A 92-kDa human immunostimulatory protein.

Fontan, Élisabeth; Briend, Emmanuel; Saklani-Jusforgues, Hélène; d’Alayer, Jacques; Vandekerckhove, Joël; Fauve, Robert M.

doi:10.1073/pnas.91.18.8353

Cited by 8 publications

(2 citation statements)

References 21 publications

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“…In SDS-PAGE gels, this apTHP migrates in the same position as the spTHP and, also, in an additional band that migrates ~10 kD farther. (In addition to apTHP, proteins with other functional activities have also been shown to fully share a primary peptide sequence with THP; these proteins, or subfractions of THP, were also isolated by methods other than salt precipitation [ 22 , 23 ].)…”

Section: Methodsmentioning

confidence: 99%

Surface Aggregation of Urinary Proteins and Aspartic Acid-Rich Peptides on the Faces of Calcium Oxalate Monohydrate Investigated by In Situ Force Microscopy

et al. 2009

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The growth of calcium oxalate monohydrate in the presence of Tamm-Horsfall protein (THP), osteopontin, and the 27-residue synthetic peptides (DDDS) 6 DDD and (DDDG) 6 DDD (D = aspartic acid, S = serine, and G = glycine) was investigated via in situ atomic force microscopy. The results show that these four growth modulators create extensive deposits on the crystal faces. Depending on the modulator and crystal face, these deposits can occur as discrete aggregates, filamentary structures, or uniform coatings. These proteinaceous films can lead to either the inhibition of or an increase in the step speeds (with respect to the impurity-free system), depending on a range of factors that include peptide or protein concentration, supersaturation, and ionic strength. While THP and the linear peptides act, respectively, to exclusively increase and inhibit growth on the " 101 ð Þface, both exhibit dual functionality on the (010) face, inhibiting growth at low supersaturation or high modulator concentration and accelerating growth at high supersaturation or low modulator concentration. Based on analyses of growth morphologies and dependencies of step speeds on supersaturation and protein or peptide concentration, we propose a picture of growth modulation that accounts for the observations in terms of the strength of binding to the surfaces and steps and the interplay of electrostatic and solvent-induced forces at the crystal surface.Keywords Biomineralization Á Crystal growth Á Kinetics Á Calcium oxalate monohydrate Á Tamm-Horsfall protein Á Scanning probe microscopy Á Kidney stones Á Osteopontin Although naturally occurring macromolecules in urine [1], such as osteopontin (OPN) and Tamm-Horsfall glycoprotein (THP), are believed to play an important role in controlling stone formation, a mechanistic understanding is missing. Numerous in vitro studies have shown that OPN strongly inhibits the nucleation and growth of calcium oxalate monohydrate-the most abundant mineral phase in urinary stones [2]. Stones are often found attached to epithelial cells and are typically aggregates of smaller crystals containing matrices of proteins, carbohydrates, and lipids [3]. A role for these macromolecular constituents in either promoting or inhibiting aggregation and/or attachment to cell membranes has been proposed [4,5]. Indeed, some in vitro studies have concluded that THP, which is the most common protein found in human urine [6] COM crystal growth [7,8] but acts to inhibit aggregation [9,10] or to limit the attachment of existing crystals to cell surfaces [11][12][13]. Others concluded that THP plays no role in COM growth [8]. A number of studies provide direct evidence that both OPN and THP themselves readily attach to the COM surface [3,14]. For example, using SDS-PAGE and Western blotting, Ryall et al. [15] observed that THP adhered to COM crystals taken from urine, and that ''scrupulous care must be taken to ensure the complete removal of extraneous THP adventitiously associated with CaOx crystals ….'' Using atomic force...

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Section: Methodsmentioning

confidence: 99%

Surface Aggregation of Urinary Proteins and Aspartic Acid-Rich Peptides on the Faces of Calcium Oxalate Monohydrate Investigated by In Situ Force Microscopy

et al. 2009

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“…Protein concentrations were determined either by the method of Bradford or by absorbance at 280 nm using an extinction coefficient of 0.352 unit⅐mg Ϫ1 ⅐cm 2 for rNEMO and of 0.181 unit⅐mg Ϫ1 ⅐cm 2 for the truncated rNEMO. Microsequencing of an internal peptide of the DnaK protein was performed as described previously (22), and the amino acid sequence comparisons were carried out using the protein data base Colibri.…”

Section: Methodsmentioning

confidence: 99%

NEMO Trimerizes through Its Coiled-coil C-terminal Domain

Agou¹,

Ye²,

Goffinont³

et al. 2002

Journal of Biological Chemistry

102

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NEMO/IB kinase (IKK)␥ is the regulatory component of the IKK complex comprising the two protein kinases, IKK␣ and IKK␤. To investigate the self-assembly properties of NEMO and to understand further the mechanism of activation of the IKK complex, we purified wildtype and mutant NEMO expressed in Escherichia coli. In the absence of its IKK partners, recombinant NEMO (rNEMO) is a metastable functional monomer correctly folded, according to its fluorescence and far-UV CD spectra, which is binding specifically to the IKK complex. A minor fraction of rNEMO was found tightly associated with DnaK (E. coli Hsp70). We also examined the interaction of NEMO with prokaryotic and eukaryotic Hsp70, and we showed that the Hsp70-NEMO complex forms a supramolecular structure probably corresponding to an assembly intermediate. Among the signaling pathways known to date, the transcription factor NF-B is one of the most intensely studied regulators of gene expression, playing a crucial role in inflammatory responses, cell proliferation, and apoptosis (1-4). NF-B transcription factors activate groups of genes in response to various stimuli, including the proflammatory cytokine tumor necrosis factor (TNF), 1 interleukin 1␤, bacterial lipopolysaccharide, and viral products. The most common form of NF-B transcription factor consists of RelA (p65) and p50 subunits (5, 6). In most non-induced cell types it is sequestered in the cytoplasm in an inactive state through its association with members of a family of inhibitory proteins known as IB. After cell stimulation, IB proteins are rapidly phosphorylated by the IB kinase complex (IKK) and are then degraded by the 26 S proteasome upon polyubiquitination (7). This degradation allows NF-B to move to the nucleus to switch on its target genes. Three components were identified as constituents of the IKK complex (Ϸ700 kDa): IKK␣, IKK␤, and NF-B essential modulator (NEMO, also called IKK␥). IKK␣ and IKK␤ sharing 52% identity possess a similar organization in functional domains including a kinase, a leucine zipper, and a helix-loop-helix domain (8 -11). Although in cells IKK␣ and IKK␤ are active both as homo-or as heterodimers promoted by their leucine zipper motifs, the heterodimer is the predominant form (12). The recent generation of IKK␣-and IKK␤-deficient mice showed different phenotypes assigning different functional roles to each catalytic subunit. Thus, whereas IKK␤ is required for the activation of NF-B, IKK␣ but not IKK␤ seems rather involved in keratinocyte differentiation (13).NEMO, the third component of the IKK complex, was originally identified by functional complementation of cells that did not respond to a variety of stimuli (14). It associates preferentially with IKK␤, and its presence is crucial for the stimuli-dependent activation of the IKK complex. NEMO, which has no known catalytic activity, contains at least four structural motifs as deduced from the primary structure analysis. The N-terminal domain contains a large coiled-coil domain (CC1, residues 93-231) carrying most of ...

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The anti-tumoral activity of human glycoprotein HGP.92: A study with the mouse Lewis-lung-tumor cell

et al. 1997

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The ability of a purified human glycoprotein (HGP.92) to exert anti-tumor activity was investigated in a mouse model using long-term readout assays. In vitro, in the presence of inflammatory mouse macrophages incubated with HGP.92, the growth of the mouse Lewis-lung-tumor cells (3LL) was decreased. This effect was concentration-dependent and required direct contact between tumor targets and HGP.92-treated macrophages. In addition, if the macrophage mono-layer was depleted of HGP.92 before addition of the target cells, no more cytostatic effect was observed. This anti-tumor activity of HGP.92-treated mouse macrophage was partially abrogated by addition of catalase in the culture medium, but not by superoxide dismutase or scavengers of the hydroxyl radical and singlet oxygen. Moreover, this tumor-cell growth reduction was not dependent on nitric oxide. In vivo, multiple i.v. injections of HGP.92 (5 times, 3 days apart) during the first week and a half exerted significant anti-tumor activity, as assessed by the reduction of both the number and the size of the lung nodules 3 weeks after i.v. inoculation of 3LL cells. Int. J. Cancer, 70:0-0, 1997. r 1997 Wiley-Liss, Inc. We have shown that a human glycoprotein of 92 kDa (HGP.92), isolated and purified from human urine, protected normal and SCID mice against an otherwise lethal inoculum of Listeria monocytogenes given intravenously; in addition, inflammatory murine macrophages incubated in vitro with HGP.92 exert anti-tumoral activity, as assessed by the growth reduction of 3LL cells (Fontan et al., 1994). Sequencing of 5 tryptic peptides of HGP.92 revealed complete homology with the amino-acid sequence of the Tamm Horsfall glycoprotein (THP), a urinary oligomeric glycopro-tein consisting of 80-to 100-kDa sub-units. Treatment of THP with urea yields a monomeric molecule of 92 kDa, suggesting that HGP.92 could be a sub-unit of THP. THP is known to be produced in the kidney, but its physiological function remains unclear (Sikri et al., 1981; Kumar et al., 1985). Some authors have described immunosuppressive activity in vitro (Brown et al., 1986; Hession et al., 1987), but some have reported immunostimulatory effects on human lymphocytes (Hunt and McGiven, 1978; Yu et al., 1993) and polymorphonuclear neutrophils (Horton et al., 1990; Yu et al., 1992). In our experimental models, native oligomeric THP did not enhance the resistance of mice infected with L. monocytogenes, and mouse macrophages incubated in vitro with THP were not cytostatic against 3LL cells. The increased resistance of HGP.92-treated SCID mice to L. monocytogenes infection, as well as the anti-tumoral activity of HGP.92-treated macrophages, provide some evidence that macro-phages may be one of the targets of HGP.92 biological activity. Many studies indicate that mouse macrophages can be activated to a tumoricidal state in vitro and in vivo by various cytokines and by non-cytokine molecules such as lipopolysaccharide (LPS) acting either alone or in synergism. We therefore monitored the ability of HGP.92 to...

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A 92-kDa human immunostimulatory protein.

Cited by 8 publications

References 21 publications

Surface Aggregation of Urinary Proteins and Aspartic Acid-Rich Peptides on the Faces of Calcium Oxalate Monohydrate Investigated by In Situ Force Microscopy

Surface Aggregation of Urinary Proteins and Aspartic Acid-Rich Peptides on the Faces of Calcium Oxalate Monohydrate Investigated by In Situ Force Microscopy

NEMO Trimerizes through Its Coiled-coil C-terminal Domain

The anti-tumoral activity of human glycoprotein HGP.92: A study with the mouse Lewis-lung-tumor cell

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