Regulation of actin filament assembly and disassembly in growth cone motility and axon guidance.

Directed outgrowth of axons is fundamental for the establishment of neuronal networks. Axon outgrowth is guided by growth cones, highly motile structures enriched in filamentous actin (F-actin) located at the axons' distal tips. Growth cones exploit F-actin-based protrusions to scan the environment for guidance cues, and they contain the sensory apparatus to translate guidance cue information into intracellular signaling cascades. These cascades act upstream of actin-binding proteins (ABP) and thereby control assembly and disassembly of F-actin. Spatiotemporally controlled F-actin dis-/assembly in growth cones steers the axon towards attractants and away from repellents, and it thereby navigates the axon through the developing nervous system. Hence, ABP that control F-actin dynamics emerged as critical regulators of neuronal network formation. In the present review article, we will summarize and discuss current knowledge of the mechanisms that control remodeling of the actin cytoskeleton in growth cones, focusing on recent progress in the field. Further, we will introduce tools and techniques that allow to study actin regulatory mechanism in growth cones.

Transient reduction in dendritic spine density in brain-specific profilin1 mutant mice is associated with behavioral deficits.

Actin filaments form the backbone of dendritic spines, the postsynaptic compartment of most excitatory synapses in the brain. Spine density changes affect brain function, and postsynaptic actin defects have been implicated in various neuropathies. It is mandatory to identify the actin regulators that control spine density. Based on previous studies, we hypothesized a role for the actin regulator profilin1 in spine formation. We report reduced hippocampal spine density in juvenile profilin1 mutant mice together with impairments in memory formation and reduced ultrasonic communication during active social behavior. Our results, therefore, underline a previously suggested function of profilin1 in controlling spine formation and behavior in juvenile mice.

Functional Redundancy of Cyclase-Associated Proteins CAP1 and CAP2 in Differentiating Neurons.

Cyclase-associated proteins (CAPs) are evolutionary-conserved actin-binding proteins with crucial functions in regulating actin dynamics, the spatiotemporally controlled assembly and disassembly of actin filaments (F-actin). Mammals possess two family members (CAP1 and CAP2) with different expression patterns. Unlike most other tissues, both CAPs are expressed in the brain and present in hippocampal neurons. We recently reported crucial roles for CAP1 in growth cone function, neuron differentiation, and neuron connectivity in the mouse brain. Instead, CAP2 controls dendritic spine morphology and synaptic plasticity, and its dysregulation contributes to Alzheimer's disease pathology. These findings are in line with a model in which CAP1 controls important aspects during neuron differentiation, while CAP2 is relevant in differentiated neurons. We here report CAP2 expression during neuron differentiation and its enrichment in growth cones. We therefore hypothesized that CAP2 is relevant not only in excitatory synapses, but also in differentiating neurons. However, CAP2 inactivation neither impaired growth cone morphology and motility nor neuron differentiation. Moreover, CAP2 mutant mice did not display any obvious changes in brain anatomy. Hence, differently from CAP1, CAP2 was dispensable for neuron differentiation and brain development. Interestingly, overexpression of CAP2 rescued not only growth cone size in CAP1-deficient neurons, but also their morphology and differentiation. Our data provide evidence for functional redundancy of CAP1 and CAP2 in differentiating neurons, and they suggest compensatory mechanisms in single mutant neurons.

Mutual functional dependence of cyclase-associated protein 1 (CAP1) and cofilin1 in neuronal actin dynamics and growth cone function.

Neuron connectivity depends on growth cones that navigate axons through the developing brain. Growth cones protrude and retract actin-rich structures to sense guidance cues. These cues control local actin dynamics and steer growth cones towards attractants and away from repellents, thereby directing axon outgrowth. Hence, actin binding proteins (ABPs) moved into the focus as critical regulators of neuron connectivity. We found cyclase-associated protein 1 (CAP1), an ABP with unknown brain function, abundant in growth cones. Super-resolution microscopy and live cell imaging combined with pharmacological approaches on hippocampal neurons from gene-targeted mice revealed a crucial role for CAP1 in actin dynamics that is critical for growth cone morphology and function. Growth cone defects in CAP1 knockout (KO) neurons compromised neuron differentiation and was associated with impaired neuron connectivity in CAP1-KO brains. Mechanistically, by rescue experiments in double KO neurons lacking CAP1 and the key actin regulator cofilin1, we demonstrated that CAP1 was essential for cofilin1 function in growth cone actin dynamics and morphology and vice versa. Together, we identified CAP1 as a novel actin regulator in growth cones that was relevant for neuron connectivity, and we demonstrated functional interdependence of CAP1 and cofilin1 in neuronal actin dynamics and growth cone function.

Glucose-regulated protein 75 determines ER-mitochondrial coupling and sensitivity to oxidative stress in neuronal cells.

The crosstalk between different organelles allows for the exchange of proteins, lipids and ions. Endoplasmic reticulum (ER) and mitochondria are physically linked and signal through the mitochondria-associated membrane (MAM) to regulate the transfer of Ca2+ from ER stores into the mitochondrial matrix, thereby affecting mitochondrial function and intracellular Ca2+ homeostasis. The chaperone glucose-regulated protein 75 (GRP75) is a key protein expressed at the MAM interface which regulates ER-mitochondrial Ca2+ transfer. Previous studies revealed that modulation of GRP75 expression largely affected mitochondrial integrity and vulnerability to cell death. In the present study, we show that genetic ablation of GRP75, by weakening ER-mitochondrial junctions, provided protection against mitochondrial dysfunction and cell death in a model of glutamate-induced oxidative stress. Interestingly, GRP75 silencing attenuated both cytosolic and mitochondrial Ca2+ overload in conditions of oxidative stress, blocked the formation of reactive oxygen species and preserved mitochondrial respiration. These data revealed a major role for GRP75 in regulating mitochondrial function, Ca2+ and redox homeostasis. In line, GRP75 overexpression enhanced oxidative cell death induced by glutamate. Overall, our findings suggest weakening ER-mitochondrial connectivity by GRP75 inhibition as a novel protective approach in paradigms of oxidative stress in neuronal cells.