A mouse pluripotent embryonal stem cell line stage-specifically regulates expression of homeo-box containing DNA sequences during differentiation in vitro

Mouse embryonal stem (ES) cells have been shown to provide a new model system suitable for the analysis of different aspects of murine development. This report gives evidence that ES cell lines are also most useful for the study of developmentally regulated gene expression in vitro. Homeo-box containing genes which are suggested to play a key role in the regulation of differentiation steps occurring during embryogenesis are stage-specifically transcribed in differentiating murine ES cells: (i) A mouse embryonal stem cell line (ES-12957) was isolated and characterized with respect to its differentiation potential. When injected subcutaneously into syngeneic mice, ES-12957 cells formed fully differentiated teratomas representing derivatives of all three germ layers. When allowed to grow in suspension cultures in vitro, the cells followed a reproducible developmental pathway forming complex organized 'embryoid bodies' which resembled mouse early postimplantation embryos. (ii) A mouse DNA sequence with homeo-box homology (MH-121) was isolated and structurally analyzed. Transcription of a 1.7 kb RNA species from this DNA sequence was demonstrated in ES-12957 cells which were differentiated in vitro. A second, previously described homeo-box gene (Mo-10) was also shown to be expressed in ES-12957 cells in a stage-specific manner. A 4-kb transcript could be identified exclusively in RNA of cells which were allowed to differentiate for 9 days. These findings support the suggestion that the homeo-box genes of mammals, like those of Drosophila, may have important functions during embryonic development

Competitive antagonism by phenylglycine derivatives at type 1 metabotropic glutamate receptors

The metabotropic glutamate receptors (mGluRs) form a family of G-protein- coupled receptors which consists of at least seven members termed mGluR1- mGluR7. These members are classified into subfamilies according to their sequence similarities, signal transduction mechanisms and agonist selectivities. mGluR1 and mGluR5 are coupled to the phosphoinositide hydrolysis/Ca2+ signal transduction and efficently respond to quisqualate. In this study, we have stably expressed mGluR1 in Chinese hamster ovary cells on which the activation of the phosphoinositide signal transduction pathway was evaluated by means of two methods and their degree of correspondence was analyzed. These two methods involve the Li+-dependent accumulation of [3H]inositol-labeled inositol phosphates or the [3H]cytidine-labeled phospholiponucleotide cytidine diphospho (CDP)- diacylglycerol (DAG). The correlation between the two measures was found to be generally uniform for the different agonists evaluated. However, the levels of CDP-DAG were found to be consistently higher. Furthermore, quisqualate showed a differential activity on the two methods behaving as a partial agonist and as a full agonist on the inositol phosphate and the CDP-DAG responses, respectively. On the same cells the activity of a series of carboxyphenylglycines recently described as possible new tools for investigating the role of mGluRs has been evaluated. Three phenylglycine derivatives were tested and found to be competitive antagonists at this mGluR subtype. They inhibited both the phosphoinositide signal transduction pathway and the release of intracellular Ca2+ induced by quisqualate the most potent agonist at mGluR1. The pharmacological nature of these compounds and their relative potencies in antagonizing mGluR1 activation are described. © 1994 Academic Press, Inc

Chromosomal mapping of the structural gene coding for the mouse cell adhesion molecule uvomorulin.

The gene coding for the mouse cell adhesion molecule uvomorulin has been mapped to chromosome 8. Uvomorulin cDNA clone F5H3 identified restriction fragment length polymorphisms in Southern blots of genomic DNA from mouse species Mus musculus domesticus and Mus spretus. By analyzing the segregation pattern of the gene in 75 offspring from an interspecific backcross a single genetic locus, Um, was defined on chromosome 8. Recombination frequency between Um and the co-segregating loci serum esterase 1 (Es-1) and tyrosine aminotransferase (Tat) places Um about 14 centimorgan (cM) distal to Es-1, and 5 cM proximal to Tat. In situ hybridization of uvomorulin [3H]cDNA to mouse metaphase chromosomes located the Um locus close to the distal end of chromosome 8 (bands C3-E1). Since uvomorulin is evolutionarily highly conserved, its chromosomal assignment adds an important marker to the mouse genetic map

The structure of cell adhesion molecule uvomorulin. Insights into the molecular mechanism of Ca2+-dependent cell adhesion

We have determined the amino acid sequence of the Ca2+-dependent cell adhesion molecule uvomorulin as it appears on the cell surface. The extracellular part of the molecule exhibits three internally repeated domains of 112 residues which are most likely generated by gene duplication. Each of the repeated domains contains two highly conserved units which could represent putative Ca2+-binding sites. Secondary structure predictions suggest that the putative Ca2+-binding units are located in external loops at the surface of the protein. The protein sequence exhibits a single membrane-spanning region and a cytoplasmic domain. Sequence comparison reveals extensive homology to the chicken L-CAM. Both uvomorulin and L-CAM are identical in 65% of their entire amino acid sequence suggesting a common origin for both CAMs

The structure of cell adhesion molecule uvomorulin. Insights into the molecular mechanism of Ca2+-dependent cell adhesion.

We have determined the amino acid sequence of the Ca2+-dependent cell adhesion molecule uvomorulin as it appears on the cell surface. The extracellular part of the molecule exhibits three internally repeated domains of 112 residues which are most likely generated by gene duplication. Each of the repeated domains contains two highly conserved units which could represent putative Ca2+-binding sites. Secondary structure predictions suggest that the putative Ca2+-binding units are located in external loops at the surface of the protein. The protein sequence exhibits a single membrane-spanning region and a cytoplasmic domain. Sequence comparison reveals extensive homology to the chicken L-CAM. Both uvomorulin and L-CAM are identical in 65% of their entire amino acid sequence suggesting a common origin for both CAMs