Qian Cai

Snapin-Regulated Late Endosomal Transport Is Critical for Efficient Autophagy-Lysosomal Function in Neurons

Neuron maintenance and survival require late endocytic transport from distal processes to the soma where lysosomes are predominantly localized. Here, we report a role for Snapin in attaching dynein to late endosomes through its intermediate chain (DIC). snapin(-/-) neurons exhibit aberrant accumulation of immature lysosomes, clustering and impaired retrograde transport of late endosomes along processes, reduced lysosomal proteolysis due to impaired delivery of internalized proteins and hydrolase precursors from late endosomes to lysosomes, and impaired clearance of autolysosomes, combined with reduced neuron viability and neurodegeneration. The phenotypes are rescued by expressing the snapin transgene, but not the DIC-binding-defective Snapin-L99K mutant. Snapin overexpression in wild-type neurons enhances late endocytic transport and lysosomal function, whereas expressing the mutant defective in Snapin-DIC coupling shows a dominant-negative effect. Altogether, our study highlights new mechanistic insights into how Snapin-DIC coordinates retrograde transport and late endosomal-lysosomal trafficking critical for autophagy-lysosomal function, and thus neuronal homeostasis.

Syntabulin–Kinesin-1 Family Member 5B-Mediated Axonal Transport Contributes to Activity-Dependent Presynaptic Assembly

The mechanism by which microtubule-based axonal transport regulates activity-dependent presynaptic plasticity in developing neurons remains mostly unknown. Our previous studies established that syntabulin is an adaptor capable of conjoining the kinesin family member 5B (KIF5B) motor and syntaxin-1. We now report that the complex of syntaxin-1–syntabulin–KIF5B mediates axonal transport of the active zone (AZ) components essential for presynaptic assembly. Syntabulin associates with AZ precursor carriers and colocalizes and comigrates with green fluorescent protein (GFP)-Bassoon-labeled AZ transport cargos within developing axons. Knock-down of syntabulin or disruption of the syntaxin-1–syntabulin–KIF5B complex impairs the anterograde transport of GFP-Bassoon out of the soma and reduces the axonal densities of synaptic vesicle (SV) clusters and FM4-64 [N-(3-triethylammoniumpropyl)-4-(p-dibutylaminostyryl)pyridinium, dibromide] loading. Furthermore, syntabulin loss of function results in a reduction in both the amplitude of postsynaptic currents and the frequency of asynchronous quantal events, and abolishes the activity-induced recruitment of new GFP-Bassoon into the axons and subsequent coclustering with SVs. Consequently, syntabulin loss of function blocks the formation of new presynaptic boutons during activity-dependent synaptic plasticity in developing neurons. These studies establish that a kinesin motor–adaptor complex is critical for the anterograde axonal transport of AZ components, thus contributing to activity-dependent presynaptic assembly during neuronal development.

The role of snapin in neurosecretion: Snapin knock-out mice exhibit impaired calcium-dependent exocytosis of large dense-core vesicles in chromaffin cells

Identification of the molecules that regulate the priming of synaptic vesicles for fusion and the structural coupling of the calcium sensor with the soluble N-ethyl maleimide sensitive factor adaptor protein receptor (SNARE)-based fusion machinery is critical for understanding the mechanisms underlying calcium-dependent neurosecretion. Snapin binds to synaptosomal-associated protein 25 kDa (SNAP-25) and enhances the association of the SNARE complex with synaptotagmin. In the present study, we abolished snapin expression in mice and functionally evaluated the role of Snapin in neuroexocytosis. We found that the association of synaptotagmin-1 with SNAP-25 in brain homogenates of snapin mutant mice is impaired. Consequently, the absence of Snapin in embryonic chromaffin cells leads to a significant reduction of calcium-dependent exocytosis resulting from a decreased number of vesicles in releasable pools. Overexpression of Snapin fully rescued this inhibitory effect in the mutant cells. Furthermore, Snapin is relatively enriched in the purified large dense-core vesicles of chromaffin cells and associated with synaptotagmin-1. Thus, our biochemical and electrophysiological studies using snapin knock-out mice demonstrate that Snapin plays a critical role in modulating neurosecretion by stabilizing the release-ready vesicles.

KIF5B motor adaptor syntabulin maintains synaptic transmission in sympathetic neurons.

Newly synthesized synaptic proteins and mitochondria are transported along lengthy neuronal processes to assist in the proper assembly of developing synapses and activity-dependent remodeling of mature synapses. Neuronal transport is mediated by motor proteins that associate with their cargoes via adaptors and travel along the cytoskeleton within neuronal processes. Our previous studies in developing hippocampal neurons revealed that syntabulin acts as a KIF5B motor adaptor and mediates anterograde transport of presynaptic cargoes and mitochondria, presynaptic assembly, and activity-induced plasticity. Here, using cultured superior cervical ganglion neurons combined with manipulation of syntabulin expression or interference with its interaction with KIF5B, we uncover a crucial role for syntabulin in the maintenance of presynaptic function. Syntabulin loss-of-function delayed the appearance of synaptic activity in developing neurons and impaired synaptic transmission in mature neurons, including reduced basal activity, accelerated synaptic depression under high-frequency firing, slowed recovery rates after synaptic vesicle depletion, and impaired presynaptic short-term plasticity. These defects correlated with reduced mitochondrial distribution along neuronal processes and were rescued by the application of ATP within presynaptic neurons. These results suggest that syntabulin supports the axonal transport of mitochondria and concomitant ATP production at presynaptic terminals. ATP supply from locally stationed mitochondria is in turn necessary for the efficient mobilization of synaptic vesicles into the readily releasable pool. These findings emphasize the critical role of KIF5B-syntabulin-mediated axonal transport in the maintenance of presynaptic function and regulation of synaptic plasticity.

Snapin Recruits Dynein to BDNF-TrkB Signaling Endosomes for Retrograde Axonal Transport and Is Essential for Dendrite Growth of Cortical Neurons

Neurotrophin signaling is crucial for neuron growth. While the signaling endosomes hypothesis is one of the accepted models, the molecular machinery that drives retrograde axonal transport of TrkB signaling endosomes is largely unknown. In particular, mechanisms recruiting dynein to TrkB signaling endosomes have not been elucidated. Here, using snapin deficient mice and gene rescue experiments combined with compartmentalized cultures of live cortical neurons, we reveal that Snapin, as a dynein adaptor, mediates retrograde axonal transport of TrkB signaling endosomes. Such a role is essential for dendritic growth of cortical neurons. Deleting snapin or disrupting Snapin-dynein interaction abolishes TrkB retrograde transport, impairs BDNF-induced retrograde signaling from axonal terminals to the nucleus, and decreases dendritic growth. Such defects were rescued by reintroducing the snapin gene. Our study indicates that Snapin-dynein coupling is one of the primary mechanisms driving BDNF-TrkB retrograde transport, thus providing mechanistic insights into the regulation of neuronal growth and survival.

Axonal autophagosomes recruit dynein for retrograde transport through fusion with late endosomes

Late endosome-loaded dynein–snapin complexes drive amphisome retrograde transport upon fusion of autophagosomes with late endosomes in distal axons.

Moving or Stopping Mitochondria: Miro as a Traffic Cop by Sensing Calcium

Mitochondrial transport in neurons and their spatial distribution among synapses are directly correlated with synaptic activity. One paper in this issue of Neuron (MacAskill et al.) and two papers recently published in Cell (Wang and Schwarz) and PNAS (Saotome et al.) provide compelling evidence that Miro serves as a calcium sensor that controls mitochondrial mobility.

Alterations in Mitochondrial Quality Control in Alzheimer's Disease.

Mitochondrial dysfunction is one of the earliest and most prominent features in the brains of Alzheimer’s disease (AD) patients. Recent studies suggest that mitochondrial dysfunction plays a pivotal role in the pathogenesis of AD. Neurons are metabolically active cells, causing them to be particularly dependent on mitochondrial function for survival and maintenance. As highly dynamic organelles, mitochondria are characterized by a balance of fusion and fission, transport, and mitophagy, all of which are essential for maintaining mitochondrial integrity and function. Mitochondrial dynamics and mitophagy can therefore be identified as key pathways in mitochondrial quality control. Tremendous progress has been made in studying changes in these key aspects of mitochondrial biology in the vulnerable neurons of AD brains and mouse models, and the potential underlying mechanisms of such changes. This review highlights recent findings on alterations in the mitochondrial dynamics and mitophagy in AD and discusses how these abnormalities impact mitochondrial quality control and thus contribute to mitochondrial dysfunction in AD.

Snapin-Mediated BACE1 Retrograde Transport Is Essential for Its Degradation in Lysosomes and Regulation of APP Processing in Neurons

β site amyloid precursor protein (APP)-cleaving enzyme 1 (BACE1) is the major β secretase for generating β-amyloid (Aβ) peptides. The acidic environment of endosomes is optimal for β secretase activity. However, the mechanisms regulating BACE1 traffic from endosomes to lysosomes for degradation are largely unknown. Here, using snapin-deficient mice combined with gene rescue experiments, we reveal that Snapin, as a dynein motor adaptor for late endosomes, mediates BACE1 retrograde transport. hAPP mutant live neurons and mouse brains exhibited BACE1 accumulation within the altered late endocytic organelles and defective lysosomal targeting due to reduced Snapin-dynein coupling. Deleting snapin or disrupting Snapin-dynein coupling reduces BACE1 transport to lysosomes for degradation, thus enhancing APP processing. Overexpressing Snapin in hAPP neurons reduces β site cleavage of APP by enhancing BACE1 turnover. Altogether, our study provides mechanistic insights into the complex regulation of BACE1 level and activity and turnover through retrograde transport, thus controlling Aβ generation in neurons.

Snapin associates with late endocytic compartments and interacts with late endosomal SNAREs

Late endocytic membrane trafficking delivers target materials and newly synthesized hydrolases into lysosomes and is critical for maintaining an efficient degradation process and cellular homoeostasis. Although some features of late endosome-lysosome trafficking have been described, the mechanisms underlying regulation of this event remain to be elucidated. Our previous studies showed that Snapin, as a SNAP25 (25 kDa synaptosome-associated protein)-binding protein, plays a critical role in priming synaptic vesicles for synchronized fusion in neurons. In the present study, we report that Snapin also associates with late endocytic membranous organelles and interacts with the late endosome-targeted SNARE (soluble N-ethylmaleimide-sensitive factor-attachment protein receptor) complex. Using a genetic mouse model, we further discovered that Snapin is required to maintain a proper balance of the late endocytic protein LAMP-1 (lysosome-associated membrane protein-1) and late endosomal SNARE proteins syntaxin 8 and Vti1b (vesicle transport through interaction with target SNAREs homologue 1b). Deleting the snapin gene in mice selectively led to the accumulation of these proteins in late endocytic organelles. Thus our present study suggests that Snapin serves as an important regulator of the late endocytic fusion machinery, in addition to its established role in regulating synaptic vesicle fusion.