RanBP9 is known to act as a scaffolding protein bringing together a variety of cell surface receptors and intracellular targets thereby regulating functions as diverse as neurite and axonal outgrowth, cell morphology, cell proliferation, myelination, gonad development, myofibrillogenesis and migration of neuronal precursors. Though RanBP9 is ubiquitously expressed in all tissues, brain is one of the organs with the highest expression levels of RanBP9. In the neurons, RanBP9 is localized mostly in the cytoplasm but also in the neurites and dendritic processes. We recently demonstrated that RanBP9 plays pathogenic role in Alzheimer’s disease. To understand the role of RanBP9 in the brain, here we generated RanBP9 null mice by gene-trap based strategy. Most of Ran−/− mice die neonatally due to defects in the brain growth and development. The major defects include smaller cortical plate (CP), robustly enlarged lateral ventricles (LV) and reduced volume of hippocampus (HI). The lethal phenotype is due to a suckling defect as evidenced by lack of milk in the stomachs even several hours after parturition. The complex somatosensory system which is required for a behavior such as suckling appears to be compromised in Ran−/− mice due to under developed CP. Most importantly, RanBP9 phenotype is similar to ERK1/2 double knockout and the neural cell adhesion receptor, L1CAM knockout mice. Both ERK1 and L1CAM interact with RanBP9. Thus, RanBP9 appears to control brain growth and development through signaling mechanisms involving ERK1 and L1CAM receptor.

The ubiquitin ligase Highwire has a conserved role in synapse formation. Here, we show that Highwire coordinates several facets of central synapse formation in the giant fiber system, including axon termination, axon pruning, and synaptic function. Despite the similarities to the fly neuromuscular junction, the role of Highwire and the underlying signaling pathways are distinct in the fly's giant fiber system. During development, branching of the giant fiber presynaptic terminal occurs and, normally, the transient branches are pruned away. However, in mutants these ectopic branches persist, indicating that Highwire promotes axon pruning. mutants also exhibit defects in synaptic function. Highwire promotes axon pruning and synaptic function cell-autonomously by attenuating a mitogen-activated protein kinase pathway including Wallenda, c-Jun N-terminal kinase/Basket, and the transcription factor Jun. We also show a novel role for Highwire in non-cell autonomous promotion of synaptic function from the midline glia. Highwire also regulates axon termination in the giant fibers, as mutant axons exhibit severe overgrowth beyond the pruning defect. This excessive axon growth is increased by manipulating Fos expression in the cells surrounding the giant fiber terminal, suggesting that Fos regulates a -synaptic signal that promotes giant fiber axon growth.

We previously demonstrated that RanBP9 overexpression increased Aβ generation and amyloid plaque burden, subsequently leading to robust reductions in the levels of several synaptic proteins as well as deficits in the learning and memory skills in a mouse model of Alzheimer's disease (AD). In the present study, we found striking reduction of spinophilin-immunoreactive puncta (52%, p<0.001) and spinophilin area (62.5%, p<0.001) in the primary cortical neurons derived from RanBP9 transgenic mice (RanBP9-Tg) compared to wild-type (WT) neurons. Similar results were confirmed in WT cortical neurons transfected with EGFP-RanBP9. At 6-months of age, the total spine density in the cortex of RanBP9 single transgenic, APΔE9 double transgenic and APΔE9/RanBP9 triple transgenic mice were similar to WT mice. However, in the hippocampus the spine density was significantly reduced (27%, p<0.05) in the triple transgenic mice compared to WT mice due to reduced number of thin spines (33%, p<0.05) and mushroom spines (22%, p<0.05). This suggests that RanBP9 overexpression in the APΔE9 mice accelerates loss of spines and that hippocampus is more vulnerable. At 12-months of age, cortex showed significant reductions in total spine density in the RanBP9 (22%, p<0.05), APΔE9 (19%, p<0.05) and APΔE9/RanBP9 (33%, p<0.01) mice compared to WT controls due to reductions in mushroom and thin spines. Similarly, in the hippocampus the total spine density was reduced in the RanBP9 (23%, p<0.05), APΔE9 (26%, p<0.05) and APΔE9/RanBP9 (39%, p<0.01) mice due to reductions in thin and mushroom spines. Most importantly, RanBP9 overexpression in the APΔE9 mice further exacerbated the reductions in spine density in both the cortex (14%, p<0.05) and the hippocampus (16%, p<0.05). Because dendritic spines are considered physical traces of memory, loss of spines due to RanBP9 provided the physical basis for the learning and memory deficits. Since RanBP9 protein levels are increased in AD brains, RanBP9 might play a crucial role in the loss of spines and synapses in AD.

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