The role of BiP in endoplasmic reticulum-associated degradation of major histocompatibility complex class I heavy chain induced by cytomegalovirus proteins.

Human cytomegalovirus (HCMV1) US11 and US2 proteins cause rapid degradation of major histocompatibility complex (MHC) molecules, apparently by ligating cellular endoplasmic reticulum (ER)-associated degradation machinery. Here, we show that US11 and US2 bind the ER chaperone BiP. Four related HCMV proteins, US3, US7, US9, and US10, which do not promote degradation of MHC proteins, did not bind BiP.

T-type CaV3.3 calcium channels produce spontaneous low-threshold action potentials and intracellular calcium oscillations.

The precise contribution of T-type Ca2+ channels in generating action potentials (APs), burst firing and intracellular Ca2+ signals needs further elucidation. Here, we show that CaV3.3 channels can trigger repetitive APs, generating spontaneous membrane potential oscillations (MPOs), and a concomitant increase in the intracellular Ca2+ concentration ([Ca2+]i) when overexpressed in NG108-15 cells.

[Dietary water intoxication: a preventable pathology].

Case Report: The authors report the case of an infant presenting with status epilepticus, apneas and hypothermia, related to hyponatremia. The cause was water intoxication, secondary to feeding a dilute formula and free water. The parents were notoriously challenged by the care of their children.

Synthesis, physical properties, multitemperature crystal structure, and 20 K synchrotron X-ray charge density of a magnetic metal organic framework structure, Mn3(C8O4H4)3(C5H11ON)2.

A new magnetic metal organic framework material has been synthesized, Mn3(C8O4H4)3(C5H11ON)2, 1. Magnetic susceptibility measurements from 2 to 400 K reveal anti-ferromagnetic ordering at approximately 4 K and a total magnetic moment of 6.0 micro(B).

A survey of patient dose and clinical factors in a full-field digital mammography system.

In this work, we analyse the dose to 5034 patients (20 137 images) who underwent mammographic examinations with a full-field digital mammography (FFDM) system over a 2-y period. The information relevant to this study has been extracted from the image Digital Imaging and Communications in Medicine (DICOM) headers. Entrance surface air-kerma (ESAK) without backscatter and average glandular dose (AGD) were estimated following the methodology proposed in the European Protocol on Dosimetry in Mammography.

Slip-rate measurements on the Karakorum Fault may imply secular variations in fault motion.

Beryllium-10 surface exposure dating of offset moraines on one branch of the Karakorum Fault west of the Gar basin yields a long-term (140- to 20-thousand-year) right-lateral slip rate of approximately 10.7 +/- 0.7 millimeters per year.

Patient dose in digital mammography.

In the present investigation, we analyze the dose of 5034 patients (20,137 images) who underwent mammographic examinations with a full-field digital mammography system. Also, we evaluate the system calibration by analyzing the exposure factors as a function of breast thickness. The information relevant to this study has been extracted from the image DICOM header and stored in a database during a 3-year period (March 2001-October 2003).

Characterization of different strains of poliovirus and influenza virus by differential scanning calorimetry.

Vaccines against poliomyelitis and influenza contain inactivated forms of poliovirus and influenza virus. These antigens are generated on an industrial scale from the purified active viruses that have been analysed in this study by DSC (differential scanning calorimetry). Multiple unfolding transitions are seen for influenza virus A/New Caledonia/20/99 (H1N1), A/Panama/2007/99 (H3N2) and B/Shangdong/7/97.

HIV-1 gp41 and gp160 are hyperthermostable proteins in a mesophilic environment. Characterization of gp41 mutants.

HIV gp41(24-157) unfolds cooperatively over the pH range of 1.0-4.0 with T(m) values of > 100 degrees C.

Human cytomegalovirus US3 chimeras containing US2 cytosolic residues acquire major histocompatibility class I and II protein degradation properties.

Human cytomegalovirus (HCMV) glycoprotein US2 increases the proteasome-mediated degradation of major histocompatibility complex (MHC) class I heavy chain (HC), class II DR-alpha and DM-alpha proteins, and HFE, a nonclassical MHC protein. US2-initiated degradation of MHC proteins apparently involves the recruitment of cellular proteins that participate in a process known as endoplasmic reticulum (ER)-associated degradation. ER-associated degradation is a normal process by which misfolded proteins are recognized and translocated into the cytoplasm for degradation by proteasomes.

Insight into the structure and function of the transferrin receptor from Neisseria meningitidis using microcalorimetric techniques.

The transferrin receptor of Neisseria meningitidis is composed of the transmembrane protein TbpA and the outer membrane protein TbpB. Both receptor proteins have the capacity to independently bind their ligand human transferrin (htf). To elucidate the specific role of these proteins in receptor function, isothermal titration calorimetry was used to study the interaction between purified TbpA, TbpB or the entire receptor (TbpA + TbpB) with holo- and apo-htf.

Binding of human cytomegalovirus US2 to major histocompatibility complex class I and II proteins is not sufficient for their degradation.

Human cytomegalovirus (HCMV) glycoprotein US2 causes degradation of major histocompatibility complex (MHC) class I heavy-chain (HC), class II DR-alpha and DM-alpha proteins, and HFE, a nonclassical MHC protein. In US2-expressing cells, MHC proteins present in the endoplasmic reticulum (ER) are degraded by cytosolic proteasomes. It appears that US2 binding triggers a normal cellular pathway by which misfolded or aberrant proteins are translocated from the ER to cytoplasmic proteasomes.

Affinity-purification of Transferrin-binding protein B under nondenaturing conditions.

The commonly used purification procedures for Transferrin-binding protein B (TbpB) are based on an affinity chromatography step using resins onto which human transferrin had been immobilized. These protocols involve protein elution using denaturing buffer solutions. Here we present an improved protocol which permits protein elution under nondenaturing conditions using chelating agents such as phosphate or compounds containing a pyrophosphate group.

Modulation of the antibody response to the HIV envelope subunit by co-administration of infectious or heat-inactivated canarypoxvirus (ALVAC) preparations.

Poxviruses are large DNA viruses capable of infecting a broad range of animal species. Infection is generally accompanied by an inflammatory response in the host, the extent of which varies considerably with the specific poxvirus and host species. Regarding ALVAC, a poxvirus derived from the canarypox vaccine strain, Kanapox, and which represents a promising immunization vehicle in humans, nothing is known about its inflammatory capacity.

Isolation, expression, and characterization of fully functional nontoxic BiP/GRP78 mutants.

Mammalian BiP/GRP78 and Escherichia coli DnaK belong to the highly conserved hsp70 family and function as molecular chaperones in the endoplasmic reticulum or the cytosol, respectively. Induction of murine BiP/GRP78 expression in E. coli leads to growth arrest and cell death, independent of the bacterial strain and vector used.

A sensitive mapping strategy for monitoring the reproducibility of glycan processing in an HIV vaccine, RGP-160, expressed in a mammalian cell line.

The external envelope glycoprotein (gp160) of HIV-1 is a candidate for vaccines against AIDS. Most of the surface of the molecule is shielded by carbohydrate and the structures and locations of these glycans may be important in defining the immunogenicity of the viral coat. Here we report a sensitive mapping strategy for profiling and analysing the N-glycosylation of gp160, based on chemical release of glycans, fluorescent labelling and HPLC analysis.

Interaction of murine BiP/GRP78 with the DnaJ homologue MTJ1.

The activity of Hsp70 proteins is regulated by accessory proteins, among which the most studied are the members of the DnaJ-like protein family. BiP/GRP78 chaperones the translocation and maturation of secreted and membrane proteins in the endoplasmic reticulum. No DnaJ-like partner has been described so far to regulate the function of mammalian BiP/GRP78.

One-step capillary isoelectric focusing for the separation of the recombinant human immunodeficiency virus envelope glycoprotein glycoforms.

One-step capillary isoelectric focusing was investigated as a rapid method to resolve the glycoforms of the heterogeneous recombinant human immunodeficiency virus (HIV) envelope glycoprotein (rgp 160sMN/LAI). The separation was performed in a poly(vinyl alcohol) (PVA) coated capillary using a mixture of ampholyte of narrow and wide pH range. A combination of saccaharose and 3-(cyclohexylamino)-1-propanesulfonic acid was shown to be the most efficient additive to avoid protein precipitation which occurs at a pH close to its pI.

Specificity of peptide-induced depolymerization of the recombinant carboxy-terminal fragment of BiP/GRP78.

In the present study, we have used a non-denaturing gel electrophoresis assay to characterize the specificity of the peptide-induced depolymerization process of the isolated recombinant C-terminal domain (C30) of the molecular chaperone BiP, in the presence of specific synthetic peptides and with the neuropeptide Substance P. In the absence of peptidic ligand, C30 self-associates readily into multiple oligomeric species. Upon peptide addition, C30 oligomers convert into dimers, then into monomers.

Substrate binding induces depolymerization of the C-terminal peptide binding domain of murine GRP78/BiP.

To investigate the role of each domain in BiP/GRP78 function, we have used a full-length recombinant BiP engineered to contain two enterokinase sites; one site is located after an N-terminal FLAG epitope, and a second site has been inserted at the junction between the N- and C-terminal domains (FLAG-BiP.ent). FLAG-BiP.

The mode of presentation and route of administration are critical for the induction of immune responses to p53 and antitumor immunity.

We have examined the immune response to full-length wild-type human p53 presented by a recombinant canarypox vector (ALVAC) and by plasmid DNA. For the ALVAC recombinant, intravenous, but not subcutaneous, intramuscular or intradermal administration, induced CD8+ CTLs that lysed tumor cells transfected with human mutant p53. Intrasplenic administration also induced CTLs.