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M Morange

Publications and source records attributed to M Morange.

At least 37 records · Page 2Linked to original sources
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↗
PubMed1997

Evidence for the involvement of mouse heat shock factor 1 in the atypical expression of the HSP70.1 heat shock gene during mouse zygotic genome activation.

The mouse HSP70.1 gene, which codes for a heat shock protein (hsp70), is highly transcribed at the onset of zygotic genome activation (ZGA). This expression, which occurs in the absence of stress, is then repressed. It has been claimed that this gene does not exhibit a stress response until the blastocyst stage. The promoter of HSP70.1 contains four heat shock element (HSE) boxes which are the binding sites of heat shock transcription factors (HSF). We have been studying the presence and localization of the mouse HSFs, mHSF1 and mHSF2, at different stages of embryo development. We show that mHSF1 is already present at the one-cell stage and concentrated in the nucleus. Moreover, by mutagenizing HSE sequences and performing competition experiments (in transgenic embryos with the HSP70.1 promoter inserted before a reporter gene), we show that, in contrast with previous findings, HSE boxes are involved in this spontaneous activation. Therefore, we suggest that HSF1 and HSE are important in this transient expression at the two-cell stage and that the absence of typical inducibility at this early stage of development results mainly from the high level of spontaneous transcription of this gene during the ZGA.

Animals↗
PubMed1997

The transformation of molecular biology on contact with higher organisms, 1960-1980: from a molecular description to a molecular explanation.

The convergence of developmental biology--embryology--and molecular biology was one of the major scientific events of the last decades of the twentieth century. The transformation of developmental biology by the concepts and methods of molecular biology has already been described. Less has been told on the reciprocal transformation of molecular biology on contact with higher organisms. The transformation of molecular biology occurred at the end of a deep crisis which affected this discipline in the sixties and seventies and which led to a cruel criticism of the preexisting models of gene regulation. Numerous new, sometimes heterodox, models were proposed to describe the level at which gene regulation took place and its underlying mechanisms. The crisis resolved itself at the beginning of the eighties with the rapid accumulation of results from genetic engineering techniques and, above all, a displacement of the descriptive level from the molecule to the cell. This displacement gave molecular biologists the 'explanandum' which had been cruelly lacking during their initial study of higher organisms. The new molecular cell biology is an interfield explanation of living phenomena, relating a description and an interpretation localized at different levels of organization.

Embryology↗
PubMed1996

Low temperature enhancement of reporter genes expression directed by human immunodeficiency virus type 1 long terminal repeat.

Bacteria and eukaryotic cells respond to cold stress by inducing and enhancing the synthesis of specific arrays of proteins. We describe here cold-induced enhancement of expression for two reporter genes; luciferase and beta-galactosidase, both under the control of HIV-1 LTR sequences, observed in mouse fibroblasts and human HeLa cells respectively. Increased expression of luciferase in fibroblasts when shifted to 25 degrees C was detectable at 30 degrees C but was not observed following cold shock at 4 degrees C. To sustain the cold-induced effect, cells had to be kept at subphysiological temperature. The observed enhancement of luciferase activity did not result from a particular site of integration of the reporter gene and was evident whether cold-stressed cells were stationary or growing. Cold-induced expression of luciferase was evidenced at the protein level, enzymatic activity and RNA level, furthermore, active transcription and translation were required for overexpression. The cold effect which has been generalized with the reporter gene beta-galactosidase appears to be a process involving, at least in part, the HIV-1 LTR sequences and might correspond to an increase in the half-life of mRNA. The cold-dependent enhanced expression of luciferase and beta-galactosidase reported here, together with data describing the activation of HIV-1 LTR by hyperthermia, point out the particular temperature sensitivity of these regulatory sequences. This potential thermal modulation may be useful in the comprehension of regulatory processes in latency and reactivation of viral expression during HIV-1 infection.

3T3 Cells↗
PubMed1995

Different carboxyl-terminal domain kinase activities are induced by heat-shock and arsenite. Characterization of their substrate specificity, separation by Mono Q chromatography, and comparison with the mitogen-activated protein kinases.

In response to heat-shock and chemical treatments, cells undergo profound biochemical changes such as modifications in protein phosphorylation in order to resist the new, unfavorable growth conditions. We have previously shown that in HeLa cells a protein kinase (HS-CTD kinase) activity is induced rapidly after a heat or sodium arsenite shock. This kinase activity is able to phosphorylate a synthetic peptide composed of four repeats of the motif Ser-Pro-Thr-Ser-Pro-Ser-Tyr, a motif highly repeated in the carboxyl-terminal domain (CTD) of the largest subunit of eukaryotic RNA polymerase II. In this paper, we designed a new experimental procedure to characterize the substrate specificity of this kinase activity. We show that HS-CTD kinase activity phosphorylates a consensus sequence (-P-X-S/T-P-) which is similar to the sequence phosphorylated by extracellular regulated protein kinases (also called mitogen-activated protein kinases). However, there is a slight but reproducible difference between these kinases in their use of serine or threonine as the phosphate acceptor. Mono Q chromatography allows the separation of five stress-induced CTD kinase activities, two of which coelute with active mitogen-activated protein kinase forms revealed by Western blotting with anti ERK1-ERK2 antibodies. The other three CTD kinase activities induced after a stress are distinct from ERK1 and ERK2 and have different enzymatic properties. The molecular nature of these HS-CTD kinases and the physiological significance of their activation during stress remain to be determined.

Amino Acid Sequence↗
PubMed1994

A novel kinase cascade triggered by stress and heat shock that stimulates MAPKAP kinase-2 and phosphorylation of the small heat shock proteins.

MAPK-activated protein kinase-2 (MAPKAP kinase-2) is activated in vitro by the p42 and p44 isoforms of MAPK (p42/p44MAPK). In several cell lines, however, MAPKAP kinase-2 is activated by sodium arsenite, heat shock, or osmotic stress and not by agonists that activate p42/p44MAPK. We have identified a MAPK-like enzyme that acts as a MAPKAP kinase-2 reactivating kinase (RK). RK is recognized by an antiserum raised against a Xenopus MAPK (Mpk2), which is most similar to HOG1 from S. cerevisiae. We also identified a RK kinase (RKK) on the basis of its ability to activate either RK or a GST-Mpk2 fusion protein. The RKK, RK, and MAPKAP kinase-2 constitute a new stress-activated signal transduction pathway in vertebrates that is distinct from the classical MAPK cascade.

Amino Acid Sequence↗
PubMed1994

GRP78 induction by cyclosporin A in human HeLa cells.

Immunosuppressive drugs such as cyclosporin A (CsA) and FK506 are known to have pleiotropic effects on cells. Here we demonstrate that treatment of HeLa cells with low concentrations of CsA (but not of FK506) induces the synthesis of a stress protein, GRP78, located inside the endoplasmic reticulum. High concentrations of CsA lead to a general decrease in protein synthesis. When cells are stressed (heat-shocked) during the CsA treatment, the synthesis of heat shock proteins is reinforced. FK506 has no detectable effects at any concentration. The mechanism of induction of GRP78 by CsA remains presently unknown. Whatever the mechanism involved, GRP78 overexpression might be responsible for some of the physiological effects of CsA.

Carrier Proteins↗
PubMed1994

Detection of heat shock element-binding activities by gel shift assay during mouse preimplantation development.

Heat shock gene expression is regulated by highly conserved sequence elements (HSE for "heat shock elements"). Some of heat shock genes display an atypical expression during preimplantation mouse development. We have examined the profile of HSE-binding activities (HSE-BA) in matured ovulated oocytes and during the preimplantation development by gel shift assay and quantified the data by PhosphorImager. In each of our experiments, the F9 embryonal carcinoma cell line that contains both constitutive and heat-induced activity has been used as a control. We determine the number of oocytes or embryos required to get reproducible signals and accurate quantification by PhosphorImager. Oocytes, one-cell, and two-cell embryos respond to heat shock by inducing a strong HSE-BA. At the four-cell stage, no HSE-BA can be induced by heat shock, which suggests that noninducibility of heat shock genes at this stage (when the general mechanism of transcription is well established) might result from a defect in HSF or in the mechanism of HSF activation. A progressive reappearance of the ability to induce HSE-BA by stress is observed between the eight-cell stage and the blastocyst stage, and this parallels the appearance of heat shock gene inducibility. Matured ovulated oocytes and the first cleavage stages of embryos do not contain any HSE-BA at normal temperature but we observed a HSE-BA at normal temperature at the morula stage, which is increased at the blastocyst stage. These data, which, to our knowledge, for the first time describe the profile of a DNA-binding activity during the mouse preimplantation development, could serve as a basis for the study of other transcription factors during early embryogenesis.

Animals↗
PubMed1994

Heat shock factor 2-like activity in mouse blastocysts.

The expression of heat shock genes is induced in all living cells by a series of proteotoxic treatments. Heat shock genes are also activated spontaneously during different phases of embryonic development. HSP89 alpha and HSC70 are expressed at a high level in the mouse blastocyst. A family of factors, called HSFs, are able to bind to the promoters of heat shock genes on upstream conserved elements (HSEs). HSF1 is unable to bind to HSE sequences in absence of stress. It is activated after a stress by post-translational modifications and conformational change. HSF2 shows common structural domains with HSF1; however, it is active at normal temperatures. Recently we showed the presence of an abundant HSE-binding activity in nonshocked blastocysts. We demonstrate here by using polyclonal antibodies that HSF2 is the major constituent of this constitutive HSE-binding activity. HSF2 might be involved in the control of heat shock gene expression during early mammalian embryogenesis.

Animals↗
PubMed1993

Ubiquitin pathway involvement in human lymphocyte gamma-irradiation-induced apoptosis.

Apoptosis (the classical type of programmed cell death) can be triggered in many cell types by widely diverse stimuli. gamma rays, at low doses, can induce apoptosis in vitro in interphase human lymphocytes. In this type of apoptosis induction, activated gene expression is necessary for the fulfillment of the death program. In this report, we present evidence for a relationship between ubiquitin gene expression or ubiquitination and gamma-irradiation-mediated apoptosis in normal circulating human lymphocytes. Using in vitro nuclear transcription assays (run-on), Northern (RNA) blot analysis, immunolocalization studies, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis after immunoprecipitation, we demonstrate that (i) the ubiquitin mRNA level is increased as a consequence of the activation of ubiquitin gene transcription 15 to 90 min after initiation of apoptosis; (ii) specifically in apoptotic cells, and not in all irradiated cells, nuclear proteins are highly ubiquitinated; and (iii) ubiquitin sequence-specific antisense oligonucleotide inhibition results in a decreased level of ubiquitinated nuclear proteins and considerably diminishes the proportion of cells exhibiting the apoptotic death pattern. Each of these results might be explained by different modifications occurring in irradiated cells. Their convergence strongly suggests that the ubiquitin gene is one of the genes with induced activity in the apoptotic death program and that ubiquitination of nuclear proteins might be involved in chromatin disorganization and oligonucleosomal fragmentation, which are among the key events occurring in apoptosis.

Apoptosis↗
PubMed1993

The discovery of cellular oncogenes.

Between 1975 and 1985 a series of experiments demonstrated that cancer, whatever its causative agent, is due to the activation, by modification or overexpression, of a family of genes highly conserved during evolution, called the cellular oncogenes. These genes participate in the control of cell division in every living cell. Their products belong to the regulatory network relaying external signals from the membranes towards the nucleus and allowing cells to adapt their division rate to the demand of the organism. These discoveries constitute what may be called the 'oncogene paradigm'. Although the existence of cellular oncogenes, assumed in early models of oncogenesis, was demonstrated as early as 1976, we will show in this article that this discovery was not sufficient for the development of the new paradigm. We will describe its slow and complex formation between 1980 and 1985 followed by its rapid acceptance by the scientific community.

Animals↗
PubMed1992

Mammalian heat shock protein families. Expression and functions.

When prokaryotic or eukaryotic cells are submitted to a transient rise in temperature or to other proteotoxic treatments, the synthesis of a set of proteins called the heat shock proteins (hsp) is induced. The structure of these proteins has been highly conserved during evolution. The signal leading to the transcriptional activation of the corresponding genes is the accumulation of denatured and/or aggregated proteins inside the cells after stressful treatment. The expression of a subset of hsp is also induced during early embryogenesis and many differentiation processes. Two different functions have been ascribed to hsp: a molecular chaperone function: chaperones mediate the folding, assembly or translocation across the intracellular membranes of other polypeptides, and a role in protein degradation: some of the essential components of the cytoplasmic ubiquitin-dependent degradative pathway are hsp. These functions of hsp are essential in every living cell. They are required for repairing the damage resulting from stress.

Animals↗
PubMed1991

Heat shock increases turnover of 90 kDa heat shock protein phosphate groups in HeLa cells.

The 90 kDa heat shock protein (hsp90) is a major phosphoprotein which associates various other cellular polypeptides such as actin, calmodulin, steroid hormone receptors and certain protein-kinases. Little is known about the function of hsp90 in recovery from stress. In this report, we describe a dramatic increase in the rate of both phosphate uptake and dephosphorylation of hsp90 in HeLa cells submitted to acute stresses. This increased turnover of hsp90 phosphate groups might reflect a greater protein binding activity of hsp90 in stressed cells.

Carrier Proteins↗
PubMed1991

Denaturation of proteins during heat shock. In vivo recovery of solubility and activity of reporter enzymes.

Using beta-galactosidase and luciferase as reporter enzymes, we have previously shown that enzymatic inactivation occurring during a heat shock is concomitant with protein insolubilization (Nguyen, V. T., Morange, M., and Bensaude, O. (1989) J. Biol. Chem. 264, 10487-10492). In this paper, we observe that pretreatment of cells with D2O and glycerol, compounds known to stabilize protein structure, leads to a parallel decrease of protein inactivation and insolubilization, suggesting that these two phenomena result most probably from heat-induced protein denaturation. We found that heat shock-promoted inactivation and insolubilization are not irreversible processes, since even in the absence of protein synthesis, beta-galactosidase solubility and luciferase solubility and activity are recovered in vivo after a heat treatment. Cognate heat shock proteins might be involved in this renaturation process.

Animals↗
PubMed1991

[IIa/IIo conversion of RNA polymerase II during heat shock].

Heat-shock treatment of cells activates a protein kinase which phosphorylates a heptapeptide analogous to the repeated motif of the C-terminal domain (CTD) of the large subunit of RNA polymerase II from mammalian cells. This is corroborated with a modification of the large subunit of this enzyme during thermal stress in HeLa cells. We have observed a shift from the IIa form (unphosphorylated) to the IIo form (phosphorylated) with a higher apparent molecular weight, during a heat-shock at 44, 45 or 46 degrees C and by a chemical stress induced by sodium arsenite. RNA polymerase II hyperphosphorylation, together with the activation of the heat-shock transcription factor, might contribute to the onset of the preferential transcription of heat-shock genes.

Arsenates↗