Synaptic vesicle distribution by conveyor belt.

The equal distribution of synaptic vesicles among synapses along the axon is critical for robust neurotransmission. Wong et al. show that the continuous circulation of synaptic vesicles throughout the axon driven by molecular motors ultimately yields this even distribution.

Dynactin subunit p150(Glued) is a neuron-specific anti-catastrophe factor.

Regulation of microtubule dynamics in neurons is critical, as defects in the microtubule-based transport of axonal organelles lead to neurodegenerative disease. The microtubule motor cytoplasmic dynein and its partner complex dynactin drive retrograde transport from the distal axon. We have recently shown that the p150(Glued) subunit of dynactin promotes the initiation of dynein-driven cargo motility from the microtubule plus-end. Because plus end-localized microtubule-associated proteins like p150(Glued) may also modulate the dynamics of microtubules, we hypothesized that p150(Glued) might promote cargo initiation by stabilizing the microtubule track. Here, we demonstrate in vitro using assembly assays and TIRF microscopy, and in primary neurons using live-cell imaging, that p150(Glued) is a potent anti-catastrophe factor for microtubules. p150(Glued) alters microtubule dynamics by binding both to microtubules and to tubulin dimers; both the N-terminal CAP-Gly and basic domains of p150(Glued) are required in tandem for this activity. p150(Glued) is alternatively spliced in vivo, with the full-length isoform including these two domains expressed primarily in neurons. Accordingly, we find that RNAi of p150(Glued) in nonpolarized cells does not alter microtubule dynamics, while depletion of p150(Glued) in neurons leads to a dramatic increase in microtubule catastrophe. Strikingly, a mutation in p150(Glued) causal for the lethal neurodegenerative disorder Perry syndrome abrogates this anti-catastrophe activity. Thus, we find that dynactin has multiple functions in neurons, both activating dynein-mediated retrograde axonal transport and enhancing microtubule stability through a novel anti-catastrophe mechanism regulated by tissue-specific isoform expression; disruption of either or both of these functions may contribute to neurodegenerative disease.

Ordered recruitment of dynactin to the microtubule plus-end is required for efficient initiation of retrograde axonal transport.

Long-range retrograde axonal transport in neurons is driven exclusively by the microtubule motor cytoplasmic dynein. The efficient initiation of dynein-mediated transport from the distal axon is critical for normal neuronal function, and neurodegenerative disease-associated mutations have been shown to specifically disrupt this process. Here, we examine the role of dynamic microtubules and microtubule plus-end binding proteins (+TIPs) in the initiation of dynein-mediated retrograde axonal transport using live-cell imaging of cargo motility in primary mouse dorsal root ganglion neurons. We show that end-binding (EB)-positive dynamic microtubules are enriched in the distal axon. The +TIPs EB1, EB3, and cytoplasmic linker protein-170 (CLIP-170) interact with these dynamic microtubules, recruiting the dynein activator dynactin in an ordered pathway, leading to the initiation of retrograde transport by the motor dynein. Once transport has initiated, however, neither the EBs nor CLIP-170 are required to maintain transport flux along the mid-axon. In contrast, the +TIP Lis1 activates transport through a distinct mechanism and is required to maintain processive organelle transport along both the distal and mid-axon. Further, we show that the EB/CLIP-170/dynactin-dependent mechanism is required for the efficient initiation of transport from the distal axon for multiple distinct cargos, including mitochondria, Rab5-positive early endosomes, late endosomes/lysosomes, and TrkA-, TrkB-, and APP-positive organelles. Our observations indicate that there is an essential role for +TIPs in the regulation of retrograde transport initiation in the neuron.

SVIP is a molecular determinant of lysosomal dynamic stability, neurodegeneration and lifespan.

Missense mutations in Valosin-Containing Protein (VCP) are linked to diverse degenerative diseases including IBMPFD, amyotrophic lateral sclerosis (ALS), muscular dystrophy and Parkinson's disease. Here, we characterize a VCP-binding co-factor (SVIP) that specifically recruits VCP to lysosomes. SVIP is essential for lysosomal dynamic stability and autophagosomal-lysosomal fusion. SVIP mutations cause muscle wasting and neuromuscular degeneration while muscle-specific SVIP over-expression increases lysosomal abundance and is sufficient to extend lifespan in a context, stress-dependent manner. We also establish multiple links between SVIP and VCP-dependent disease in our Drosophila model system. A biochemical screen identifies a disease-causing VCP mutation that prevents SVIP binding. Conversely, over-expression of an SVIP mutation that prevents VCP binding is deleterious. Finally, we identify a human SVIP mutation and confirm the pathogenicity of this mutation in our Drosophila model. We propose a model for VCP disease based on the differential, co-factor-dependent recruitment of VCP to intracellular organelles.

Dual separable feedback systems govern firing rate homeostasis.

Firing rate homeostasis (FRH) stabilizes neural activity. A pervasive and intuitive theory argues that a single variable, calcium, is detected and stabilized through regulatory feedback. A prediction is that ion channel gene mutations with equivalent effects on neuronal excitability should invoke the same homeostatic response. In agreement, we demonstrate robust FRH following either elimination of Kv4/Shal protein or elimination of the Kv4/Shal conductance. However, the underlying homeostatic signaling mechanisms are distinct. Eliminating Shal protein invokes Krüppel-dependent rebalancing of ion channel gene expression including enhanced slo, Shab, and Shaker. By contrast, expression of these genes remains unchanged in animals harboring a CRISPR-engineered, Shal pore-blocking mutation where compensation is achieved by enhanced IKDR. These different homeostatic processes have distinct effects on homeostatic synaptic plasticity and animal behavior. We propose that FRH includes mechanisms of proteostatic feedback that act in parallel with activity-driven feedback, with implications for the pathophysiology of human channelopathies.

Angiotensin II AT1 receptor blockade decreases lipopolysaccharide-induced inflammation in the rat adrenal gland.

Peripheral administration of bacterial endotoxin [lipopolysaccharide (LPS)] to rodents produces an innate immune response and hypothalamic-pituitary-adrenal axis stimulation. Renin-angiotensin-aldosterone system inhibition by angiotensin II AT1 receptor blockade has antiinflammatory effects in the vasculature. We studied whether angiotensin II receptor blockers (ARBs) prevent the LPS response. We focused on the adrenal gland, one organ responsive to LPS and expressing a local renin-angiotensin-aldosterone system. LPS (50 microg/kg, ip) produced a generalized inflammatory response with increased release of TNF-alpha and IL-6 to the circulation, enhanced adrenal aldosterone synthesis and release, and enhanced adrenal cyclooxygenase-2, IL-6, and TNF-alpha gene expression. ACTH and corticosterone release were also increased by LPS. Pretreatment with the ARB candesartan (1 mg/kg.d, sc for 3 d before the LPS administration) decreased LPS-induced cytokine release to the circulation, adrenal aldosterone synthesis and release, and cyclooxygenase-2 and IL-6 gene expression. Candesartan did not prevent the LPS-induced ACTH and corticosterone release. Our results suggest that AT1 receptors are essential for the development of the full innate immune and stress responses to bacterial endotoxin. The ARB decreased the general peripheral inflammatory response to LPS, partially decreased the inflammatory response in the adrenal gland, prevented the release of the pro-inflammatory hormone aldosterone, and protected the antiinflammatory effects of glucocorticoid release. An unrestricted innate immune response to the bacterial endotoxin may have deleterious effects for the organism and may lead to development of chronic inflammatory disease. We postulate that the ARBs may have therapeutic effects on inflammatory conditions.

Axonal transport: cargo-specific mechanisms of motility and regulation.

Axonal transport is essential for neuronal function, and many neurodevelopmental and neurodegenerative diseases result from mutations in the axonal transport machinery. Anterograde transport supplies distal axons with newly synthesized proteins and lipids, including synaptic components required to maintain presynaptic activity. Retrograde transport is required to maintain homeostasis by removing aging proteins and organelles from the distal axon for degradation and recycling of components. Retrograde axonal transport also plays a major role in neurotrophic and injury response signaling. This review provides an overview of axonal transport pathways and discusses their role in neuronal function.

Dynactin is required for transport initiation from the distal axon.

Dynactin is a required cofactor for the minus-end-directed microtubule motor cytoplasmic dynein. Mutations within the highly conserved CAP-Gly domain of dynactin cause neurodegenerative disease. Here, we show that the CAP-Gly domain is necessary to enrich dynactin at the distal end of primary neurons. While the CAP-Gly domain is not required for sustained transport along the axon, we find that the distal accumulation facilitates the efficient initiation of retrograde vesicular transport from the neurite tip. Neurodegenerative disease mutations in the CAP-Gly domain prevent the distal enrichment of dynactin thereby inhibiting the initiation of retrograde transport. Thus, we propose a model in which distal dynactin is a key mediator in promoting the interaction among the microtubule, dynein motor, and cargo for the efficient initiation of transport. Mutations in the CAP-Gly domain disrupt the formation of the motor-cargo complex, highlighting the specific defects in axonal transport that may lead to neurodegeneration.

Retrograde axonal transport: pathways to cell death?

Active transport along the axon is crucial to the neuron. Motor-driven transport supplies the distal synapse with newly synthesized proteins and lipids, and clears damaged or misfolded proteins. Microtubule motors also drive long-distance signaling along the axon via signaling endosomes. Although positive signaling initiated by neurotrophic factors has been well-studied, recent research has focused on stress-signaling along the axon. Here, the connections between axonal transport alterations and neurodegeneration are discussed, including evidence for defective transport of vesicles, mitochondria, degradative organelles, and signaling endosomes in models of amyotrophic lateral sclerosis, Huntington's, Parkinson's and Alzheimer's disease. Defects in transport are sufficient to induce neurodegeneration, but recent progress suggests that changes in retrograde signaling pathways correlate with rapidly progressive neuronal cell death.