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Biomedical subjects

M Morange

Publications and source records attributed to M Morange.

At least 19 recordsLinked to original sources
PubMed2001

The bacterial nucleoside N(6)-methyldeoxyadenosine induces the differentiation of mammalian tumor cells.

Contrary to bacterial DNA, mammalian DNA contains very little if any N(6)-methyldeoxyadenosine (MDA). The possible biological effect of this nucleoside on eukaryotic cells has been studied on different tumor cell lines. Addition of MDA to C6.9 glioma cells triggers a differentiation process and the expression of the oligodendroglial marker 2',3'-cyclic nucleotide 3'phosphorylase (CNP). The biological effects of N(6)-methyldeoxyadenosine were not restricted to C6.9 glioma cells since differentiation was also observed on pheochromocytoma and teratocarcinoma cell lines and on dysembryoplastic neuroepithelial tumor cells. The precise mechanism by which MDA induces cell differentiation remains unclear, but is related to cell cycle modifications. These data point out the potential interest of N(6)-methyldeoxyadenosine as a novel antitumoral and differentiation agent. They also raise the intriguing question of the loss of adenine methylation in mammalian DNA. Furthermore, the finding that a methylated nucleoside found in bacterial DNA induces a biological process might have implications in gene therapy approaches when plasmid DNAs are injected into humans.

2',3'-Cyclic Nucleotide 3'-Phosphodiesterase↗
PubMed2001

Overexpression of murine small heat shock protein HSP25 interferes with chondrocyte differentiation and decreases cell adhesion.

Although multiple functions for the small heat shock protein HSP25 have been proposed, its specific role during developmental and differentiation processes is not known. Cartilage is one of the tissues in which HSP25 is specifically and highly expressed during development. C1 cells, able to form aggregates in vitro, can be induced to differentiate into chondrocytes. In this study, we generated two stable transfected clones overexpressing HSP25 at two different levels. Cell morphology and growth rate were modified in both clones, although the actin content and distribution did not seem to be altered. Overexpressing clones had more difficulties in coalescing, leading to smaller aggregates and they did not differentiate into chondrocytes. Subsequently, these aggregates tended to dissociate into loose masses of dying cells. The strength of all these effects was directly correlated to the level of HSP25 overexpression. These data suggest that overexpressing HSP25 decreases cellular adhesion and interferes with chondrocyte differentiation.

Actin Cytoskeleton↗
PubMed2000

Cyclosporin A induces an atypical heat shock response.

Cyclosporin A is a widely used immunosuppressive drug having toxic side effects, in particular on kidneys and liver, as a result of its action on different molecular targets. Here we demonstrate that low doses of CsA are able to induce the expression of the heat shock protein HSP27 and its hyperphosphorylation. It also activates the two heat shock transcription factors, HSF1 and HSF2. Since these factors have been shown to be activated by proteasome inhibition, we tested the hypothesis that the inhibitory action of CsA on the proteasome might be responsible for the activation of HSFs and the subsequent expression of HSP27. The increase in multiubiquitinated proteins as well as the stabilization of p53 following CsA addition argues in favor of this hypothesis. The kidney BSC-1 cells are highly responsive to the addition of CsA: the possible link between HSP27 induction and hyperphosphorylation and nephrotoxicity is discussed.

Animals↗
PubMed2000

Hsp25 and the p38 MAPK pathway are involved in differentiation of cardiomyocytes.

The small heat-shock protein HSP25 is expressed in the heart early during development, and although multiple roles for HSP25 have been proposed, its specific role during development and differentiation is not known. P19 is an embryonal carcinoma cell line which can be induced to differentiate in vitro into either cardiomyocytes or neurons. We have used P19 to examine the role of HSP25 in differentiation. We found that HSP25 expression is strongly increased in P19 cardiomyocytes. Antisense HSP25 expression reduced the extent of cardiomyocyte differentiation and resulted in reduced expression of cardiac actin and the intermediate filament desmin and reduced level of cardiac mRNAs. Thus, HSP25 is necessary for differentiation of P19 into cardiomyocytes. In contrast, P19 neurons did not express HSP25 and antisense HSP25 expression had no effect on neuronal differentiation. The phosphorylation of HSP25 by the p38/SAPK2 pathway is known to be important for certain of its functions. Inhibition of this pathway by the specific inhibitor SB203580 prevented cardiomyocyte differentiation of P19 cells. In contrast, PD90589, which inhibits the ERK1/2 pathway, had no effect. Surprisingly, cardiogenesis was only sensitive to SB203580 during the first 2 days of differentiation, before HSP25 expression increases. In contrast to the effect of antisense HSP25, SB203580 reduced the level of expression of the mesodermal marker Brachyury-T during differentiation. Therefore, we propose that the p38 pathway acts on an essential target during early cardiogenesis. Once this initial step is complete, HSP25 is necessary for the functional differentiation of P19 cardiomyocytes, but its phosphorylation by p38/SAPK2 is not required.

Calcium-Calmodulin-Dependent Protein Kinases↗
PubMed2000

Gene function.

The problem of gene function--of the relationships between hereditary material and the characteristics of organisms--preceded the rediscovery of Mendel's laws and accompanied the development of genetics in the XXth century. Molecular biologists replaced the simple gene-character relationship by two relationships: the first, between genes and proteins, was well defined, whereas the second between proteins and the complex structural and functional characteristics of organisms remained unknown. I will describe in this article the experimental approaches which helped to characterize during the last twenty years the relationships between proteins and characters. Four principles of macromolecular organization emerged from these studies: conservation of the elementary components during evolution, existence of pathways and networks, pleiotropy and redundancy. These principles are the explanation of the surprising experimental observations that have been made in recent years. The existence of these principles makes problematic any prediction on the consequences of gene modification. It both sounds the death-knell of the simplistic reductionist approach of many biologists, whereby genes were considered as responsible for specific functions, and definitely prevents the distribution of genes in separate, well defined categories.

Animals↗
PubMed2000

Heat shock and arsenite induce expression of the nonclassical class I histocompatibility HLA-G gene in tumor cell lines.

The nonclassical histocompatibility class I gene HLA-G has a tissue-restricted expression. To explore mechanisms involved in HLA-G transcriptional regulation, we have investigated the effect of stress, including heat shock and arsenite treatment, on HLA-G expression in tumor cell lines. We show that stress induces an increase of the level of the different HLA-G alternative transcripts without affecting other MHC class I HLA-A, -B, -E, and -F transcripts. A heat shock element (HSE) that binds to heat shock factor 1 (HSF1) on stress conditions was further identified within the HLA-G promoter. Considering the ability of HLA-G to modulate the function of immunocompetent cells, we hypothesize a new feature of HLA-G as a signal regulating the immune response to stress.

Arsenites↗
PubMed2000

The distribution of heat shock proteins in the nervous system of the unstressed mouse embryo suggests a role in neuronal and non-neuronal differentiation.

Heat shock proteins (Hsps) act as molecular chaperones and are generally constitutively expressed in the absence of stress. Hsps are also inducible by a variety of stressors whose effects could be disastrous on the brain. It has been shown previously that Hsps are differentially expressed in glial and neuronal cells, as well as in the different structures of the brain. This differential expression has been related to specific functions distinct from their general chaperone function, such as intracellular transport. We investigated here the constitutive expression of 5 Hsps (the small Hsp, Hsp25, the constitutive Hsc70 and Hsp90beta, the mainly inducible Hsp70 and Hsp90alpha), and of a molecular chaperone, TCP-1alpha during mouse nervous system development. We analyzed, by immunohistochemistry, their distribution in the central nervous system and in the ganglia of the peripheral nervous system from day 9.5 (E9.5) to day 17.5 (E17.5) of gestation. Hsps are expressed in different cell classes (neuronal, glial, and vascular). The different proteins display different but often overlapping patterns of expression in different regions of the developing nervous system, suggesting unique roles at different stages of neural maturation. Their putative function in cell remodeling during migration or differentiation and in protein transport is discussed. Moreover we consider Hsp90 function in cell signaling and the role of Hsp25 in apoptosis protection.

Animals↗
PubMed2000

François Jacob's lab in the seventies: the T-complex and the mouse developmental genetic program.

The existence of a genetic program of development was proposed by molecular biologists in the nineteen-sixties. Historians and philosophers of science have since thoroughly criticized this notion. To fully appreciate its significance, it is interesting to consider the research which was pursued during this period by molecular biologists who proposed this notion. This study focuses on François Jacob's work and on the model of development supported by his lab in the early seventies, the T-complex model. This episode of Jacob's scientific activity has since been forgotten. Characterization of this model shows that the notion of program was used in a metaphoric way and that it did not put any constraint on the work pursued in the lab at that time. Some attention is devoted to the origin of this metaphor in the context of the nineteen-seventies.

Animals↗
PubMed1999

Genomic structure and chromosomal localization of the mouse Hsf2 gene and promoter sequences.

The mouse heat shock factor 2 (HSF2) cDNA was previously cloned by homology to HSF1, the heat shock factor involved in the cellular response to stress [Sarge, K.D., Zimarino, V., Holm, K., Wu, C., Morimoto, R.I., Cloning and characterization of two mouse heat shock factors with distinct inducible and constitutive DNA-binding ability. Genes Dev. 5 (1991) 1902-1911]. HSF2 is active in restricted cell types during pre- and post-implantation stages of development, and only in male germ cells of adult mice. However, the function of this factor remains elusive. We report here the cloning of the mouse Hsf2 gene and its genomic structure. We show that the gene is composed of 13 exons of variable sizes spanning at least 43kb in the genome. The transcription start site has been determined, and upstream sequences with promoter activity have been identified by their ability to direct the expression of a luciferase reporter gene in transfected cells. A preliminary analysis of the proximal promoter sequence determined that the TATA box is absent, but that a GC-rich region with several potential binding sites for transcription factors is present. The gene has been mapped to mouse chromosome 10 by in-situ hybridization on metaphase chromosomes.

Animals↗
PubMed1999

[Temporal regulation of gene expression].

Molecular biology gives a static--not a dynamic--vision of the mechanisms regulating gene expression. Genetics already gave to time a limited place in the explanation of living phenomena. Such a static vision is supported by the techniques--such as X-ray crystallography--used by the biologists. However time is an important parameter in the control of gene expression during the cellular response to external signals, during life and aging of organisms or even in the succession of living forms which takes place in evolution. Models are slowly moving, due to the eruption of new technologies giving access to the fast events which occur inside living cells. A new dynamic vision is progressively replacing the old one. The consequences of these changes on the form of the future biology remain still unknown.

Aging↗
PubMed1998

Characterization of the residues phosphorylated in vitro by different C-terminal domain kinases.

The C-terminal part of the largest subunit of eukaryotic RNA polymerase II is composed solely of the highly repeated consensus sequence Tyr1-Ser2-Pro3-Thr4-Ser5-Pro6-Ser7. This domain, called the C-terminal domain (CTD), is phosphorylated mostly at serine residues during transcription initiation, but the precise role of this phosphorylation remains controversial. Several protein kinases are able to phosphorylate this sequence in vitro. The aim of this work was to define the positions of the amino acids phosphorylated by four of these CTD kinases (transcription factor (TF) IIH-kinase, DNA-dependent protein kinase, and the mitogen-activated protein kinases ERK1 and ERK2) and to compare the specificity of these different protein kinases. We show that TFIIH kinase and the mitogen-activated protein kinases phosphorylate only serine 5 of the CTD sequence, whereas DNA-dependent protein kinase phosphorylates serines 2 and 7. Among the different CTD kinases, only TFIIH kinase is appreciably more active on two repeats of the consensus sequence than on one motif. These in vitro results can provide some clues to the nature of the protein kinases responsible for the in vivo phosphorylation of the RNA polymerase CTD. In particular, the ratio of phosphorylated serine to threonine observed in vivo cannot be explained if TFIIH kinase is the only protein kinase involved in the phosphorylation of the CTD.

Amino Acid Sequence↗
PubMed1998

Hsp and chaperone distribution during endochondral bone development in mouse embryo.

The process of endochondral bone formation was examined with regard to expression of seven heat shock proteins (Hsps): two small Hsps, the constitutive and the inducible forms of the 70 and the 90 Hsp families, the collagen chaperone Hsp47, and a cytosolic chaperone, TCP-1alpha, using immunohistochemistry. Around day 15.5 of embryogenesis the calcification of the long endochondral bones occurs through progressive replacement of the cartilaginous scaffold (rich in type II collagen) with an ossified matrix (rich in type I collagen), and thus a longitudinal section of limb bone recapitulates all the steps of chondrogenesis and the early steps of osteogenesis. We observed that all these Hsps and chaperones are differentially expressed during bone development in a stage-specific pattern reaching very high levels at some specific stages. The involvement of chaperones during these important differentiation steps will be discussed.

Animals↗
PubMed1997

MAP kinase activation by cyclosporine A.

Short treatment of HeLa cells with cyclosporine A led to the activation in the crude cell extracts of a MAP kinase-like activity. Fractionation by chromatography on a Mono Q column allowed the separation of two activities co-eluting with the MAP kinases ERK1 and ERK2. The activation of these two MAP kinases was demonstrated in Western Blotting by the appearance, after CsA treatment, of two new slowly migrating forms on SDS electrophoretic gels. A similar activation was also obtained in renal epithelial BSC-1 cells and 3T3 fibroblasts. MAP kinase activation might result from a perturbation of calcium homeostasis induced by CsA treatment.

Blotting, Western↗
PubMed1997

Function and regulation of heat shock factor 2 during mouse embryogenesis.

The spontaneous expression of heat shock genes during development is well documented in many animal species, but the mechanisms responsible for this developmental regulation are only poorly understood. In vertebrates, additional heat shock transcription factors, distinct from the heat shock factor 1 (HSF1) involved in the stress response, were suggested to be involved in this developmental control. In particular, the mouse HSF2 has been found to be active in testis and during preimplantation development. However, the role of HSF2 and its mechanism of activation have remained elusive due to the paucity of data on its expression during development. In this study, we have examined HSF2 expression during the postimplantation phase of mouse development. Our data show a developmental regulation of HSF2, which is expressed at least until 15.5 days of embryogenesis. It becomes restricted to the central nervous system during the second half of gestation. It is expressed in the ventricular layer of the neural tube which contains mitotically active cells but not in postmitotic neurons. Parallel results were obtained for mRNA, protein, and activity levels, demonstrating that the main level of control was transcriptional. The detailed analysis of the activity of a luciferase reporter gene under the control of the hsp70.1 promoter, as well as the description of the protein expression patterns of the major heat shock proteins in the central nervous system, show that HSF2 and heat shock protein expression domains do not coincide. This result suggests that HFS2 might be involved in other regulatory developmental pathways and paves the way to new functional approaches.

Animals↗
PubMed1997

HSP gene expression and HSF2 in mouse development.

During the pre-implantation phase of development, the mouse embryo synthesizes HSC70, and HSP90 alpha and beta at a very high rate. After implantation, the expression of HSPs appears non-coordinated and is not uniform in the different tissues. The expression of inducible HSPs appears later in development than that of constitutive members of the family. HSP25 is highly expressed early in heart and muscle development, but also in some structure of the central nervous system. HSC70 and HSP90 beta are expressed ubiquitously, but their expression reaches very high levels in the nervous system (neural tracks) and during bone morphogenesis (in the hypertrophic chondrocytes). The mechanisms involved in HSP expression during mouse embryogenesis are probably diverse, involving tissue-specific sequences. Although the DNA-binding activity and expression of the second heat shock transcription factor, HSF2, seems to be developmentally regulated, becoming detectable at the blastocyst stage and reaching a peak at day 10 of development, there is no obvious correlation between the level of this factor and the expression of HSPs. HSF2 might be involved in the onset of expression of HSPs, regulate (inhibit) their expression, or control the expression of other developmental genes yet to be discovered.

Animals↗