A novel ligand-independent function of the estrogen receptor is essential for osteocyte and osteoblast mechanotransduction

Bone senses and adapts to meet mechanical needs by means of an extensive mechanotransduction network comprising osteocytes (former osteoblasts entrapped in mineral) and their cytoplasmic projections through which osteocytes communicate with osteoblasts and osteoclasts on the bone surface. Mechanical stimulation promotes osteocyte (and osteoblast) survival by activating the extracellular signal-regulated kinases, ERKs. Estrogens have similar effects and, intriguingly, the adaptive response of bone to mechanical forces is defective in mice lacking estrogen receptor (ER) alpha or ERbeta. We report that ERKs are not activated by stretching in osteocytic and osteoblastic cells in which both ERalpha and ERbeta have been knocked out or knocked down and this is reversed partially by transfection of either one of the two human ERs and fully by transfection of both receptors. ERK activation in response to stretching is also recovered by transfecting the ligand-binding domain (E) of either receptor or an ERalpha mutant that does not bind estrogens. Furthermore, mechano-responsiveness is restored by transfecting the Ealpha targeted to the plasma membrane, but not to the nucleus, whereas ERalpha mutants with impaired plasma membrane localization or binding to caveolin-1 fail to confer ERK activation in response to stretching. Lastly, the ER antagonist ICI 182,780 abrogates ERK activation and the anti-apoptotic effect of mechanical stimulation. We conclude that in addition to their role as ligand-dependent mediators of the effects of estrogens, the ERs participate in the transduction of mechanical forces into pro-survival signaling in bone cells, albeit in a ligand-independent manner.     

Proteome, salicylic acid, and jasmonic acid changes in cucumber plants inoculated with Trichoderma asperellum strain T34

Trichoderma spp. is one of the most commonly used biological control agents against plant pathogens. This fungus produces changes in plant metabolism, thus increasing growth and enhancing resistance to biotic and abiotic stresses. However, its modes of action remain to be defined. In the first hours of interaction between cucumber plant roots and Trichoderma asperellum strain T34, salicylic and jasmonic acid levels and typical antipathogenic peroxidase activity increase in the cotyledons to different degrees depending on the applied concentration of the fungi. The use of 2-DE protein profiling and MS analysis allowed us to identify 28 proteins whose expression was affected in cotyledons after cucumber root colonization by Trichoderma applied at high concentrations: 17 were found to be up-regulated while 11 were down-regulated. Proteins involved in ROS scavenging, stress response, isoprenoid and ethylene biosynthesis, and in photosynthesis, photorespiration, and carbohydrate metabolism were differentially regulated by Trichoderma. The proteome changes found in this study help to give an understanding of how Trichoderma-treated plants become more resistant to pathogen attacks through the changes in expression of a set of defence-oriented proteins which can directly protect the plant or switch the metabolism to a defensive, nonassimilatory state.     

Role of cofactors B (TBCB) and E (TBCE) in tubulin heterodimer dissociation

Tubulin folding cofactors B (TBCB) and E (TBCE) are alpha-tubulin binding proteins that, together with Arl2 and cofactors D (TBCD), A (TBCA or p14) and C (TBCC), participate in tubulin biogenesis. TBCD and TBCE have also been implicated in microtubule dynamics through regulation of tubulin heterodimer dissociation. Understanding the in vivo function of these proteins will shed light on the Kenny-Caffey/Sanjad-Sakati syndrome, an important human disorder associated with TBCE. Here we show that, when overexpressed, TBCB depolymerizes microtubules. We found that this function is based on the ability of TBCB to form a binary complex with TBCE that greatly enhances the efficiency of this cofactor to dissociate tubulin in vivo and in vitro. We also show that TBCE, TBCB and alpha-tubulin form a ternary complex after heterodimer dissociation, whereas the free beta-tubulin subunit is recovered by TBCA. These complexes might serve to escort alpha-tubulin towards degradation or recycling, depending on the cell requirements.     

Spatial variation of shell morphometrics in the subantarctic land snail <Emphasis Type="Italic">Notodiscus hookeri</Emphasis> from Crozet and Kerguelen Islands

The land snail Notodiscus hookeri, widely distributed in subantarctic islands, shows a large intraspecific variation in shell morphology. In the present work, shell size and form of individuals from populations located in Crozet and Kerguelen archipelagos were investigated by means of multivariate statistics. Variation in shell morphometrics was analysed after the partitioning of the overall variation into size and shape components by means of a principal component-based approach. Shell size shows a significant spatial heterogeneity, which seems essentially related to environmental pressures. Previous works pointed to a greater conchological variation between populations from Kerguelen but present observations show that intra-island variances are not significantly different in the two islands studied. Variation in shell shape splits the populations into two main entities because of different allometric relationships between two shell height components and all other measurements. However, using geographical affinities of populations as instrumental variables shows that more complex environmental features interfered in population clustering.     

Intercalation of proflavine and a platinum derivative of proflavine into double-helical Poly(A)

The equilibria and kinetics of the interactions of proflavine (PR) and its platinum-containing derivative [PtCl(tmen)(2)HNC(13)H(7)(NHCH(2)CH(2))(2)](+) (PRPt) with double-stranded poly(A) have been investigated by spectrophotometry and Joule temperature-jump relaxation at ionic strength 0.1 M, 25 degrees C, and pH 5.2. Spectrophotometric measurements indicate that base-dye interactions are prevailing. T-jump experiments with polarized light showed that effects due to field-induced alignment could be neglected. Both of the investigated systems display two relaxation effects. The kinetic features of the reaction are discussed in terms of a two-step series mechanism in which a precursor complex DS(I) is formed in the fast step, which is then converted to a final complex in the slow step. The rate constants of the fast step are k(1) = (2.5 +/- 0.4) x 10(6) M(-1) s(-1), k(-1) = (2.4 +/- 0.1) x 10(3) s(-1) for poly(A)-PR and k(1) = (2.3 +/- 0.1) x 10(6) M(-1) s(-1), k(-1) = (1.6 +/- 0.2) x 10(3) s(-1) for poly(A)-PRPt. The rate constants for the slow step are k(2) = (4.5 +/- 0.5) x 10(2) s(-1), k(-2) = (1.7 +/- 0.1) x 10(2) s(-1) for poly(A)-PR and k(2) = 9.7 +/- 1.2 s(-1), k(-2) = 10.6 +/- 0.2 s(-1) for poly(A)-PRPt. Spectrophotometric measurements yield for the equilibrium constants and site size the values K = (4.5 +/- 0.1) x 10(3) M(-1), n = 1.3 +/- 0.5 for poly(A)-PR and K = (2.9 +/- 0.1) x 10(3) M(-1), n = 2.3 +/- 0.6 for poly(A)-PRPt. The values of k(1) are similar and lower than expected for diffusion-limited reactions. The values of k(-1) are similar as well. It is suggested that the formation of DS(I) involves only the proflavine residues in both systems. In contrast, the values of k(2) and k(-2) in poly(A)-PRPt are much lower than in poly(A)-PR. The results suggest that in the complex DS(II) of poly(A)-PRPt both proflavine and platinum residues are intercalated. In addition, a very slow process was detected and ascribed to the covalent binding of Pt(II) to the adenine.     

2-Methyl-2-hydroxymethylchromenes from Artemisia campestris subsp. glutinosa

From the non volatile neutral part of a hexane extract from Artemisia campestris subsp. glutinosa, two new 2-methyl-2-hydroxymethyl chromenes h     

MEK1 inactivates Myt1 to regulate Golgi membrane fragmentation and mitotic entry in mammalian cells

The pericentriolar stacks of Golgi cisternae are separated from each other in G2 and fragmented extensively during mitosis. MEK1 is required for Golgi fragmentation in G2 and for the entry of cells into mitosis. We now report that Myt1 mediates MEK1's effects on the Golgi complex. Knockdown of Myt1 by siRNA increased the efficiency of Golgi complex fragmentation by mitotic cytosol in permeabilized and intact HeLa cells. Myt1 knockdown eliminated the requirement of MEK1 in Golgi fragmentation and alleviated the delay in mitotic entry due to MEK1 inhibition. The phosphorylation of Myt1 by MEK1 requires another kinase but is independent of RSK, Plk, and CDK1. Altogether our findings reveal that Myt1 is inactivated by MEK1 mediated phosphorylation to fragment the Golgi complex in G2 and for the entry of cells into mitosis. It is known that Myt1 inactivation is required for CDK1 activation. Myt1 therefore is an important link by which MEK1 dependent fragmentation of the Golgi complex in G2 is connected to the CDK1 mediated breakdown of Golgi into tubules and vesicles in mitosis.     

Heliangolides from Leucanthemopsis pulverulenta

The structure of hispanolide (4,5-cis-3-oxogermacranolide) and three other related germacranolides, isolated from Leucanthemopsis pulverulenta, have been established by a series of chemical transformations and spectral data.     

Elaboración y validación de un nuevo cuestionario de hábitos alimentarios para pacientes con sobrepeso y obesidad

IntroductionWhen designing a comprehensive and personalized treatment for weight control, the information provided by obese patients on their dietary habits must be gathered correctly. Our objective was to develop and validate a new questionnaire on dietary habits for overweight and obese patients that could be used clinically.Material and methodsThe new questionnaire was developed and validated through a three-step process. Firstly, an existing questionnaire was reviewed. Secondly, a new questionnaire was developed, which included new items. Thirdly, scales to aid interpretation of the results in clinical practice were developed. Construct validity was assessed using factor analysis. The internal consistency reliability was analyzed.ResultsThe validity and reliability of the new questionnaire was confirmed. The values of the patients' responses to the new questionnaire were related to variables such as age, sex, body mass index, and whether or not the patients were under treatment for hypertension, diabetes mellitus or dyslipidemia.ConclusionThe validity and reliability of this questionnaire and the scales that aid their interpretation make it a useful tool for initial assessment and monitoring of changes in eating habits in response to treatment both individually and collectively in the Spanish population.     

The stromal‐vascular fraction of adipose tissue contributes to major differences between subcutaneous and visceral fat depots

Adipose tissue represents a complex tissue both in terms of its cellular composition, as it includes mature adipocytes and the various cell types comprising the stromal‐vascular fraction (SVF), and in relation to the distinct biochemical, morphological and functional characteristics according to its anatomical location. Herein, we have characterized the proteomic profile of both mature adipocyte and SVF from human visceral adipose tissue (VAT) and subcutaneous adipose tissue (SAT) fat depots in order to unveil differences in the expression of proteins which may underlie the distinct association of VAT and SAT to several pathologies. Specifically, 24 proteins were observed to be differentially expressed between SAT SVF versus VAT SVF from lean individuals. Immunoblotting and RT‐PCR analysis confirmed the differential regulation of the nuclear envelope proteins lamin A/C, the membrane‐cytoskeletal linker ezrin and the enzyme involved in retinoic acid production, aldehyde dehydrogenase 1A2, in the two fat depots. In sum, the observation that proteins with important cell functions are differentially distributed between VAT and SAT and their characterization as components of SVF or mature adipocytes pave the way for future research on the molecular basis underlying diverse adipose tissue‐related pathologies such as metabolic syndrome or lipodystrophy.