AMPK adapts metabolism to developmental energy requirement during dendrite pruning in Drosophila.

To reshape neuronal connectivity in adult stages, Drosophila sensory neurons prune their dendrites during metamorphosis using a genetic degeneration program that is induced by the steroid hormone ecdysone. Metamorphosis is a nonfeeding stage that imposes metabolic constraints on development. We find that AMP-activated protein kinase (AMPK), a regulator of energy homeostasis, is cell-autonomously required for dendrite pruning. AMPK is activated by ecdysone and promotes oxidative phosphorylation and pyruvate usage, likely to enable neurons to use noncarbohydrate metabolites such as amino acids for energy production. Loss of AMPK or mitochondrial deficiency causes specific defects in pruning factor translation and the ubiquitin-proteasome system. Our findings distinguish pruning from pathological neurite degeneration, which is often induced by defects in energy production, and highlight how metabolism is adapted to fit energy-costly developmental transitions.

Functions of Microtubule Disassembly during Neurite Pruning.

Large-scale neurite pruning, the developmentally regulated degeneration of axons or dendrites, is an important specificity mechanism during neuronal circuit formation. Pruning is usually restricted to single neurite branches and can occur by local degeneration or retraction. How this spatial regulation is achieved, and what triggers degeneration locally, are still poorly understood. At the cellular level, pruning involves local cytoskeleton disassembly before branch removal. Recent evidence suggests that microtubule disassembly is the local trigger and that the specific local microtubule organization of axons or dendrites determines where and how neurites degenerate. Based on these data, we propose a general model for spatial pruning regulation by microtubules and discuss how microtubule-associated proteins such as Tau could contribute to these regulatory aspects.

Spatial regulation of microtubule disruption during dendrite pruning in Drosophila.

Large-scale neurite pruning is an important specificity mechanism during neuronal morphogenesis. Drosophila sensory neurons prune their larval dendrites during metamorphosis. Pruning dendrites are severed in their proximal regions, but how this spatial information is encoded is not clear. Dendrite severing is preceded by local breakdown of dendritic microtubules through PAR-1-mediated inhibition of Tau. Here, we investigated spatial aspects of microtubule breakdown during dendrite pruning. Live imaging of fluorescently tagged tubulin shows that microtubule breakdown first occurs at proximal dendritic branchpoints, followed by breakdown at more distal branchpoints, suggesting that the process is triggered by a signal emanating from the soma. In fly dendrites, microtubules are arranged in uniformly oriented arrays where all plus ends face towards the soma. Mutants in kinesin-1 and -2, which are required for uniform microtubule orientation, show defects in microtubule breakdown and dendrite pruning. Our data suggest that the local microtubule organization at branchpoints determines where microtubule breakdown occurs. Local microtubule organization may therefore contribute spatial information for severing sites during dendrite pruning.

PAR-1 promotes microtubule breakdown during dendrite pruning in Drosophila.

Pruning of unspecific neurites is an important mechanism during neuronal morphogenesis. Drosophila sensory neurons prune their dendrites during metamorphosis. Pruning dendrites are severed in their proximal regions. Prior to severing, dendritic microtubules undergo local disassembly, and dendrites thin extensively through local endocytosis. Microtubule disassembly requires a katanin homologue, but the signals initiating microtubule breakdown are not known. Here, we show that the kinase PAR-1 is required for pruning and dendritic microtubule breakdown. Our data show that neurons lacking PAR-1 fail to break down dendritic microtubules, and PAR-1 is required for an increase in neuronal microtubule dynamics at the onset of metamorphosis. Mammalian PAR-1 is a known Tau kinase, and genetic interactions suggest that PAR-1 promotes microtubule breakdown largely via inhibition of Tau also in Drosophila Finally, PAR-1 is also required for dendritic thinning, suggesting that microtubule breakdown might precede ensuing plasma membrane alterations. Our results shed light on the signaling cascades and epistatic relationships involved in neurite destabilization during dendrite pruning.

The serpin PN1 is a feedback regulator of FGF signaling in germ layer and primary axis formation.

Germ layer formation and primary axis development rely on Fibroblast growth factors (FGFs). In Xenopus, the secreted serine protease HtrA1 induces mesoderm and posterior trunk/tail structures by facilitating the spread of FGF signals. Here, we show that the serpin Protease nexin-1 (PN1) is transcriptionally activated by FGF signals, suppresses mesoderm and promotes head development in mRNA-injected embryos. An antisense morpholino oligonucleotide against PN1 has the opposite effect and inhibits ectodermal fate. However, ectoderm and anterior head structures can be restored in PN1-depleted embryos when HtrA1 and FGF receptor activities are diminished, indicating that FGF signals negatively regulate their formation. We show that PN1 binds to and inhibits HtrA1, prevents degradation of the proteoglycan Syndecan 4 and restricts paracrine FGF/Erk signaling. Our data suggest that PN1 is a negative-feedback regulator of FGF signaling and has important roles in ectoderm and head development.