Biomedical subjects

R Donato

Publications and source records attributed to R Donato.

At least 37 records · Page 2Linked to original sources

PubMed1999

Role of the C-terminal extension in the interaction of S100A1 with GFAP, tubulin, the S100A1- and S100B-inhibitory peptide, TRTK-12, and a peptide derived from p53, and the S100A1 inhibitory effect on GFAP polymerization.

Whereas native and recombinant S100A1 inhibited GFAP assembly, a truncated S100A1 lacking the last six C-terminal residues (Phe88-Ser93) (S100A1Delta88-93) proved unable to do so. The inhibitory effects of native and recombinant S100A1 on GFAP assembly were blocked by both TRTK-12, a synthetic peptide derived from the alpha-subunit of the actin capping protein, CapZ, and a synthetic peptide derived from the tumor-suppressor protein, p53, in a dose-dependent manner. By fluorescent spectroscopy, TRTK-12 and the p53 peptide, like GFAP and tubulin, caused a dose- and Ca2+-dependent blue-shift of the fluorescence maximum of acrylodan-S100A1. In contrast, GFAP, tubulin, TRTK-12, or the p53 peptide caused no significant changes in the fluorescence spectrum of acrylodan-S100A1Delta88-93. By chemical crosslinking, both TRTK-12 and the p53 peptide strongly reduced or blocked the formation of GFAP-S100A1 or tubulin-S100A1 complexes, respectively, and S100A1Delta88-93 was unable to complex with tubulin, whereas a remarkably reduced complexation of GFAP with the truncated protein was observed. All the above observations show that the C-terminal extension of S100A1 is an essential part of the S100A1 site implicated in the recognition of GFAP, tubulin, p53, and the alpha-subunit of CapZ.

Animals↗

PubMed1999

S100 calcium binding protein affects neuronal electrical discharge activity by modulation of potassium currents.

S100 calcium binding protein has been associated with a variety of intra- and extracellular calcium-mediated functions, including learning and memory. We have previously localized S100-immunoreactive neurons correlated with spontaneous discharge activity in the central nervous system of the mollusc, Helix pomatia. In this study, we further investigated the effects of S100 (S100B and S100A1) on electrical discharge activity and membrane currents of Helix neurons, using current- and voltage-clamp techniques. Extracellular application of disulphide-linked S100B (S100B-s-s) in pico- to nanogram/ml concentrations was found to hyperpolarize the membrane resting potential, to inhibit spontaneous discharge activity of action potentials, to alter the stimulus response behaviour from tonic to phasic, to decrease the duration and increase the afterhyperpolarization of action potentials, and to reduce the cell input resistance. Measurement of membrane currents revealed that the total outward current was increased by S100B-s-s. Separation of outward currents showed that three types of potassium currents were altered: (i) an inward rectifying current, (ii) a calcium-activated potassium outward current, both increased by S100B-s-s, and (iii) a delayed, voltage-dependent potassium outward current which was decreased by the protein. The transient potassium outward and the calcium inward currents were not affected by S100B-s-s. Immunocytochemistry showed intracellular labelling of the cytoplasm after extracellular application of the protein, indicating internalization and suggesting an internal site of action. Injection of S100A1 mimicked the effects of S100B-s-s on discharge activity and action potentials. We conclude from our experiments that S100 calcium binding protein, by modulation of potassium currents, may play a role as a neuromodulator in nervous functions.

Action Potentials↗

PubMed1999

S100B and S100A1 proteins in bovine retina:their calcium-dependent stimulation of a membrane-bound guanylate cyclase activity as investigated by ultracytochemistry.

The Ca2(+)-binding proteins of the EF-hand type, S100B and S100A1, were detected in the outer segment of bovine retina photoreceptors where they are localized to disc membranes, as investigated by immunofluorescence and immunogold cytochemistry. S100B and S100A1 stimulate a membrane-bound guanylate cyclase activity associated with photoreceptor disc membranes in dark-adapted retina in a Ca2(+)-dependent manner, although with different Ca2+ requirements, as investigated by an ultracytochemical approach. Other retinal cell types express S100B and S100A1 as well. S100B is detected in the outer limiting membrane, fine cell processes in the outer nuclear layer and the outer plexiform layer, cell bodies in the inner nuclear layer and the ganglion cell layer, and the inner limiting membrane, whereas S100A1 has a more discrete distribution. S100B and S100A1 also stimulate a membrane-bound guanylate cyclase activity in photoreceptor cell bodies and Muller cells, but their effect appears independent of the light- or dark-adapted state of the retina and is observed at relatively high Ca2+ concentrations. These data represent the ultrastructural counterpart of recent biochemical observations implicating S100B and, possibly, S100A1 in the Ca2(+)-dependent stimulation of a photoreceptor membrane-bound guanylate cyclase activity [T. Duda, R. M. Goraczniak and R. K. Sharma (1996) Molecular characterization of S100A1-S1000B protein in retina and its activation mechanism of bovine photoreceptor guanylate cyclast. Biochemistry 35, 6263-6266; A. Margulis, N. Pozdnyakov and A. Sitaramayya (1996) Activation of bovine photoreceptor guanylate cyclast by S100 proteins. Biochem. Biophys. Res. Commun. 218, 243-247]. Our data suggest that at least S100B may take part in the regulation of a membrane-bound guanylate cyclase-based signalling pathway in both photoreceptors and Muller cells.

Animals↗

PubMed1999

Regulation of the p75 neurotrophin receptor in a rat myogenic cell line (L6).

Neurotrophins are expressed in muscle cells both during development and postnatally. Furthermore, during development muscle cells express high levels of the common p75 neurotrophin receptor, which binds all neurotrophins. Only fragmentary and controversial data are available regarding the responsiveness of muscle cells to neurotrophins and the importance of low-affinity p75 receptor in muscle development. The present study investigates in vitro the immunocytochemical expression of p75 in a rat myogenic cell line (L6) at various time points and in response to different coating substrates as a first step in elucidating the regulation of p75 in muscle. We found that in L6 myoblasts, p75 is expressed only at very early stages of maturation and its levels of expression are regulated by the nature of the coating substrates. p75 expression decreases in cells growing on substrates more suitable for myoblast fusion into myotubes. Time course analysis indicates a reverse correlation between myoblast fusion into myotubes and the levels of p75 expression. Myotubes were always p75 negative. Substrates not suitable for the fusion process induced a prolonged presence of p75 in myoblasts with an increase of their apoptosis. We conclude that expression of p75, at least in this in vitro condition, is regulated by the stages of myoblast differentiation and the nature of the coating substrates. According to the observed time- and substrate-related evidences, future studies should investigate in vivo both the regulation of p75 in the myoblast fusion and the effects and the importance of neurotrophins binding during myoblast differentiation.

Animals↗

PubMed1999

Replicating myoblasts and fused myotubes express the calcium-regulated proteins S100A1 and S100B.

We investigated the expression and the subcellular localization of S100A1 and S100B, two Ca(2+)-binding proteins of the EF-hand type, in replicating myoblasts and fused myotubes. Northern blot and reverse transcriptase-polymerase chain reaction analyses revealed the presence of S100A1 mRNA and S100B mRNA respectively, in myoblasts. Immunofluorescence and immunogold electron microscopy were used to localize individual proteins in myoblasts and myotubes. In the present report we document that: (1) in replicating myoblasts S100B is localized to intracellular membranes, including Golgi membranes, vimentin intermediate filaments (IFs) and microtubule (MT) structures; (2) in the same cells S100A1 is found associated with intracellular membranes; (3) following treatment of replicating myoblasts with colchicine, a fraction of S100B remains colocalized with bundled and collapsed vimentin IFs, whereas another fraction follows the destiny of endoplasmic membranes; (4) under the same conditions S100A1, like a fraction of S100B, follows the collapse of the endoplasmic reticulum around the nucleus; and (5) in fused myotubes S100A1 is found diffusely in the cytoplasm, whereas S100B is mostly found associated with vimentin IFs. These data suggest that in the skeletal myogenic cell line used in the present study S100A1 and S100B might share binding sites on or close to intracellular membranes, but display a significant degree of target specificity with respect of IFs and MTs. The results of these analyses suggest that expression of S100B in skeletal muscle cells may be developmentally regulated and lend support to the possibility that S100B might regulate the MT and IF dynamics.

Amino Acid Sequence↗

PubMed1999

The calcium-modulated proteins, S100A1 and S100B, as potential regulators of the dynamics of type III intermediate filaments.

The Ca2+-modulated, dimeric proteins of the EF-hand (helix-loop-helix) type, S100A1 and S100B, that have been shown to inhibit microtubule (MT) protein assembly and to promote MT disassembly, interact with the type III intermediate filament (IF) subunits, desmin and glial fibrillary acidic protein (GFAP), with a stoichiometry of 2 mol of IF subunit/mol of S100A1 or S100B dimer and an affinity of 0.5-1.0 microM in the presence of a few micromolar concentrations of Ca2+. Binding of S100A1 and S100B results in inhibition of desmin and GFAP assemblies into IFs and stimulation of the disassembly of preformed desmin and GFAP IFs. S100A1 and S100B interact with a stretch of residues in the N-terminal (head) domain of desmin and GFAP, thereby blocking the head-to-tail process of IF elongation. The C-terminal extension of S100A1 (and, likely, S100B) represents a critical part of the site that recognizes desmin and GFAP. S100B is localized to IFs within cells, suggesting that it might have a role in remodeling IFs upon elevation of cytosolic Ca2+ concentration by avoiding excess IF assembly and/or promoting IF disassembly in vivo. S100A1, that is not localized to IFs, might also play a role in the regulation of IF dynamics by binding to and sequestering unassembled IF subunits. Together, these observations suggest that S100A1 and S100B may be regarded as Ca2+-dependent regulators of the state of assembly of two important elements of the cytoskeleton, IFs and MTs, and, potentially, of MT- and IF-based activities.

Calcium↗

PubMed1998

Association of S100B with intermediate filaments and microtubules in glial cells.

Previous in vitro studies have shown that the Ca2+-regulated S100B protein modulates the assembly-disassembly of microtubules (MTs) and type III intermediate filaments (IFs). In the present report, by double immunofluorescence cytochemistry S 100B was localized to both GFAP/vimentin IFs and MTs as well as to centrosomes in U251 glial cells. In cells treated with the MT-depolymerizing agent, colchicine, S100B remained associated with the rearranged GFAP IFs throughout the cell and, at the cell periphery, vimentin IFs. In cells treated with the MT stabilizing agent, taxol, S100B followed partly the rearrangement of MTs and partly the rearrangement of IFs. Under the latter condition, bundles of MTs with their associated S100B appeared surrounded and/or flanked by rearranged IFs with their associated S100B. Colocalization of S100B with closely arranged IFs and MTs was best evident in cells manipulated with taxol and in triton-cytoskeletons. In these cases, MTs and their associated S100B appeared surrounded and/or flanked by and/or intermingled with IFs and their associated S100B. Also, a preferential association of S100B with GFAP vs. vimentin IFs could be observed near the nucleus where colocalization of S100B with MTs was also maximal. Condensation of IFs and alteration of the MT network caused by treatment of cells with the phosphatase inhibitor, okadaic acid, resulted in a concomitant condensation/alteration of the S100B immunoreactivity. The present results lend support to the possibility that S100B may be an important factor implicated in the regulation of the dynamics of MTs and IFs.

Calcium-Binding Proteins↗

PubMed1998

Annexin VI binds S100A1 and S100B and blocks the ability of S100A1 and S100B to inhibit desmin and GFAP assemblies into intermediate filaments.

Annexin VI, a member of a family of Ca(2+)-dependent phospholipid- and membrane-binding proteins, interacts with the Ca(2+)-regulated EF-hand proteins, S100A1 and S100B, and blocks the ability of these two proteins to inhibit the assembly of desmin and glial fibrillary acidic protein (GFAP) into intermediate filaments in a Ca(2+)- and dose-dependent manner. S100A1 and S100B each possess one annexin VI binding site, characterized by an affinity for annexin VI in the submicromolar range. Binding of annexin VI to either S100 protein occurs at a site that appears to differ in some parts from that recognizing desmin and GFAP. As S100A1 and S100B exist in solution as homodimers in which the two monomers are related by a 2-fold symmetry axis, each of the above S100 homodimers likely crosslinks two annexin VI molecules, a situation that appears typical of all the annexin-S100 protein complexes described thus far. However, whereas in the cases of other annexin-S100 complexes the C-terminal extension of the S100 molecule appears indispensable for annexin binding, the annexin VI binding site cannot be restricted to the S100A1 and S100B C-terminal extension. We speculate that the annexin VI site on S100A1/B may only partially overlap to the desmin/GFAP site. In contrast, no effects of annexin V on the ability of S100A1 or S100B to affect the desmin and GFAP assemblies could be documented, although binding of annexin V to S100A1 and S100B could be detected at relatively high Ca2+ concentrations. The present data suggest that annexin VI might regulate S100A1 and S100B activities and vice versa.

Animals↗

PubMed1997

Annexin-immunoreactive proteins in the nervous system and eye of the gastropods, Aplysia and Helix.

We studied the distribution of annexin I- and annexin V-like proteins in the eye and central nervous system of the snails, Aplysia californica and Helix pomatia by immunocytochemistry. Annexin I-immunoreactive material in Aplysia californica was localized in sensory and corneal cells of the eye and in distinct neurons of the cerebral, buccal, and abdominal ganglia, where it was exclusively located in bag cells. Annexin V-immunoreactive neurons were restricted to the pleural ganglia of Aplysia californica. In Helix pomatia annexin I-immunoreactive neurons were present in the cerebral, buccal, visceral, and left and right parietal ganglia, whereas annexin V-immunoreactivity neurons were present in left and right pleural, left and right parietal, visceral, and buccal ganglia. Annexin VI-immunoreactivity was absent in both gastropods studied. Our study shows a cell specific localization of annexin-like proteins in the central nervous system and eye of molluscs. The cell types containing the immunoreactive proteins suggests that the annexin-like proteins may be involved in intracellular signaling mechanisms, which ultimately may modulate egg-laying and circadian rhythmicity.

Animals↗

PubMed1997

Leukemia inhibitory factor enhances the regeneration of transected rat sciatic nerve and the function of reinnervated muscle.

The cytokine leukemia inhibitory factor (LIF) favors the survival and growth of axons in vitro and in vivo. Fibronectin has been shown to enhance nerve regeneration when added in combination with various growth factors including LIF. The goal of this study was to evaluate the effect of LIF plus fibronectin on the regeneration of transected nerve and functional recovery of reinnervated skeletal muscle, in one experimental model of peripheral nerve repair, at two recovery times. The rat sciatic nerve was cut at mid-thigh level and a silicone cuff containing either saline (control), LIF, or LIF plus fibronectin (L+F) was used to bridge the proximal and distal nerve stumps leaving a 1 cm gap between them. Rats were then explored at 6 or 12 weeks following the initial surgery. Regenerating nerves were assessed by measuring the diameter of myelinated axons, conduction velocity, and number of myelinated fibers. Muscle reinnervation was assessed by measuring muscle mass, force of contraction, and histologically for changes in muscle fiber type (type I and type II). In this report we demonstrate that at 6 weeks there were significant increases in 1) nerve conduction velocity, 2) myelinated axon diameter, and 3) number of myelinated axons over that of control (saline-treated) animals. Both LIF groups demonstrated a shift in type II muscle fiber area compared to saline-treated controls, with the L+F group having a significant increase in muscle mass. At 12 weeks there was an improved recovery over and above that demonstrated at 6 weeks. Muscle mass was 65% and 42% greater than control for LIF and L+F, respectively. Force of contraction, conduction velocity, myelinated fiber number, and diameter were also significantly greater for both LIF- and L+F-treated rats than saline-treated rats. These results demonstrate that LIF significantly improves the regeneration of damaged peripheral nerves and the preservation of muscle viability, resulting in greatly enhanced recovery of skeletal muscle function.

Analysis of Variance↗

PubMed1997

Pyrazolo[1,5-a]pyrido[3,4-e]pyrimidin-6-ones. II. Synthesis and in vitro antimicrobial evaluation.

A series of Pyrazolo[1,5-a]pyrido[3,4-e]pyrimidin-6-ones (4a-p) was prepared by a simple synthetic procedure based on the reaction of hydroxylamine or methoxyamine with 2,3-substituted ethyl 7-dimethylaminovinyl pyrazolo[1,5-a]pyrimidin-6-carboxylates (3a-p). The antimicrobial activity of the obtained compounds was evaluated on a series of standard strains of Gram positive, Gram negative bacteria and fungi. None of the tested compounds showed significant activity.

Anti-Bacterial Agents↗

PubMed1996

S-100 (alpha and beta) binding peptide (TRTK-12) blocks S-100/GFAP interaction: identification of a putative S-100 target epitope within the head domain of GFAP.

Alignment of previously characterized S-100 (alpha and beta)-binding peptides (J. Biol. Chem. 270, 14651-14658) has enabled the identification of a putative S-100 target epitope within the head domain of glial fibrillary acidic protein (GFAP). The capacity of a known peptide inhibitor of S-100 protein (TRTK-12), homologous to this region, to perturb the interaction of S-100 (alpha and beta) and GFAP (J. Biol. Chem 268, 12669-12674) was investigated. Fluorescence spectrophotometry and chemical cross-linking analyses determined TRTK-12 to disrupt S-100:GFAP interaction in a dose- and Ca(2+_dependent manner. TRTK-12 also inhibited S-100's ability to block GFAP assembly and to mediate disassembly of preformed glial filaments. Each of these events was strictly dependent upon the presence of calcium and inhibitory peptide, maximal inhibition occurring at a concentration of TRTK-12 equivalent to the molar amount of S-100 monomer present. Together with our recent report demonstrating TRTK-12 also blocks the interaction of S-100 protein with the actin capping protein, CapZ, these results suggest TRTK-12 functions as a pleiotropic inhibitor of S-100 function. Availability of a functional inhibitor of S-100 will assist the further characterization of S-100 protein function in vitro and in vivo. Moreover, this report provides additional evidence supportive of a role for S-100 as a multi-faceted regulator of cytoskeletal integrity.

2-Naphthylamine↗

PubMed1996

Characterization of type III intermediate filament regulatory protein target epitopes: S-100 (beta and/or alpha) binds the N-terminal head domain; annexin II2-p11(2) binds the rod domain.

We have investigated the interaction of S-100 proteins (beta and/or alpha) and annexin II2-p11(2) with glial fibrillary acidic protein (GFAP) and desmin to have further information on the mechanisms whereby S-100 proteins and annexin II2-p11(2) affect assembly/disassembly of GFAP and desmin intermediate filaments (IFs). Analyses were conducted on either native IF subunits, GFAP or desmin rod domain, or headless GFAP or desmin. Our data indicate that: (i) S-100 proteins bind to GFAP and desmin N-terminal head domain; (ii) annexin II2-p11(2) binds to GFAP rod domain; (iii) annexin II2-p11(2) does not interact with desmin nor affects desmin assembly. The present data suggest that the ability of S-100 proteins to inhibit GFAP and desmin assemblies and to promote the disassembly of preformed GFAP and desmin IFs depends on occupation of a site on the N-terminal head domain of these IF subunit. It is known that the N-terminal head domain is critical for the progression from the stage of GFAP and desmin dimers/tetramers to that of large oligomers. On the other hand, the ability of annexin II2-p11(2) to stimulate GFAP assembly under conditions where this latter is normally hampered (e.g., at alkaline pH values) might depend on annexin II2-p11(2)-induced changes in the structure of GFAP rod domain, possibly as a consequence of charge modifications. By contrast, the inability of annexin II2-p11(2) to bind to desmin would depend on desmin resistance to charge modifications.

Animals↗

PubMed1996

Time-resolved fluorescence of S-100a protein: effect of Ca2+, Mg2+ and unilamellar vesicles of egg phosphatidylcholine.

Phase-modulation fluorescence lifetime measurements were used to study the single Trp residue of the Ca(2+)-binding protein S-100a both in the absence and in the presence of Ca2+ and/or Mg2+. Trp fluorescence decay for the protein was satisfactorily described by Lorentzian lifetime distributions centered around two components (approximately 4 ns and 0.5 ns). Lifetime values were unchanged by 2 mM Ca2+, but the fractional intensity associated with longer lifetime increased up to 75%. In the presence of Mg2+, the Ca2+ induced increase of the fractional intensity associated with longer lifetime was only 57%. For the protein in buffer, about the 85% of the recovered anisotropy was associated to a rotational correlation time of 6.7 ns. After the addition of Ca2+, this value was increased to 16.08 ns. In the presence of Mg2+, Ca+2 increased the rotational correlation time to 33.75 ns. Similar studies were performed with S-100a interacting with egg phosphatidylcholine vesicles (SUV). Our data suggest that the conformation of the protein may be influenced by structural features of the lipidic membrane. Moreover, data obtained in the presence of Mg2+ indicate some interaction between lipids and S-100, likely mediated by this ion.

Animals↗

PubMed1996

Effects of calcium-binding proteins (S-100a(o), S-100a, S-100b) on desmin assembly in vitro.

S-100a(o), the alpha alpha isoform of a subfamily of Ca(2+)-binding proteins of the EF-hand type expressed in cardiac and skeletal muscle cells, is reported to inhibit the assembly of the intermediate filament subunit desmin and to stimulate the disassembly of desmin intermediate filaments in the presence of micromolar levels of free Ca(2+). These effects are dose-dependent with respect to the S-100a(o) concentration and maximal at a desmin/S-100a(o) (dimer) molar ratio of approximately 2. Other members of the S-100 subfamily [S-100a (alpha beta) and S-100b (beta beta) and the unfractionated mixture of S-100a plus S-100b produce qualitatively similar effects on desmin assembly, with a potency that depends on the fraction of S-100alpha subunit (the most potent) or S-100beta subunit (the least potent) present in the S-100 isoforms tested. A binding stoichiometry of 2 mol of desmin/mol of S-100a(o) (dimer) and an affinity in the submicromolar range are calculated. The S-100beta subunit also interacts with desmin, but with a lower affinity compared with S-100alpha. By contrast, the S-100-like proteins calcyclin and p11 neither interact with desmin nor affect desmin assembly. The present data suggest that S-100a(o) might play a role in the regulation of the state of assembly of desmin intermediate filaments.

Animals↗

PubMed1996

Pyrazolo[1,5-a]pyrido[3,4-e]pyrimidin-6-ones. I. Synthesis and in vitro antimicrobial evaluation.

A series of pyrazolo[1,5-a]pyrido[3,4-e]pyrimidin-6-ones (9-36) was prepared by a simple synthetic procedure based on the reaction of ethyl 7-dimethylaminovinyl pyrazolo[1,5-a]pyrimidin-6-carboxylates (1-8) and reagents (a-n) containing a primary amino group. When this set of compounds was subjected to "in vitro" antimicrobial screening, it showed no significant activity.

Anti-Bacterial Agents↗

PubMed1995

Detection of membrane-bound guanylate cyclase activity in rat C6 glioma cells at different growth states following activation by natriuretic peptides.

We studied the activity and the ultracytochemical localization of membrane-bound guanylate cyclase (GC) after stimulation with rat atrial natriuretic peptide (rANP), porcine brain natriuretic peptide (pBNP), rat brain natriuretic peptide (rBNP), or porcine C-type natriuretic peptide (CNP) in rat C6 glioma cells during proliferation or following exposure of confluent cells to dibutyryl cyclic AMP (db-cAMP) or retinoic acid (RA). Under our experimental conditions all peptides were activators of GC as demonstrated by the accumulation of cGMP within cells. During proliferation of C6 cells, the amounts of cGMP remained approximately constant. However, at subconfluency, confluency and postconfluency, the GC reaction product was located at different sites in C6 cells. At subconfluency, GC reaction product was on membranes of protoplasmic extensions, at postconfluency, GC reaction product was in association with membranes of cell bodies, and at confluency, both localizations of GC reaction product were detected. Incubation of confluent cells in culture medium containing db-cAMP or RA induced the appearance of long and slender protoplasmic extensions. Under these conditions, the GC reaction product was localized exclusively to these processes. These data suggest that GC is differentially located depending on the state of growth of glial cells, and that in differentiating glial cells GC is preferentially located in cell processes.

Animals↗

PubMed1995

Annexin II2-p11(2) (calpactin I) stimulates the assembly of GFAP in a calcium- and pH-dependent manner.

Annexin II2-p11(2) (calpactin I) was tested as a potential regulator of GFAP assembly into glial filaments (GF), following the observation that it interacts with GFAP and cosediments with GF in a sedimentation assay. Under conditions where GFAP assembly is reduced, e.g., at pH values > 6.8, annexin II2-p11(2) stimulates GF formation in a Ca(2+)- and dose-dependent manner. Concomitantly, an ever larger fraction of annexin II2-p11(2) can be recovered in GF pellets as the pH is raised from 6.8 to 7.35. Monomeric annexin II also stimulates GFAP assembly, although with a smaller efficacy as compared to annexin II2-p11(2), but does not cosediment with GF to a large extent, whereas p11 neither cosediments with GF nor affects GFAP assembly. On the other hand, the in vitro reconstituted annexin II2-p11(2) heterotetramer mimics native annexin II2-p11(2), and perturbation of the integrity of annexin II2-p11(2) by a mild treatment with alpha-chymotrypsin results in the nearly complete abolition of the stimulatory effect of annexin II2-p11(2) on GFAP assembly. These data suggest that annexin II2-p11(2) might be involved in the regulation of the state of assembly of GF, possibly in concert with other proteins.

Animals↗