Claude Gicquaud

Stabilization of actin by phalloidin: A differential scanning calorimetric study

We have used differential scanning calorimetry to study the effects of phalloidin on F and G actin stability. For F actin, saturating concentrations of phalloidin induced an important shift on the transition temperature, Tm, from 69.5 degrees C to 83.5 degrees C. However, the calorimetric enthalpy remained unchanged. Using lower phalloidin concentrations, monomers linked to phalloidin, as well as neighboring unlinked monomers, were both stabilized. Contrary to previous reports, phalloidin was also shown to affect G actin, shifting its Tm from 59.5 degrees C to 75 degrees C. Two mechanisms are proposed to explain this finding: first, it could indicate a real interaction of phalloidin with G actin, and second, heating of the specimen during the scan could have induced polymerization of some G actin to the F form. The resulting F polymer would then interact with phalloidin, thus shifting the equilibrium between G and F actin towards the polymeric form.

An ultrastructural analysis of early fertilization events in the giant scallop, Placopecten magellanicus(Mollusca, Pelecypoda)

Summary We examined the ultrastuctural events occurring during early fertilization in the giant scallop, Placopecten magellanicus. The unfertilized oocytes had not completed their meiotic maturation and were blocked at the metaphase I stage. At fertilization, there was a local loss of microvilli, and a fertilization cone of 6 μm in length by 4 μm in diameter was produced. As the spermatozoon penetrated the oocyte, its head rotated by 90 to 180°. The condensed electron-opaque state of the sperm chromatin transformed into a dispersed form of male pronucleus; this event was coupled with the beginning of the formation of the pronuclear envelope and the development of the sperm aster. The male pronucleus was completely formed 60 min postfertilization, before completion of meiosis, and spots of electron-dense chromatin still remained. During sperm nuclear reorganization, meiotic maturation was resumed. At anaphase I, 30 min after fertilization, the dyad chromosomes moved into a cytoplasmic protrusion that becam...

Research on the mechanism of interaction between actin and membrane lipids

Using an in vitro system involving pure actin and liposomes, we have established that actin may interact with membrane lipids without any intermediate proteins, and that the mechanism of interaction depends upon the concentration of divalent cation. In the absence of divalent cation, actin increases membrane permeability. Low concentrations (1 mM) of divalent cation potentialize this interaction. In the presence of high divalent cation concentration, actin deposits on the surface of liposomes in a crystalline organization and reduces the membrane microviscosity as shown by the polarization of fluorescence of the DPH probe. We propose that actin interacts with lipids by hydrophobic association which is facilitated by initial electrostatic binding.

Surface film pressure of actin: interactions with lipids in mixed monolayers

The interactions of actin with neutral lipid films made from DLPC, and with positively charged films built from DLPC and stearylamine (SA), have been characterized by the monolayer technique. Injection of actin underneath an expanded lipid film produces an increase in the surface pressure that is consistent with a penetration of the lipid molecules by actin. This adsorption of actin to the lipid is more pronounced either with positively charged films or with Mg(2+) present in the sub-phase, suggesting that the mechanism involves an electrostatic attraction. During compression, the actin molecules are squeezed out into the sub-phase, carrying along some lipid molecules; this suggests a strong affinity of the lipids for actin. An analysis of the dilational modulus shows that when actin is found as monomers at the interface, the mixed actin-lipid film undergoes three phase changes upon compression. On the other hand, when actin is polymerized at the interface, the actin and the lipid form a rigid film for which the compressibility is mostly dominated by actin.