The CaMKII/MLC1 Axis Confers Ca-Dependence to Volume-Regulated Anion Channels (VRAC) in Astrocytes.

Astrocytes, the main glial cells of the central nervous system, play a key role in brain volume control due to their intimate contacts with cerebral blood vessels and the expression of a distinctive equipment of proteins involved in solute/water transport. Among these is MLC1, a protein highly expressed in perivascular astrocytes and whose mutations cause megalencephalic leukoencephalopathy with subcortical cysts (MLC), an incurable leukodystrophy characterized by macrocephaly, chronic brain edema, cysts, myelin vacuolation, and astrocyte swelling. Although, in astrocytes, MLC1 mutations are known to affect the swelling-activated chloride currents (ICl,) mediated by the volume-regulated anion channel (VRAC), and the regulatory volume decrease, MLC1's proper function is still unknown.

Megalencephalic Leukoencephalopathy with Subcortical Cysts Disease-Linked MLC1 Protein Favors Gap-Junction Intercellular Communication by Regulating Connexin 43 Trafficking in Astrocytes.

Astrocytes, the most numerous cells of the central nervous system, exert critical functions for brain homeostasis. To this purpose, astrocytes generate a highly interconnected intercellular network allowing rapid exchange of ions and metabolites through gap junctions, adjoined channels composed of hexamers of connexin (Cx) proteins, mainly Cx43. Functional alterations of Cxs and gap junctions have been observed in several neuroinflammatory/neurodegenerative diseases.

Megalencephalic Leukoencephalopathy with Subcortical Cysts Protein-1 (MLC1) Counteracts Astrocyte Activation in Response to Inflammatory Signals.

Megalencephalic leukoencephalopathy with subcortical cysts protein-1 (MLC1) is a membrane protein expressed by perivascular astrocytes. MLC1 mutations cause MLC, an incurable leukodystrophy characterized by macrocephaly, brain edema, cysts, myelin vacuolation, and astrocytosis, leading to cognitive/motor impairment and epilepsy. Although its function is unknown, MLC1 favors regulatory volume decrease after astrocyte osmotic swelling and down-regulates intracellular signaling pathways controlling astrocyte activation and proliferation.

Megalencephalic leukoencephalopathy with subcortical cysts protein-1 regulates epidermal growth factor receptor signaling in astrocytes.

Mutations in the MLC1 gene, which encodes a protein expressed in brain astrocytes, are the leading cause of MLC, a rare leukodystrophy characterized by macrocephaly, brain edema, subcortical cysts, myelin and astrocyte vacuolation. Although recent studies indicate that MLC1 protein is implicated in the regulation of cell volume changes, the exact role of MLC1 in brain physiology and in the pathogenesis of MLC disease remains to be clarified. In preliminary experiments, we observed that MLC1 was poorly expressed in highly proliferating astrocytoma cells when compared with primary astrocytes, and that modulation of MLC1 expression influenced astrocyte growth.

Native metastable prefibrillar oligomers are the most neurotoxic species among amyloid aggregates.

Many proteins belonging to the amyloid family share the tendency to misfold and aggregate following common steps, and display similar neurotoxicity. In the aggregation pathway different kinds of species are formed, including several types of oligomers and eventually mature fibers. It is now suggested that the pathogenic aggregates are not the mature fibrils, but the intermediate, soluble oligomers.

Phosphorylation and nitration of tyrosine residues affect functional properties of Synaptophysin and Dynamin I, two proteins involved in exo-endocytosis of synaptic vesicles.

Phosphorylation and nitration of protein tyrosine residues are thought to play a role in signaling pathways at the nerve terminal and to affect functional properties of proteins involved in the synaptic vesicle (SV) exo-endocytotic cycle. We previously demonstrated that the tyrosine residues in the C-terminal domain of the SV protein Synaptophysin (SYP) are targets of peroxynitrite (PN). Here, we have characterized the association between SYP and c-src tyrosine kinase demonstrating that phosphorylation of Tyr(273) in the C-terminal domain of SYP is crucial in mediating SYP binding to and activation of c-src.

MLC1 trafficking and membrane expression in astrocytes: role of caveolin-1 and phosphorylation.

Megalencephalic leukoencephalopathy with subcortical cysts (MLC) is a rare congenital leukodystrophy caused by mutations in the MLC1 gene that encodes a membrane protein of unknown function. In the brain MLC1 protein is mainly expressed in astrocyte end-feet, localizes in lipid rafts and associates with the dystrophin glycoprotein complex (DGC). Using pull-down and co-fractionation assays in cultured human and rat astrocytes, we show here that MLC1 intracellular domains pull-down the DGC proteins syntrophin, dystrobrevin, Kir4.

Peroxynitrite induces tyrosine residue modifications in synaptophysin C-terminal domain, affecting its interaction with src.

Peroxynitrite is a potent oxidant that contributes to tissue damage in neurodegenerative disorders. We have previously reported that treatment of rat brain synaptosomes with peroxynitrite induced post-translational modifications in pre- and post-synaptic proteins and stimulated soluble N-ethylmaleimide sensitive fusion proteins attachment receptor complex formation and endogenous glutamate release. In this study we show that, following peroxynitrite treatment, the synaptic vesicle protein synaptophysin (SYP) can be both phosphorylated and nitrated in a dose-dependent manner.

Biochemical characterization of MLC1 protein in astrocytes and its association with the dystrophin-glycoprotein complex.

MLC1 gene mutations have been associated with megalencephalic leukoencephalopathy with subcortical cysts (MLC), a rare neurologic disorder in children. The MLC1 gene encodes a membrane protein (MLC1) with unknown function which is mainly expressed in astrocytes. Using a newly developed anti-human MLC1 polyclonal antibody, we have investigated the biochemical properties and localization of MLC1 in cultured astrocytes and brain tissue and searched for evidence of a relationship between MLC1 and proteins of the dystrophin-glycoprotein complex (DGC).

Synaptic remodeling induced by axotomy of superior cervical ganglion neurons: Involvement of metalloproteinase-2.

We previously demonstrated the involvement of the dystrophin-dystroglycan (Dys-DG) complex in the stabilization of intraganglionic synapses in rodent superior cervical ganglion (SCG) by investigating changes in the organization of their post-synaptic apparatus induced either by ganglionic neuron axotomy or by the lack of Dys in genetically dystrophic mdx mice, or by the combination of the two. A role of the matrix metalloproteinases (MMPs) MMP-2 and MMP-9 in the degradation of DG and, hence, in disrupting the connection between the extracellular matrix (ECM) and the cortical cytoskeleton, has recently been proposed. We hypothesized that the degradation by MMPs of ECM proteins and DG in ganglionic neurons may be involved in injury-induced synaptic detachment observed in rodent SCG.

Immunodetection of partially glycosylated isoforms of alpha-dystroglycan by a new monoclonal antibody against its beta-dystroglycan-binding epitope.

The alpha/beta dystroglycan (DG) complex links the extracellular matrix to the actin cytoskeleton. The extensive glycosylation of alpha-DG is believed to be crucial for the interaction with its extracellular matrix-binding partners. We characterized a monoclonal antibody, directed against the beta-DG-binding epitope ( approximately positions 550-565), which recognizes preferentially hypoglycosylated alpha-DG.