Neural cell adhesion molecules in rat endocrine tissues and tumor cells: distribution and molecular analysis

The adhesive properties of neural cell adhesion molecules (NCAMs) can be modified by alternative splicing of the primary transcript or posttranslational modifications. In the present study, we describe distinct forms of alternative splicing and posttranslational modification of the extracellular domain of NCAM of various endocrine tissues and derived tumor cells of the rat. Using an antiserum detecting the immunoglobulin-like domains of NCAM as well as a monoclonal antibody recognizing the NCAM-specific polysialic acid (PSA), we observed a similar staining pattern in adrenals, pituitary, and neoplastic endocrine cells. In endocrine tumor cells [pheochromocytoma (PC12), insulinoma (RINA2), and pituitary tumor cells (GH3)], NCAM immunoreactivity was most intense at contact sites between the cells. The immunocytochemical data were substantiated by results of in situ hybridization histochemistry. Specifically, higher levels of NCAM mRNA were detected in the adrenal cortex than in the medulla. In the pituitary, NCAM mRNA was more abundant in the anterior and intermediate lobes than in the neural lobe. The sequence of NCAM mRNAs in endocrine cells was analyzed by polymerase chain reaction and S1 nuclease protection assays. We found that major exons 4-13 of the NCAM mRNA in endocrine tissues and related tumor cell lines were homologous to those in the brain. However, PC12, RINA2, and GH3 tumor cells; normal rat pituitaries; and adrenals contained different amounts of NCAM mRNA with an alternative extra exon, termed VASE (also called pi in mouse) between constitutive exons 7 and 8. In addition, in pituitaries, we detected an alternative exon in splice site a between the constitutive exons 12 and 13, termed a15, with or without an AAG triplett. These sites are thought to be important for the adhesive properties of NCAM. Therefore, these results suggest that modifications of NCAM may be important for adhesive interactions in normal and neoplastic endocrine cells

Leydig cells express neural cell adhesion molecules in vivo and in vitro

The neural cell adhesion molecule (NCAM) polypeptides are expressed by numerous tissues during embryonic development, where they are involved in cell-cell interactions. In the adult, NCAM expression is confined to a few cell types, including neurons and peptide-hormone-producing cells. Here we demonstrate that the Leydig cells of the adult rat, mouse, and hamster testes express NCAM as well. Western blotting showed that an NCAM of approximately 120 kDa was present in the adult testes of all three species investigated. This form was also found in freshly isolated mouse Leydig cells and in Leydig cells after 2 days in culture. After 4 days in culture, mouse Leydig cells expressed additional NCAM isoforms of approximately 140 and 180 kDa, indicating changes in alternative splicing of NCAM primary transcripts. Also, NCAM mRNA of all isoforms, as detected by S1-nuclease protection assays, increased with time in culture. The expression of the cell adhesion molecule NCAM by adult Leydig cells may explain the aggregation of Leydig cells in clusters in rodent testes, which could be a prerequisite for functional coordination of groups of Leydig cells. Furthermore, the presence of this neural and endocrine marker may indicate a closer relationship between Leydig cells and neural and peptide-hormone-producing cells than is considered to exist at the present time

Insights into GABA receptor signalling in TM3 Leydig cells

gamma-Aminobutyric acid (GABA) is an emerging signalling molecule in endocrine organs, since it is produced by endocrine cells and acts via GABA(A) receptors in a paracrine/autocrine fashion. Testicular Leydig cells are producers and targets for GABA. These cells express GABA(A) receptor subunits and in the murine Leydig cell line TM3 pharmacological activation leads to increased proliferation. The signalling pathway of GABA in these cells is not known in this study. We therefore attempted to elucidate details of GABA(A) signalling in TM3 and adult mouse Leydig cells using several experimental approaches. TM3 cells not only express GABA(A) receptor subunits, but also bind the GABA agonist {[}H-3] muscimol with a binding affinity in the range reported for other endocrine cells (K-d = 2.740 +/- 0.721 nM). However, they exhibit a low B-max value of 28.08 fmol/mg protein. Typical GABA(A) receptor-associated events, including Cl- currents, changes in resting membrane potential, intracellular Ca2+ or cAMP, were not measurable with the methods employed in TM3 cells, or, as studied in part, in primary mouse Leydig cells. GABA or GABA(A) agonist isoguvacine treatment resulted in increased or decreased levels of several mRNAs, including transcription factors (c-fos, hsf-1, egr-1) and cell cycle-associated genes (Cdk2, cyclin D1). In an attempt to verify the cDNA array results and because egr-1 was recently implied in Leydig cell development, we further studied this factor. RT-PCR and Western blotting confirmed a time-dependent regulation of egr-1 in TM3. In the postnatal testis egr-1 was seen in cytoplasmic and nuclear locations of developing Leydig cells, which bear GABA(A) receptors and correspond well to TM3 cells. Thus, GABA acts via an untypical novel signalling pathway in TM3 cells. Further details of this pathway remain to be elucidated. Copyright (c) 2005 S. Karger AG, Base

Continuous Equilibrium in Affine and Information-Based Capital Asset Pricing Models

We consider a class of generalized capital asset pricing models in continuous time with a finite number of agents and tradable securities. The securities may not be sufficient to span all sources of uncertainty. If the agents have exponential utility functions and the individual endowments are spanned by the securities, an equilibrium exists and the agents' optimal trading strategies are constant. Affine processes, and the theory of information-based asset pricing are used to model the endogenous asset price dynamics and the terminal payoff. The derived semi-explicit pricing formulae are applied to numerically analyze the impact of the agents' risk aversion on the implied volatility of simultaneously-traded European-style options.Comment: 24 pages, 4 figure

Folding-competent and folding-defective forms of Ricin A chain have different fates following retrotranslocation from the endoplasmic reticulum

We report that a toxic polypeptide retaining the potential to refold upon dislocation from the endoplasmic reticulum (ER) to the cytosol (ricin A chain; RTA) and a misfolded version that cannot (termed RTAΔ), follow ER-associated degradation (ERAD) pathways in Saccharomyces cerevisiae that substantially diverge in the cytosol. Both polypeptides are dislocated in a step mediated by the transmembrane Hrd1p ubiquitin ligase complex and subsequently degraded. Canonical polyubiquitylation is not a prerequisite for this interaction because a catalytically inactive Hrd1p E3 ubiquitin ligase retains the ability to retrotranslocate RTA, and variants lacking one or both endogenous lysyl residues also require the Hrd1p complex. In the case of native RTA, we established that dislocation also depends on other components of the classical ERAD-L pathway as well as an ongoing ER–Golgi transport. However, the dislocation pathways deviate strikingly upon entry into the cytosol. Here, the CDC48 complex is required only for RTAΔ, although the involvement of individual ATPases (Rpt proteins) in the 19S regulatory particle (RP) of the proteasome, and the 20S catalytic chamber itself, is very different for the two RTA variants. We conclude that cytosolic ERAD components, particularly the proteasome RP, can discriminate between structural features of the same substrate

Study of ^194 Ir via thermal neutron capture and (d,p) reactions

Levels of ^194 Ir were studied using thermal neutron capture reaction. A pair spectrometer was used to measure the high-energy gamma-ray spectrum from thermal-neutron capture in enriched ^193 Ir target over the energy range 4640 - 6100 keV. The low-energy gamma-radiation from the reaction was studied with crystal diffraction spectrometers, and conversion electrons were observed with magnetic spectrometers. The high-sensitivity measurements at the Grenoble reactor, evaluated for transition energies up to 500 keV, are compared with lower-sensitivity measurements at the Wuerenlingen and Salaspils reactors. The comparison helped to obtain reliable isotopic identification for a number of ^194 Ir lines. The multipolarity admixtures for 29 gamma-transitions were determined on the basis of conversion lines from different electron subshells. Prompt and delayed gamma-gamma coincidences were measured using semiconductor and scintillation detectors. The ^193 Ir(d,p) high-resolution spectra, observed with a magnetic spectrometer, are given. All these data contributed to establishing a detailed level scheme of ^194 Ir. Additional data and the interpretation of the results in terms of current models will be presented in a forthcoming paper

Proučavanje 194Ir uhvatom termičkih neutrona I (d, p) reakcijom

Levels of 194Ir were studied using thermal neutron capture reaction. A pair spectrometer was used to measure the high-energy γ-ray spectrum from thermal-neutron capture in enriched 193Ir target over the energy range 4640 - 6100 keV. The low-energy γ-radiation from the reaction was studied with crystal diffraction spectrometers, and conversion electrons were observed with magnetic spectrometers. The high-sensitivity measurements at the Grenoble reactor, evaluated for transition energies up to 500 keV, are compared with lower-sensitivity measurements at the Wuerenlingen and Salaspils reactors. The comparison helped to obtain reliable isotopic identification for a number of 194Ir lines. The multipolarity admixtures for 29 γ-transitions were determined on the basis of conversion lines from different electron subshells. Prompt and delayed γ-γ coincidences were measured using semiconductor and scintillation detectors. The 193Ir(d,p) high-resolution spectra, observed with a magnetic spectrometer, are given. All these data contributed to establishing a detailed level scheme of 194Ir. Additional data and the interpretation of the results in terms of current models will be presented in a forthcoming paper.Proučavala su se stanja u 194Ir reakcijama 193Ir(n, γ) i 193Ir(d, p). Mjerenja uhvata termičkih neutrona načinjena su uz reaktore u Grenoblu, Wuerenlingenu i Salapsisu. Za mjerenja γ-zračenja visoke energije upotrebljavao se spektrometar parova, a za niske energije difraktometar. Konverzijske elektrone se mjerilo magnetskim spektrometrom. Mjerenja reakcije (d, p) visokog razlučivanja izvedena su magnetskim spektrometrom. Usporedbe tih mjerenja omogućile su pouzdano izotopno prepoznavanje prijelaza u 194 Ir, a spektri konverzijskih elektrona i određivanje multipolnosti prijelaza. Dobiveni su podaci osnova sheme raspada 194Ir