Delay of EGF-Stimulated EGFR Degradation in Myotonic Dystrophy Type 1 (DM1)

Funding Information: This research was supported by the Isabel Gemio Foundation (P18–13) and was also partially supported by the “Fondo Europeo de Desarrollo Regional” (FEDER) from the European Union. E.A.-C. was supported by a pre-doctoral fellowship of Valhondo Calaff Foundation. S.C.-C. and E.U.-C. were supported by FPU fellowships (FPU19/04435 and FPU16/00684, respectively) from the Ministerio de Ciencia, Innovación y Universidades, Spain. M.P.-B. and A.G.-B. received fellowships from the “Plan Propio de Iniciación a la Investigación, Desarrollo Tecnológico e Innovación (Universidad de Extremadura). M.N.-S. was supported by the “Ramon y Cajal” Program (RYC-2016–20883), and P.G.-S., was funded by “Juan de la Cierva Incorporación” Program (IJC2019–039229-I), Spain. S.M.S.Y.-D. was supported by the Isabel Gemio Foundation and CIBERNED (CB06/05/0041). J.M.F received research support from the Isabel Gemio Foundation and the “Instituto de Salud Carlos” III, CIBERNED (CB06/05/0041). Publisher Copyright: © 2022 by the authors.Myotonic dystrophy type 1 (DM1) is an autosomal dominant disease caused by a CTG repeat expansion in the 3′ untranslated region of the dystrophia myotonica protein kinase gene. AKT dephosphorylation and autophagy are associated with DM1. Autophagy has been widely studied in DM1, although the endocytic pathway has not. AKT has a critical role in endocytosis, and its phosphorylation is mediated by the activation of tyrosine kinase receptors, such as epidermal growth factor receptor (EGFR). EGF-activated EGFR triggers the internalization and degradation of ligand–receptor complexes that serve as a PI3K/AKT signaling platform. Here, we used primary fibroblasts from healthy subjects and DM1 patients. DM1-derived fibroblasts showed increased autophagy flux, with enlarged endosomes and lysosomes. Thereafter, cells were stimulated with a high concentration of EGF to promote EGFR internalization and degradation. Interestingly, EGF binding to EGFR was reduced in DM1 cells and EGFR internalization was also slowed during the early steps of endocytosis. However, EGF-activated EGFR enhanced AKT and ERK1/2 phosphorylation levels in the DM1-derived fibroblasts. Therefore, there was a delay in EGF-stimulated EGFR endocytosis in DM1 cells; this alteration might be due to the decrease in the binding of EGF to EGFR, and not to a decrease in AKT phosphorylation.publishersversionpublishe

Dust environment and dynamical history of a sample of short-period comets: II. 81P/Wild 2 and 103P/Hartley 2

Aims. This paper is a continuation of the first paper in this series, where we presented an extended study of the dust environment of a sample of short-period comets and their dynamical history. On this occasion, we focus on comets 81P/Wild 2 and 103P/Hartley 2, which are of special interest as targets of the spacecraft missions Stardust and EPOXI. Methods. As in the previous study, we used two sets of observational data: a set of images, acquired at Sierra Nevada and Lulin observatories, and the Afρ data as a function of the heliocentric distance provided by the amateur astronomical association Cometas-Obs. The dust environment of comets (dust loss rate, ejection velocities, and size distribution of the particles) was derived from our Monte Carlo dust tail code. To determine their dynamical history we used the numerical integrator Mercury 6.2 to ascertain the time spent by these objects in the Jupiter family Comet region. Results. From the dust analysis, we conclude that both 81P/Wild 2 and 103P/Hartley 2 are dusty comets, with an annual dust production rate of 2.8 × 109 kg yr-1 and (0.4-1.5) × 109 kg yr-1, respectively. From the dynamical analysis, we determined their time spent in the Jupiter family Comet region as ~40 yr in the case of 81P/Wild 2 and ~1000 yr for comet 103P/Hartley 2. These results imply that 81P/Wild 2 is the youngest and the most active comet of the eleven short-period comets studied so far, which tends to favor the correlation between the time spent in JFCs region and the comet activity previously discussed

Surface-enhanced vibrational study of azabipiridyl and its Co(II), Ni(II) and Cu(II) complexes

The FT-Raman spectra and surface-enhanced Raman scattering (SERS) on silver colloid of the azabipiridyl ligand and its Co(II), Ni(II) and Cu(II) complexes are reported. A resonance Raman (RR) study of the solids and a surface-enhanced infrared absorption (SEIRA) study were also accomplished. The spectral analysis indicates that there is a remarkable enhancement of the Raman spectra induced by the presence of an Ag surface. The complex-surface interaction is verified by the appearance of a ν(Ag-N) band at about 216 cm-1. A general energy increasing of most of the molecular modes by surface effect is associated to a charge transfer from the colloidal silver surface, thus, leading to an important contribution of the charge transfer mechanism to the SERS enhancement, which is corroborated by the resonant effect observed at different excitation lines. From the analysis of the SERS spectra a preferential planar parallel orientation of the complexes on the surface is deduced, in such a way t

Molecular Recognition of Epothilones by Microtubules and Tubulin Dimers Revealed by Biochemical and NMR Approaches

The binding of epothilones to dimeric tubulin and to microtubules has been studied by means of biochemical and NMR techniques. We have determined the binding constants of epothilone A (EpoA) and B (EpoB) to dimeric tubulin, which are 4 orders of magnitude lower than those for microtubules, and we have elucidated the conformation and binding epitopes of EpoA and EpoB when bound to tubulin dimers and microtubules in solution. The determined conformation of epothilones when bound to dimeric tubulin is similar to that found by X-ray crystallographic techniques for the binding of EpoA to the Tubulin/RB3/TTL complex; it is markedly different from that reported for EpoA bound to zinc-induced sheets obtained by electron crystallography. Likewise, only the X-ray structure of EpoA bound to the Tubulin/RB3/TTL complex at the luminal site, but not the electron crystallography structure, is compatible with the results obtained by STD on the binding epitope of EpoA bound to dimeric tubulin, thus confirming that the allosteric change (structuring of the M-loop) is the biochemical mechanism of induction of tubulin assembly by epothilones. TR-NOESY signals of EpoA bound to microtubules have been obtained, supporting the interaction with a transient binding site with a fast exchange rate (pore site), consistent with the notion that epothilones access the luminal site through the pore site, as has also been observed for taxanes. Finally, the differences in the tubulin binding affinities of a series of epothilone analogues has been quantitatively explained using the newly determined binding pose and the COMBINE methodology.</p

Ten-year follow-up of an anatomical hydroxyapatite-coated total hip prosthesis

We report our results after ten year follow-up of 107 consecutive ABG-I hip prostheses implanted between June 1990 and December 1992: Only 84 prostheses were still in the study after ten years, but only six patients had undergone surgical revision. We can consider our clinical outcomes as excellent, with a whole-implant survival rate greater than 96%, a mean Merle D’Aubigne and Postel score increasing from 7.97 before operation to 16.17 at ten year follow-up, and a personal subjective assessment as excellent or good in 82.14% of patients. However, radiographic outcomes are more worrying: around 90% of patients show a stress-shielding phenomenon and granulomatous lesions in the proximal femur, and more than 82% suffer polyethylene wear greater than one  millimetre (mean 1.68 mm). We think that zirconia stem heads and hooded antiluxation PE inserts are determining factors in the process of PE wear and, secondarily, in cancellous bone resorption and bone osteolysis