Aß42-oligomer Interacting Peptide (AIP) neutralizes toxic amyloid-ß42 species and protects synaptic structure and function.

The amyloid-ß42 (Aß42) peptide is believed to be the main culprit in the pathogenesis of Alzheimer disease (AD), impairing synaptic function and initiating neuronal degeneration. Soluble Aß42 oligomers are highly toxic and contribute to progressive neuronal dysfunction, loss of synaptic spine density, and affect long-term potentiation (LTP). We have characterized a short, L-amino acid Aß-oligomer Interacting Peptide (AIP) that targets a relatively well-defined population of low-n Aß42 oligomers, rather than simply inhibiting the aggregation of Aß monomers into oligomers. Our data show that AIP diminishes the loss of Aß42-induced synaptic spine density and rescues LTP in organotypic hippocampal slice cultures. Notably, the AIP enantiomer (comprised of D-amino acids) attenuated the rough-eye phenotype in a transgenic Aß42 fly model and significantly improved the function of photoreceptors of these flies in electroretinography tests. Overall, our results indicate that specifically "trapping" low-n oligomers provides a novel strategy for toxic Aß42-oligomer recognition and removal.

Potassium-chloride cotransporter 3 interacts with Vav2 to synchronize the cell volume decrease response with cell protrusion dynamics.

Loss-of-function of the potassium-chloride cotransporter 3 (KCC3) causes hereditary motor and sensory neuropathy with agenesis of the corpus callosum (HMSN/ACC), a severe neurodegenerative disease associated with defective midline crossing of commissural axons in the brain. Conversely, KCC3 over-expression in breast, ovarian and cervical cancer is associated with enhanced tumor cell malignancy and invasiveness. We identified a highly conserved proline-rich sequence within the C-terminus of the cotransporter which when mutated leads to loss of the KCC3-dependent regulatory volume decrease (RVD) response in Xenopus Laevis oocytes. Using SH3 domain arrays, we found that this poly-proline motif is a binding site for SH3-domain containing proteins in vitro. This approach identified the guanine nucleotide exchange factor (GEF) Vav2 as a candidate partner for KCC3. KCC3/Vav2 physical interaction was confirmed using GST-pull down assays and immuno-based experiments. In cultured cervical cancer cells, KCC3 co-localized with the active form of Vav2 in swelling-induced actin-rich protruding sites and within lamellipodia of spreading and migrating cells. These data provide evidence of a molecular and functional link between the potassium-chloride co-transporters and the Rho GTPase-dependent actin remodeling machinery in RVD, cell spreading and cell protrusion dynamics, thus providing new insights into KCC3's involvement in cancer cell malignancy and in corpus callosum agenesis in HMSN/ACC.

Intracellular hypertonicity is responsible for water flux associated with Na+/glucose cotransport.

Detection of a significant transmembrane water flux immediately after cotransporter stimulation is the experimental basis for the controversial hypothesis of secondary active water transport involving a proposed stoichiometry for the human Na(+)/glucose cotransporter (SGLT1) of two Na(+), one glucose, and 264 water molecules. Volumetric measurements of Xenopus laevis oocytes coexpressing human SGLT1 and aquaporin can be used to detect osmotic gradients with high sensitivity. Adding 2 mM of the substrate alpha-methyl-glucose (alphaMG) created mild extracellular hypertonicity and generated a large cotransport current with minimal cell volume changes. After 20, 40, and 60 s of cotransport, the return to sugar-free, isotonic conditions was accompanied by measurable cell swelling averaging 0.051, 0.061, and 0.077 nl/s, respectively. These water fluxes are consistent with internal hypertonicities of 1.5, 1.7, and 2.2 mOsm for these cotransport periods. In the absence of aquaporin, the measured hypertonicites were 4.6, 5.0, and 5.3 mOsm for the same cotransport periods Cotransport-dependent water fluxes, previously assumed to be water cotransport, could be largely explained by hypertonicities of such amplitudes. Using intracellular Na(+) injection and Na(+)-selective electrode, the intracellular diffusion coefficient for Na(+) was estimated at 0.29 +/- 0.03 x 10(-5) cm(2) s(-1). Using the effect of intracellular alphaMG injection on the SGLT1-mediated outward current, the intracellular diffusion coefficient of alphaMG was estimated at 0.15 +/- 0.01 x 10(-5) cm(2) s(-1). Although these intracellular diffusion coefficients are much lower than in free aqueous solution, a diffusion model for a single solute in an oocyte would require a diffusion coefficient three times lower than estimated to explain the local osmolyte accumulation that was experimentally detected. This suggests that either the diffusion coefficients were overestimated, possibly due to the presence of convection, or the diffusion in cytosol of an oocyte is more complex than depicted by a simple model.

The human tumour suppressor gene SLC5A8 expresses a Na+-monocarboxylate cotransporter.

The orphan cotransport protein expressed by the SLC5A8 gene has been shown to play a role in controlling the growth of colon cancers, and the silencing of this gene is a common and early event in human colon neoplasia. We expressed this protein in Xenopus laevis oocytes and have found that it transports small monocarboxylic acids. The electrogenic activity of the cotransporter, which we have named SMCT (sodium monocarboxylate transporter), was dependent on external Na(+) and was compatible with a 3 : 1 stoichiometry between Na(+) and monocarboxylates. A portion of the SMCT-mediated current was also Cl(-) dependent, but Cl(-) was not cotransported. SMCT transports a variety of monocarboxylates (similar to unrelated monocarboxylate transport proteins) and most transported monocarboxylates demonstrated K(m) values near 100 microm, apart from acetate and d-lactate, for which the protein showed less affinity. SMCT was strongly inhibited by 1 mm probenecid or ibuprofen. In the absence of external substrate, a Na(+)-independent leak current was also observed to pass through SMCT. SMCT activity was strongly inhibited after prolonged exposure to high external concentrations of monocarboxylates. The transport of monocarboxylates in anionic form was confirmed by the observation of a concomitant alkalinization of the cytosol. SMCT, being expressed in colon and kidney, represents a novel means by which Na(+), short-chain fatty acids and other monocarboxylates are transported in these tissues. The significance of a Na(+)-monocarboxylate transporter to colon cancer presumably stems from the transport of butyrate, which is well known for having anti-proliferative and apoptosis-inducing activity in colon epithelial cells.