Summary Dendritic spines undergo actin-based growth and shrinkage during synaptic plasticity. The actin depolymerizing factor (ADF)/cofilin family of actin-associated proteins plays important roles in spine plasticity. Elevated ADF/cofilin activities often lead to reduced spine size and immature spine morphology, but can enhance synaptic potentiation in some cases. Therefore ADF/cofilin may exert distinct effects on postsynaptic structure and function. Here we report that ADF/cofilin-mediated actin dynamics regulate AMPA receptor (AMPAR) trafficking during synaptic potentiation, which is distinct from actin's structural role in spine morphology. We find that elevated ADF/cofilin activity markedly enhances surface addition of AMPARs after chemically-induced LTP (cLTP), whereas inhibition of ADF/cofilin abolishes AMPAR addition. Our data further show that cLTP elicits a temporal sequence of ADF/cofilin dephosphorylation and phosphorylation that underlies AMPAR trafficking and spine enlargement. These findings suggest a novel role for temporally-regulated ADF/cofilin activities in postsynaptic modifications of receptor number and spine size during synaptic plasticity.

Cell-cell interactions organize lens fiber cells into highly ordered structures to maintain transparency. However, signals regulating such interactions have not been well characterized. We report here that ephrin-A5, a ligand of the Eph receptor tyrosine kinases, plays a key role in lens fiber cell shape and cell-cell interactions. Lens fiber cells in mice lacking ephrin-A5 function appear rounded and irregular in cross-section, in contrast to their normal hexagonal appearance in WT lenses. Cataracts eventually develop in 87% of ephrin-A5 KO mice. We further demonstrate that ephrin-A5 interacts with the EphA2 receptor to regulate the adherens junction complex by enhancing recruitment of ␤-catenin to N-cadherin. These results indicate that the Eph receptors and their ligands are critical regulators of lens development and maintenance.␤-catenin ͉ Eph receptor ͉ N-cadherin C ataract, or the opacification of the lens, is the leading cause of visual impairment and blindness worldwide (1). The molecular events underlying lens development and the processes by which the lens maintains transparency over a lifetime are unclear (2). In addition, the cellular and biochemical mechanisms underlying the pathological changes leading to cataract remain poorly understood.The lens is composed of a single layer of epithelial cells on the anterior surface, which, over a lifetime, divide and differentiate into the underlying lens fiber cells that comprise the bulk of the lens (3, 4). Initially during lens development, primary lens fiber cells differentiate and elongate from the posterior pole. In later embryogenesis and throughout life, secondary lens fiber cells differentiate from lens epithelial cells located at the equator. In cross section, the lens fiber cells resemble flattened hexagons with two broad and four short sides (3). These cells are organized in a highly ordered and closely packed manner, and interact through extensive intercellular adhesion complexes including gap and adherens junctions (5). Fiber cell gap junctions are composed of connexins (Cx) 46 and 50 (6), inactivation of which leads to the degeneration of the inner fiber cells and the development of cataract in mice (7,8). Mutations in human Cx genes have also been associated with cataractogenesis (9, 10). As the lens is completely enclosed by an acellular, avascular capsule, it is believed that these cell-cell junctions are critical for providing nutrient transport, removal of metabolic wastes, and maintenance of homeostasis (11,12). In addition to gap junctions, widespread adherens junctions containing N-cadherin and its associated protein ␤-catenin exist between lens fiber cells (13-16), and may play important roles in lens development and function.Although cell-cell interaction is critical for maintaining lens transparency, little is known about the molecular mechanisms underlying these interactions. We have identified an unexpected regulator of lens fiber cell-cell interaction, the axon guidance molecule ephrin-A5 (17)(18)(19), and have shown that the loss...

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Congenital hyperinsulinism/hyperammonemia (HI/HA) syndrome is caused by an activation mutation of glutamate dehydrogenase 1 (GDH1), a mitochondrial enzyme responsible for the reversible interconversion between glutamate and α-ketoglutarate. The syndrome presents clinically with hyperammonemia, significant episodic hypoglycemia, seizures, and a frequent incidences of developmental and learning defects. Clinical research has implicated that although some of the developmental and neurological defects may be attributed to hypoglycemia, some characteristics cannot be ascribed to low glucose and as hyperammonemia is generally mild and asymptomatic, there exists the possibility that altered GDH1 activity within the brain leads to some clinical changes. GDH1 is allosterically regulated by many factors, and has been shown to be inhibited by the ADP-ribosyltransferase sirtuin 4 (SIRT4), a mitochondrially localized sirtuin. Here we show that SIRT4 is localized to mitochondria within the brain. SIRT4 is highly expressed in glial cells, specifically astrocytes, in the postnatal brain and in radial glia during embryogenesis. Furthermore, SIRT4 protein decreases in expression during development. We show that factors known to allosterically regulate GDH1 alter gliogenesis in CTX8 cells, a novel radial glial cell line. We find that SIRT4 and GDH1 overexpression play antagonistic roles in regulating gliogenesis and that a mutant variant of GDH1 found in HI/HA patients accelerates the development of glia from cultured radial glia cells.

Neurons have a limited capacity for heat shock protein (HSP) induction and are vulnerable to the pathogenic consequence of protein misfolding and aggregation as seen in age-related neurodegenerative diseases. Sirtuin 1 (SIRT1), an NAD(+) -dependent lysine deacetylase with important biological functions, has been shown to sustain the DNA-binding state of HSF1 for HSP induction. Here we show that differentiation and maturation of embryonic cortical neurons and N2a neuroprogenitor cells is associated with decreases in SIRT1 expression and heat shock-dependent induction of HSP70 protein. Tests of a pharmacological activator and an inhibitor of SIRT1 affirm the regulatory role of SIRT1 in HSP70 induction. Protein cross-linking studies show that nuclear SIRT1 and HSF1 form a co-migrating high molecular weight complex upon stress. The use of retroviral vectors to manipulate SIRT1 expression in N2a cells show that shRNA-mediated knock down of SIRT1 causes spontaneous neurite outgrowth coincident with reduced growth rate and decreased induction of hsp70-reporter gene, whereas SIRT1 over-expression blocks the induced neural differentiation of N2a cells. Our results suggest that decreased SIRT1 expression is conducive to neuronal differentiation and this decrease contributes to the attenuated induction of HSPs in neurons.