the Kötlujökull. The possible cause of formation of a glacier-foot is discussed, and the moraines of the Kötlujökull are compared with certain pleistocene push-moraines.

Sedimentation rates calculated on surface sediment samples from different site of the Atlantic and Pacific Oceans (Table 1)

Radiocarbon ages on CaCO3 from deep-sea cores offer constraints on the nature of the CaCO3 dissolution process. The idea is that the toll taken by dissolution on grains within the core top bioturbation zone should be in proportion to their time of residence in this zone. If so, dissolution would shift the mass distribution in favor of younger grains, thereby reducing the mean radiocarbon age for the grain ensemble. We have searched in vain for evidence supporting the existence of such an age reduction. Instead, we find that for water depths of more than 4 km in the tropical Pacific the radiocarbon age increases with the extent of dissolution. We can find no satisfactory steady state explanation and are forced to conclude that this increase must be the result of chemical erosion. The idea is that during the Holocene the rate of dissolution of CaCO3 has exceeded the rain rate of CaCO3. In this circumstance, bioturbation exhumes CaCO3 from the underlying glacial sediment and mixes it with CaCO3 raining from the sea surface.

Overdeepened glacial basins as archives for the Quaternary landscape evolution of the Alps

The Alps and the Alpine foreland have been shaped by repeated glaciations during Quaternary glacial-interglacial cycles. Extent, timing and impact on landscape evolution of these glaciations are, however, poorly constrained due to the fragmentary character of terrestrial archives. In this context, the sedimentary infills of subglacially eroded, ‘overdeepened’, basins may serve as important archives to complement the Quaternary stratigraphy over several glacial-interglacial cycles. In this thesis, the infills of deep subglacial basins in the Lower Glatt valley (N Switzerland) are explored to better constrain the Middle- to Late Pleistocene environmental change. Five drill cores gave direct insight into to the up to ~200 m thick valley fill at the study site and allowed for detailed analysis of sedimentary facies, age and architecture of the basin fills. A first focus is set on the sedimentology of coarse-grained diamicts with sorted interbeds overlying bedrock in the trough center, which mark the onset of deposition in many glacial bedrock troughs. Evidence from macro- and microsedimentology suggests that these sediments are emplaced subglacially and reflect deposition, reworking and deformation in response to repeated coupling and decoupling of the ice-bed interface promoted by high basal water pressures. Overlying these subglacial sediments, large volumes of sandy glacio-deltaic, fine-grained glacio-lacustrine and lacustrine sediments document sedimentation during glacier retreat from the basins. On these thick valley fill sequences the applicability and reliability of luminescence dating is investigated in a second step on the basis of experiments with several different luminescence signals, protocols and experiments to assess the signal stability. The valley fill of the Lower Glatt valley is then grouped into nine depositional cycles (Formations A-I), which are related to the Birrfeld Glaciation (~MIS2), the Beringen Glaciation (~MIS6), and up to three earlier Middle Pleistocene glaciations, tentatively correlated to the Hagenholz, Habsburg, and Möhlin Glaciations, according to the regional glaciation history. The complex bedrock geometry and valley fill architecture are shown to be the result of multiple erosion and infilling cycles and reflect the interplay of subglacial erosion, glacial to lacustrine infilling of overdeepened basins, and fluvial down-cutting and aggradation in the non-overdeepened valley fill. Evidence suggests that in the study area deep bedrock incision, and/or partial re-excavation, occurred mainly during the Beringen and Hagenholz Glaciation, while older structures may have existed. Together with the observation of minor, ‘inlaid’ glacial basins, dynamic changes in the magnitude and focus of subglacial erosion over time are documented.

The chirurgical works of Percivall Pott, with his last corrections. : to which are added, a short account of the life of the author, a method of curing the hydrocele by injection, and occasional notes and observations /

Published in London in 1771 and 1808.

Transactions of the third International congress for the history of religions.

Edited under the general direction of the Papers committee by Mr. P. S. Allen ... assisted by Mr. J. de M. Johnson."