Specific Aims: Syntaphilin (SNPH) is a neuron-specific and axon-targeted protein initially identified in our lab as a candidate inhibitor of presynaptic function (Lao et al., Neuron 2000). Our effort in generating SNPH KO mice has led to the discovery of a role for SNPH in the control of axonal mitochondrial motility (Kang et al., Cell 2008). Our study reveals that SNPH is required for maintaining a large number of axonal mitochondria in a stationary state through an interaction with the microtubule (MT)-based cytoskeleton. Deletion of the snph gene in mice results in a substantially higher proportion of axonal mitochondria in mobile state than that found in wild-type neurons, and reduced total and inter-bouton mitochondria density within axons. The snph mutant neurons exhibit enhanced short-term synaptic facilitation during prolonged stimulation by affecting calcium signaling at presynaptic boutons. This phenotype is fully rescued by reintroducing the snph gene into the mutant neurons. Thus, SNPH acts as a static anchor for docking/retaining mitochondria in axons and at synapses. These findings reveal for the first time a neuron-specific protein capable of docking axonal mitochondria within axons and near synapses. By applying a proteomic approach combined with time-lapse imaging, we further revealed that dynein light chain LC8 enhances the mitochondrial docking through its binding to SNPH (Chen et al., J Neuroscience 2009). SNPH recruits LC8 to axonal mitochondria via the 7-residue (ERAIQTD) LC8-binding motif and this interaction is independent of the dynein motor complex. Elevated LC8 expression in wild-type neurons inhibits axonal mitochondrial mobility. In contrast, this effect is not observed in snph-null neurons, suggesting the role of LC8 via its interaction with SNPH. CD spectrum analysis revealed that LC8 enhances mitochondrial docking by stabilizing an helix coiled-coil within the MT-binding domain of SNPH, thus providing new mechanistic insight into how SNPH and LC8 coordinately immobilize axonal mitochondria. Synaptic structure and function are highly plastic and undergo activity-dependent remodeling, thereby altering mitochondrial mobility and distribution. Axonal mitochondria exhibit the complex mobility pattern by coupling two opposing molecular motors kinesins and dynein and by attaching to docking/anchoring receptor SNPH. Identification of SNPH as a docking protein provides the molecular target for regulating the mobility and distribution of axonal mitochondria in response to neuronal activity. Our ongoing studies using the snph KO mouse will provide molecular details on how SNPH regulates mitochondrial transport and presynaptic function. In particular, our research will provide a molecular basis for addressing whether the motors and docking receptor share a single system of regulation or are modulated through distinct signal pathways. Mitochondrial dysfunction, altered mitochondrial dynamics and mobility, and perturbation of their turnover are involved in the pathology of several major neurodegenerative diseases including Parkinsons, Alzheimers, Huntingtons disease and amyotrophic lateral sclerosis (ALS). For example, ALS associated SOD1G93A and Huntingtons-linked Htt72Q transgenic mice display abnormal mitochondrial movement in neurons where mitochondria move more slowly, stop more frequently and travel shorter distances. However, whether defective mitochondrial transport plays a role in axonal degeneration remains largely unknown. Deleting the snph gene in mice recruits a majority of axonal mitochondria into an actively motile state. Our ongoing research is addressing (1) whether enhanced mobility of axonal mitochondria by crossing the snph (-/-) mice with the aforementioned diseased mouse lines contributes to efficient removal of dysfunctional mitochondria from synapses and distal axons and (2) whether enhanced mitochondrial mobility benefits the turnover of dysfunctional mitochondria via the mitophagy pathway or membrane fusion/fission dynamics. These studies will provide cellular and genetic clues as to whether manipulating mitochondrial transport and turnover may leads to new therapeutic approaches. Pursuing these investigations will advance our knowledge of fundamental processes that may affect human neurological disorders and is thus the very essence of the mission of the National Institute of Neurological Disorders and Stroke. Papers published from the lab related to the project: Guifang Lao, Volker Scheuss, Claudia M. Gerwin, Qingning Su, Sumiko Mochida, Jens Rettig, and Zu-Hang Sheng (2000). Neuron 25, 191-201. Qian Cai, Claudia Gerwin, and Zu-Hang Sheng (2005). Journal of Cell Biology 170, 959-969. Jian-Sheng Kang,Jin-Hua Tian, Philip Zald, Ping-Yue Pan, Cuiling Li, Chuxia Deng, and Zu-Hang Sheng (2008). Cell 132, 137-148. Yan-Min Chen, Claudia Gerwin, and Zu-Hang Sheng. (2009). Journal of Neuroscience 29, 9428-9437. Qian Cai, Zu-Hang Sheng (2009). Experimental Neurology 218, 257-267. Huan Ma, Qian Cai, Wenbo Lu, Zu-Hang Sheng (co-corresponding author), and Sumiko Mochida (2009). Journal of Neuroscience 29, 13019-13029. Qian Cai, Zu-Hang Sheng (2009). Neuron 61, 493-496.
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