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Les parents d’ailleurs, d’une autre culture, envisagent la difference et les troubles de leur enfant en lien avec leurs propres representations culturelles et familiales, tout en investissant la culture d’accueil et les soins de leur enfant. L’accueil de populations migrantes au CAMSP nous permet de reflechir a la place de la migration, de la perte de l’assise identitaire, langagiere et familiale, dans le lien etabli avec nous. Trois situations cliniques nous permettront de nous interroger sur la place et les effets des modalites d’annonce du diagnostic, sur nos propres representations culturelles et theoriques dans la construction du dispositif de soins, avec la question de l’alterite en arriere-plan.

The sericin 1 primary transcript of the silkworm Bombyx mori is differentially spliced via a tissue- and developmentally-regulated process. From a middle silk gland cDNA library, we have elucidated the sequence of one of the four mRNAs, the 4.0 kb Ser1B mRNA. Determination of alternative or constitutive exons and intron-exon boundaries allowed us to establish the nine exon-eight intron structure of the Ser1 gene. From these and previous data, it was possible to deduce the sequence of the sericins 1 and to predict the secondary structure and physiochemical properties of the different regions of the proteins.

International audience; Serial analysis of gene expression (SAGE) profiles, from posterior and median cells of the silk gland of Bombyx mori, were analyzed and compared, so as to identify their respective distinguishing functions. The annotation of the SAGE libraries was performed with a B. mori reference tag collection, which was extracted from a novel set of Bombyx ESTs, sequenced from the 3' side. Most of the tags appeared at similar relative concentration within the two libraries, and corresponded with region-specific and highly abundant silk proteins. Strikingly, in addition to tags from silk protein mRNAs, 19 abundant tags were found (≥0.1%), in the median cell library, which were absent in the posterior cell tag collection. With the exception of tags from SP1 mRNA, no PSG specific tags were found in this subset class. The analysis of some of the MSG-specific transcripts, suggested that middle silk gland cells have diversified functions, in addition to their well characterized role in silk sericins synthesis and secretion.

Three alleles of the sericin (Ser) 2-encoding gene (Ser2), called L, C and mC, were isolated from a Bombyx mori genomic library, and two related ones, called mCL and Cv, were also characterized in B. mori European strains. The Ser2 gene gives rise to two middle silk gland mRNAs by differential splicing. The size of a short mRNA (3.1 kb) is constant, but the length of a longer one ranges from 5 to 6.4 kb depending on the Ser2 allele. These length variations probably result from unequal recombinations in a region which contains about 30 well conserved 45-bp repeats coding for a Ser-like peptide. Furthermore, the L allele (and probably the mCL one) contains a 4.4-kb retrotransposon, resembling the copia-like ones of Drosophila.

We studied a protein from the midgut of the silkworm Bombyx mori characterized by its ability to bind the prosthetic group of chlorophyll, that confers fluorescent properties to this protein. Several techniques, 2D electrophoresis purification, MS-MS and Maldi-TOF peptide sequencing, RT-PCR and nucleotide sequencing were used to obtain the nucleotide sequence and the deduced amino acid sequence. The coding sequence was compared to the gene sequence to define the number and size of introns and exons. The gene spanned 45.5 kb of DNA and consisted of 46 exons. The cDNA encoded a protein of 2721 amino acids. The protein was identified as a lipocalin with novel features. Most lipocalins are proteins with high affinity to small lipophilic molecules, with a molecular size in the 25 kDa range and a well conserved tertiary structure. The apoprotein described here revealed 15 lipocalin like structures, in line. We called this protein a polycalin (pentadecacalin).

Mag is a retrotransposon found as an insert in the Sericin 2 gene. It is present in a few copies--4 to 15--dispersed in the genome of different strains of Bombyx mori as well as in Bombyx mandarina. Flanked by a 5 bp target sequence with no sequence specificity, it is bordered by direct repeats of 77 nucleotides. Despite their unusual short size, these terminal repeats and their immediately adjacent sequences present all the signals necessary for transcription into genomic RNA and for reverse transcription. Mag contains two overlapping open reading frames which are organized as the gag and pol genes of retroviruses and encode putative nucleic acid binding peptide, protease, reverse transcriptase, RNase H and endonuclease in this order. Sequence comparison of these proteins places mag within the gypsy group of LTR retrotransposons next to the echinoderm element SURL.

Spectroscopic studies (nuclear magnetic resonance, circular dichroism and infrared) have been carried out on chicken erythrocyte histone H5 and on three peptides cleaved therefrom: 1-31, 32-197 and 58-197. It is shown that at ionic strengths above o.1M part of the H5 molecule takes up a globular conformation containing 14% alpha helix but no beta sheet structure. Several details of the circular dichroism and nuclear magnetic resonace spectra indicate that the globular region is located in the N-terminal half of the molecule and this proposal is supported by the observation that the peptide 32-197 is largely incapable of folding and the peptide 59-197 is completely incapable of folding. Structural similarities and differences between histone H5 and histone H1 are discussed.

The sericins are a family of major cocoon proteins specifically synthesized in the middle silk gland of the silkworm Bombyx mori. The 5' part of one sericin gene had been cloned and described by Okamoto et al. (1982, J. Biol. Chem. 257, 15192-15199). Using a differential screening procedure of Bombyx genomic libraries, we obtained the 3' part of this gene. We demonstrate that it consists of a single gene extending over 24 kb, present in two allelic forms in hybrid strains. This gene encodes for four mRNAs which result from a unique transcript by an alternative splicing mechanism. This explains, at least partially, the diversity of the sericins found in the cocoon.

Poly (A)-containing RNA was isolated from the posterior silkgland of Bombyx mori just after the fourth molt, when the gland is not producing fibroin (stage V0), and at the middle of the fifth larval instar, when the fibroin is massively synthesized (stage V6). The hybridization kinetics of these mRNA with their complementary DNA (cDNA) were remarkably different. The mRNA from stage V0 consisted of two abundance classes, comprising 38 and 2 915 different species, respectively. At stage V6, fibroin mRNA (FmRNA) was separated from the rest of the poly (A)-containing RNA (F-mRNA) by centrifugation on sucrose gradients, and both preparations were analyzed separately. The kinetic complexity of FmRNA was very low as compared to its actual size. This result agrees with the existence of a short repetitive sequence accounting for 60 p. 100 of the molecule. Stage V6 F-mRNA was resolved into four classes containing 1, 20, 319 and about 2 600 species, respectively. We carried out cross-hybridization with each of the two cDNA classes from stage V0 and stage V6 F-mRNA. They demonstrated that almost all the sequences present at stage V0 were also present at stage V6, but that their abundance class distribution was modified. Our data show that the specialization of the silkgland cell for fibroin production is characterized by massive accumulation of FmRNA and another unknown species, and that stage variations of mRNA populations are related to relative quantitative variations rather than to qualitative ones.

Four mRNA of 10.5, 9.0, 4.0, and 2.8 kb are made from the sericin Ser1 gene by alternative maturation of a unique mRNA precursor. By means of RNA blots and in situ hybridization, we investigated variations in the distribution of these mRNA during the last larval instar in different territories of the middle silkgland. Taken together, the results from these two techniques show that 150 out of the 266 cells of this region of the organ express the Ser1 gene, but accumulate distinct mature mRNA species. Of these 150 cells 42 are specialized in a processing pathway resulting in the production of the 2.8-kb Ser1 mRNA throughout the larval instar. The 108 others perform successively three distinct splicing pathways leading to a development-dependent accumulation of, respectively, the 4.0-, the 10.5-, and the 9.0-kb mRNA. This suggests the occurrence of two switches in the splicing capacities of these cells during the fifth instar. The middle silkgland cells also express another sericin gene (Ser2) which encodes two mRNA of 5.4 and 3.1 kb, also arising by differential splicing. At the beginning of development, all the middle silkgland cells express this gene but, as development proceeds, expression becomes restricted to only the anterior cells. The biological consequence of this topological and temporal regulation of the mode of expression of these two genes is the sequential secretion and layering of the different sericins around the silk thread.

The conformation and stability of artificial complexes between chicken erythrocyte DNA and homologous histones FV and F2a2 was studied by circular dichroism (CD) and thermal denaturation followed by both absorbance and CD measurements. The complexes are made after a stepwise potassium fluoride gradient dialysis without urea and studied at low ionic strength (10−3 M). 1. 1) No structural changes of the DNA can be detected up to r = 0.2 with FV and r = 0.6 for F2a2. With FV at higher values of r the CD spectrum is altered, indicating the organization of DNA and histones in some kind of aggregate. 2. 2) The conformation of histone molecules inside the complexes is not related to the ionic strength of the medium but to an effective ionic environment close to 0.1 M. This ionic strength would also correspond to the melting temperature of histone-covered DNA. 3. 3) From the analysis of the absorbance melting profile the length of DNA covered with an histone molecule can be estimated. A good agreement is found between the negative charge of this piece of DNA and the net positive charge of the histone. 4. 4) Since the CD transition at 227 nm occurs before the second absorbance transition at 280 nm, the DNA is stabilized no longer by native histone but partially or fully denatured histones. The helical regions of the histone molecule are not involved in the binding process, which appears to be almost purely coulombian and most likely related to some structural fit between the pattern of negative charges in the DNA helix and that of positive charges along the peptide chain.

Analysis of the DNA of isolated nucleosomes suggests that virtually all genomic DNA sequences are organized in this basic chromatin subunit. In this report, we demonstrate that although histones reside on the transcriptionally active ovalbumin genes in the oviduct, the organization of proteins about this gene renders it highly sensitive to deoxyribonuclease I (deoxyribonucleate 5'-oligonucleotidohydrolase, EC 3.1.4.5). Treatment of oviduct nuclei from the laying hen with pancreatic deoxyribonuclease I results in the preferential digestion of over 70% of the ovalbumin sequences when only 10% of the total nuclear DNA has been solubilized. Treatment of liver nuclei does not reveal selective sensitivity of these genes to DNase I. Furthermore, regions of DNA not actively transcribed, such as the endogenous leukosis virus genes in the oviduct, are not selectively degraded by this enzyme. Similar digestions with micrococcal nuclease, however, reveal no specific digestion of transcriptionally active chromatin. These data confirm the observations of H. Weintraub and M. Groudine [(1976) Science 193, 848-856] and suggest we are dealing with an aspect of structure that may be necessary to permit transcription of the chromatin complex.

Galleria cocoon proteins have been extracted by different solubilizing agents. Nine protein bands were observed by gel electrophoresis, with molecular weights ranging from 18 to 420 kD. Three silk proteins of 24, 29 and 30 kD were extracted only in the presence of β-mercaptoethanol, suggesting that they are covalently linked by disulfide bonds to the large fibroin. They are likely to be the products of the highly abundant mRNA of the posterior silk gland cells. In vitro translation analysis of this mRNA yielded 24, 29 and 30 kD proteins. Thus, as in Bombyx , the Galleria silk is composed of several subunits, including fibroin and low molecular weight polypeptides. However, the genes coding for fibroin or low molecular weight silk proteins in Bombyx and Galleria do not show nucleotide base homology.

Summary After splitting the molecule of chicken erythrocyte specific histone (F v ) with cyanogen bromide we have isolated three fractions, by chromatography on a carboxy-methyl-cellulose column eluted with an ionic strength gradient, buffered at pH 4,3 : 2 N-terminal fractions A 1 and A 2 and the C-terminal fraction B. The results of amino-acid analysis and N-terminal determination of the tryptic peptides of the two N-terminal fractions are in complete agreement with the microheterogeneity which has been recently suggested by Greenaway and Murray : at amino-acid n o 15, a glutamyl residue in the fraction A 1 , and an arginyl residue in the fraction A 2 have been found.

Reconstituted nucleohistones were obtained by mixing in given conditions acid extracted histones and eukaryotic DNA. The histone/DNA ratio (w/w) was in the range 0.35 - 0.95. With the four histones (H2A2B) we have been able to obtain subunits (nucleosomes or upsilon-bodies). The variation of cirsular dichroism signal with temperature at 280 nm was measured to follow structural changes of the DNA inside the complex. The true change of ellipticity (see article) of histone-bound DNA regions, is similar for reconstituted nucleohistone and H1-depleted chromatin, and is therefore a physical probe of the presence of nucleosomes.

The characterization of a new silk protein mRNA (P25 mRNA) in posterior silkgland cells (PSG) and the developmental variations of its cell molecular concentration versus that of fibroin mRNA are described. A 80% pure P25 cDNA was obtained by class separation of total nonfibroin cDNA from PSG and used to identify the mRNA in blotted PSG mRNA as a single 1100 nucleotide long species. When purified from agarose gel and translated in a reticulocyte cell-free system, P25 mRNA yielded a 25-kD polypeptide (P25), identical to a 25-kD protein of the cocoon in terms of pI value and partial peptide mapping pattern. Moreover, this protein comigrated with an abundant polypeptide of the posterior silkgland (PSG) and of the middle silkgland (MSG). When tritiated leucine was injected in vivo, labeled P25 showed up in the PSG after a 2-hr pulse but appeared in the MSG only after 24 hr of labeling. Since MSG cells were found to be devoid of P25 mRNA, we concluded that P25 is exclusively synthesized in the PSG, that it accumulates in the MSG lumen and that it is spun out in the same way as fibroin. Specific probes were used to measure the concentrations of P25 mRNA and also fibroin mRNA in PSG total RNA by hybridization with an excess of cDNA. Both species are highly degraded in the few hours following the physiological arrest of feeding which precedes the fourth molting period. Their subsequent accumulation during the fifth intermolt is triggered by food uptake and proceeds in such a way that a constant 1:1 molar ratio is maintained during the period of silk secretion.

Poly(A) containing RNA was isolated from the posterior silk gland of Bombyx mori at the end of the fourth molt when the gland does not produce fibroin (stage V 0 ) and at the middle of the fifth larval instar when fibroin synthesis is massive (stage V 6 ). The kinetics of hybridization of these mRNAs with their complementary DNA (cDNA) were markedly different. The mRNA from V 0 stage consists of two abundance classes comprising, respectively, 38 and 2915 different sequences. V 6 mRNA-cDNA hybridization could not be analyzed properly by standard calculations since we found that the messenger RNA coding for fibroin (FmRNA) was about eight times less transcribed than the other mRNAs. This low yield of reverse transcription was assumed to be due to the exceptional length of FmRNA (16,000 nucleotides) as compared to the average length of the other silk gland mRNAs (1400 nucleotides). Therefore we separated the FmRNA from the rest of polyadenylated RNA (F ⊝ mRNA) by sucrose gradients and analyzed the two preparations separately. The kinetic complexity of FmRNA was very low for its size and can be accounted for by a 66-nucleotide-long driving sequence. This result agrees with the existence of a short repetitive sequence known to represent 60% of the molecule. The V 6 F ⊝ mRNA preparation was resolved into four classes containing, respectively, 1, 20, 319, and approximately 2600 sequences. Cross hybridizations have been performed with each of the cDNA abundance classes from V 0 stage and V 6 F ⊝ mRNA. They demonstrated that almost all the sequences present at the V 0 stage are also detected at V 6 stage, but their distribution among the abundance classes was modified. The relative concentration of 26 species of the abundant class and 291 species of the rare class of the V 0 stage was increased at V 6 and therefore appears to be implicated in the building of the fibroin synthesizing machinery, whereas 12 mRNA species from the V 0 abundant class decreased in relative concentration and are probably subject to another mode of control. Finally a large number of mRNA species remained in a stable low concentration at both stages and may be attributed to housekeeping functions. Our data show that the spectacular silk gland cell adaptation to fibroin production is characterized by massive accumulation of FmRNA and of an unknown mRNA species, and that stage variations in mRNA populations are related to relative quantitative variations rather than qualitative differences.

The influence of cortisol on the relation between plasma-free and platelet-bound serotonin in venous blood was examined in patients with rheumatoid arthritis treated with cortisone in excess and in patients not treated with cortisone, as well as in a normal person after i.v. injection of 50 mg cortisol. The correlation between ceruloplasmin and plasma-free or platelet-bound serotonin was investigated in patients with Wilson's disease. Cortisone significantly reduced the increased free serotonin level in rheumatoid arthritis. Patients with Wilson's disease had normal plasma-free serotonin levels, indicating that a low value of ceruloplasmin does not produce increased plasma-free serotonin levels. Platelet-bound serotonin values were diminished in these patients.