Phosphorylation of the eIF4E-binding protein PHAS-I after exposure of PC12 cells to EGF and NGF

AbstractPHAS-I or the eIF4E-binding protein 1 regulates the cap-binding activity of eIF4E by sequestering eIF4E. Binding of eIF4E to PHAS-I is regulated by phosphorylation of PHAS-I. PC12 cells were used to study the signal transduction pathway leading to phosphorylation of PHAS-I. Both EGF and NGF induced phosphorylation of PHAS-I. Wortmannin, a PI-3 kinase inhibitor, staurosporine, a PKC inhibitor, and rapamycin, a FRAP inhibitor all blocked the phosphorylation of PHAS-I. Of the three inhibitors, only wortmannin was able to inhibit MAPK phosphorylation. This excludes a role for MAPK in NGF- and EGF-induced PHAS-I phosphorylation in PC12 cells. Apparently, PHAS-I was phosphorylated in a PI-3 kinase-, PKC-, and FRAP-dependent manner after EGF or NGF stimulation. Only PI-3 kinase and FRAP are involved in the regulation of the basal level of PHAS-I phosphorylation

Angular and energy dependence of ion bombardment of Mo/Si multilayers

The process of ion bombardment is investigated for the fabrication of Mo/Si multilayer x-ray mirrors using e-beam evaporation. The ion treatment is applied immediately after deposition of each of the Si layers to smoothen the layers by removing an additional thickness of the Si layer. In this study the parameters of Kr+ ion bombardment have been optimized within the energy range 300 eV-2 keV and an angular range between 20 degrees and 50 degrees. The optical performance of the Mo/Si multilayers is determined by absolute measurements of the near-normal-incidence reflectivity at 14.4 nm wavelength. The multilayer structures are analyzed further with small-angle reflectivity measurements using both specular reflectivity and diffuse x-ray scattering. The optimal smoothening parameters are obtained by determining the effect of ion bombardment on the interface roughness of the Si layer. The optimal conditions are found to be 2 keV at 50 degrees angle of incidence with respect to the surface. These settings result in 47% reflectivity at 85 degrees (lambda = 14.4 nm) for a 16-period Mo/Si multilayer mirror, corresponding to an interface roughness of 0.21 nm rms. Analysis shows that the interface roughness is determined by ion induced viscous flow, an effect which increases with ion energy as well as angle of incidence. In order to determine the effect of intermixing of the Si and Mo atoms, the penetration depth of the Kr+ ions is calculated as a function of ion energy and angle of incidence. Furthermore, the angular dependence of the etch yield, obtained from the in situ reflectivity measurements, is investigated in order o determine the optimal ion beam parameters for the production of multilayer mirrors on curved substrates. (C) 1997 American Institute of Physics

Succes en tegenslag bij vaccinonderzoek, van mazelen tot malaria en AIDS

In 1974 werd ik benoemd tot hoogleraar moleculaire biologie bij deze universiteit. Een van de aantrekkelijke dingen om naar Utrecht te komen was de aanwezigheid van het IMB, het Instituut voor Moleculaire Biologie. Een door het College van Bestuur ingestelde en gefinancierde laboratoriumfaciliteit in Transitorium 3, waar een hoeveelheid kostbare apparatuur ten behoeve van moleculair biologisch onderzoek geconcentreerd was. Coryfeeën van toen, Henk Jansz, Gerard van Arkel, Laurent van Deenen, Peter de Haan, Peter Elbers en David de Wied waren werkgroepleiders en mij werd gevraagd om als relatieve buitenstaander voorzitter te worden, bijgestaan door Harry van Someren als beheerder

Mo/Si multilayer optics for micro-lithography

Applied Science

Design of an Extended Image Field Soft-X-Ray Projection System

A soft-x-ray projection system has been designed, which consists of spherical components to be coated with multilayer reflection coatings. In the design, a two-mirror system and a spherical reflection mask, the optical aberrations were minimized. The design enables a resolution of sub-100 nm over a circular image field with a diameter of 4 mm at a wavelength of 10.5 nm. The assembly tolerances of the system and the fabrication tolerances of the substrates have been calculated. The surface roughness determined from our superpolished quartz substrates amounts to 0.3 nm, which is sufficient to meet the required specifications

Purification of free eukaryotic initiation factors eIF-4A and eIF-4D on cibacron blue F3G-A

The eukaryotic initiation factors eIF-4A and eIF-4D are almost exclusively found in the supernatant of a reticulocyte lysate. The basic steps of general purification schemes failed because these factors do not bind to Sepharoseheparin or, in the case of eIF-4A, to phosphocellulose and a new procedure had to be devised. Cibacron Blue F3G-A proved to be a successful alternative in the purification of these factors from the excessive load of proteins found in the supernatant

Ultraviolet-crosslinking reveals specific affinity of eukaryotic initiation factors for Semliki Forest virus mRNA

Eukaryotic initiation factors (elF) associate readily with ³²P-labeled Semliki Forest virus (SFV) mRNA in vitro, forming complexes which can be crosslinked by 254 nm ultraviolet irradiation. After ribonuclease digestion, the initiation factors were released and analysed by gel electrophoresis. Autoradiography revealed proteins by virtue of crosslinked ³²P-labeled mRNA fragments, elF-4A, -4B and -4C as well as three subunits of elF-3 could be crosslinked with SFV mRNA. None of these proteins bound to ribosomal RNAs

Minimal energy foldings of eukaryotic mRNAs form a separate leader domain

We have investigated the minimal energy foldings of 38 mature mRNAs, including the globin family, the insulins, the growth hormones and interleukin-2, and have compared these foldings with those of fully and partly randomised sequences. The mRNAs differ from the random sequences in that they form a separate leader hairpin of 40–60 nucleotides, with the initiation codon typically located downstream of this hairpin, followed by a main fold in which a region flanking the initiation codon is basepaired with the trailer: resulting in a close proximity of the 5′ and 3′ end of the mRNA. The formation of this conformation depends not only—or primarily—on the structure of the leader, but on both the leader and trailer sequence and their interaction with the coding sequence. Thus if, as the frequent occurrence of this pattern suggests, the secondary structure of the leader regions plays a role in the initiation of translation, possibly accounting for the specificity of initiation and the different translational efficiencies of various mRNAs, we expect that these features may be influenced both by leader and trailer mutants