Adenylate cyclase activation and its effects on intracellular cAMP in infected KB cells during trypsin-induced infectious Senda? virus production (author's transl)]

KB cells infected by Senda? virus can produce infectious virus if they are trypsinated twice over 24 h. Adenylate cyclase activity in infected KB cells is higher and more strongly activated by trypsin than that of control cells, but intracellular concentration of cAMP is the same, except during a short time after trypsinations, especially after the second trypsination which causes infectious virus production. During this short time, intracellular cAMP is slightly higher in infected cells. This miseffect of adenylate cyclase activation on intracellular cAMP concentrations might be related to an increased cell permeability caused by trypsin.     

Structural disorder in plant proteins: where plasticity meets sessility

Plants are sessile organisms. This intriguing nature provokes the question of how they survive despite the continual perturbations caused by their constantly changing environment. The large amount of knowledge accumulated to date demonstrates the fascinating dynamic and plastic mechanisms, which underpin the diverse strategies selected in plants in response to the fluctuating environment. This phenotypic plasticity requires an efficient integration of external cues to their growth and developmental programs that can only be achieved through the dynamic and interactive coordination of various signaling networks. Given the versatility of intrinsic structural disorder within proteins, this feature appears as one of the leading characters of such complex functional circuits, critical for plant adaptation and survival in their wild habitats. In this review, we present information of those intrinsically disordered proteins (IDPs) from plants for which their high level of predicted structural disorder has been correlated with a particular function, or where there is experimental evidence linking this structural feature with its protein function. Using examples of plant IDPs involved in the control of cell cycle, metabolism, hormonal signaling and regulation of gene expression, development and responses to stress, we demonstrate the critical importance of IDPs throughout the life of the plant.     

Crystal structure and biochemical characterization of the transmembrane PAP2 type phosphatidylglycerol phosphate phosphatase from <Emphasis Type="Italic">Bacillus subtilis</Emphasis>

Type 2 phosphatidic acid phosphatases (PAP2s) can be either soluble or integral membrane enzymes. In bacteria, integral membrane PAP2s play major roles in the metabolisms of glycerophospholipids, undecaprenyl-phosphate (C₅₅-P) lipid carrier and lipopolysaccharides. By in vivo functional experiments and biochemical characterization we show that the membrane PAP2 coded by the Bacillus subtilis yodM gene is the principal phosphatidylglycerol phosphate (PGP) phosphatase of B. subtilis. We also confirm that this enzyme, renamed bsPgpB, has a weaker activity on C₅₅-PP. Moreover, we solved the crystal structure of bsPgpB at 2.25 Å resolution, with tungstate (a phosphate analog) in the active site. The structure reveals two lipid chains in the active site vicinity, allowing for PGP substrate modeling and molecular dynamic simulation. Site-directed mutagenesis confirmed the residues important for substrate specificity, providing a basis for predicting the lipids preferentially dephosphorylated by membrane PAP2s.     

Mise en évidence d'une activitéα-amylasique dans les lysosomes d'Aspergillus oryzae

Summary The -amylase of mycelial cells ofAspergillus oryzae exists in a particular form in 8000 g pellet. The lysosomal localization of acid phosphatase is confirmed by electron microscopy. The purification of lysosomes by discontinuous gradient of sucrose in D₂O shows that -amylase activity is bound to these particles.     

Place de la prolyl hydroxylase dans la synthèse du collagène de cellules KB

Summary KB cells, which synthetized collagen at a low rate, shown a prolyl hydroxylase activity at the same rate that fibroblast. The relationship between collagen synthesis and prolyl hydroxylase activity in these cells was discussed.

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Ce travail a bénéficié d'une aide du CNRS (AI n^o 031218 et RCP n^o 08533) et de l'U.E.R. de Biologie Humaine de l'Université de Lyon.     

Behaviour of the prolyl hydroxylase to collagen synthesis in KB cells]

KB cells, which synthetized collagen at a low rate, shown a prolyl hydroxylase activity at the same rate that fibroblast. The relationship between collagen synthesis and prolyl hydroxylase activity in these cells was discussed.     

Targeted gene correction of α1-antitrypsin deficiency in induced pluripotent stem cells

Human induced pluripotent stem cells (iPSCs) represent a unique opportunity for regenerative medicine because they offer the prospect of generating unlimited quantities of cells for autologous transplantation, with potential application in treatments for a broad range of disorders. However, the use of human iPSCs in the context of genetically inherited human disease will require the correction of disease-causing mutations in a manner that is fully compatible with clinical applications. The methods currently available, such as homologous recombination, lack the necessary efficiency and also leave residual sequences in the targeted genome. Therefore, the development of new approaches to edit the mammalian genome is a prerequisite to delivering the clinical promise of human iPSCs. Here we show that a combination of zinc finger nucleases (ZFNs) and piggyBac technology in human iPSCs can achieve biallelic correction of a point mutation (Glu342Lys) in the α(1)-antitrypsin (A1AT, also known as SERPINA1) gene that is responsible for α(1)-antitrypsin deficiency. Genetic correction of human iPSCs restored the structure and function of A1AT in subsequently derived liver cells in vitro and in vivo. This approach is significantly more efficient than any other gene-targeting technology that is currently available and crucially prevents contamination of the host genome with residual non-human sequences. Our results provide the first proof of principle, to our knowledge, for the potential of combining human iPSCs with genetic correction to generate clinically relevant cells for autologous cell-based therapies.     

Inhibition sélective de quelques enzymes respiratoires chezEscherichia coli

Summary It is possible to dissociate the activity of different enzymatic systems of the bacteriumEscherichia coli by the action of high hydrostatic pressures. This activity is studied in the microbes after having been subjected to high hydrostatic pressure. Each enzymatic system is inhibited at different pressures. The enzymes of the bacteria submitted to pressures lower than the inhibiting limit show a residual enzymatic activity; sometimes this activity appears after a phase of lag.     

Variations des concentrations de cAMP intracellulaire de différents types de cellules après infection par le virus Sendaï ou le virus de la stomatite vésiculeuse

Summary In KB cells, MRC5 and adult skin fibroblasts infected by low doses of Sendaï virus, intracellular cyclic AMP levels rose and fell in the first hours following infection, then remained lower than basal level during at least 2 days in KB cells and adult skin fibroblasts. When compared to other viruses or cAMP inducers previously described, this effect appeared specific of Sendaï virus. Mechanisms and roles of cAMP variations are discussed. VSV-infected KB cells showed slightly decreased cAMP levels during the first hours following infection.

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Ce travail a bénéficié de l'aide du CNRS (RCP No. 08533) et d'une aide aux équipes de Jeunes Chercheurs de la Mission à la Recherche du Ministère des Universités (bénéficiaire J. Farjanel).

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Les auteurs remercient Madame le Professeur O. Gaudin pour ses conseils et ses encouragements.