DIP-2 suppresses ectopic neurite sprouting and axonal regeneration in mature neurons.

Neuronal morphology and circuitry established during early development must often be maintained over the entirety of animal lifespans. Compared with neuronal development, the mechanisms that maintain mature neuronal structures and architecture are little understood. The conserved disco-interacting protein 2 (DIP2) consists of a DMAP1-binding domain and two adenylate-forming domains (AFDs). We show that the Caenorhabditis elegans DIP-2 maintains morphology of mature neurons. dip-2 loss-of-function mutants display a progressive increase in ectopic neurite sprouting and branching during late larval and adult life. In adults, dip-2 also inhibits initial stages of axon regeneration cell autonomously and acts in parallel to DLK-1 MAP kinase and EFA-6 pathways. The function of DIP-2 in maintenance of neuron morphology and in axon regrowth requires its AFD domains and is independent of its DMAP1-binding domain. Our findings reveal a new conserved regulator of neuronal morphology maintenance and axon regrowth after injury

DIP-2 suppresses ectopic neurite sprouting and axonal regeneration in mature neurons.

Neuronal morphology and circuitry established during early development must often be maintained over the entirety of animal lifespans. Compared with neuronal development, the mechanisms that maintain mature neuronal structures and architecture are little understood. The conserved disco-interacting protein 2 (DIP2) consists of a DMAP1-binding domain and two adenylate-forming domains (AFDs). We show that the Caenorhabditis elegans DIP-2 maintains morphology of mature neurons. dip-2 loss-of-function mutants display a progressive increase in ectopic neurite sprouting and branching during late larval and adult life. In adults, dip-2 also inhibits initial stages of axon regeneration cell autonomously and acts in parallel to DLK-1 MAP kinase and EFA-6 pathways. The function of DIP-2 in maintenance of neuron morphology and in axon regrowth requires its AFD domains and is independent of its DMAP1-binding domain. Our findings reveal a new conserved regulator of neuronal morphology maintenance and axon regrowth after injury

A Farnesyltransferase Acts to Inhibit Ectopic Neurite Formation in C. elegans.

Genetic pathways that regulate nascent neurite formation play a critical role in neuronal morphogenesis. The core planar cell polarity components VANG-1/Van Gogh and PRKL-1/Prickle are involved in blocking inappropriate neurite formation in a subset of motor neurons in C. elegans. A genetic screen for mutants that display supernumerary neurites was performed to identify additional factors involved in this process. This screen identified mutations in fntb-1, the β subunit of farnesyltransferase. We show that fntb-1 is expressed in neurons and acts cell-autonomously to regulate neurite formation. Prickle proteins are known to be post-translationally modified by farnesylation at their C-terminal CAAX motifs. We show that PRKL-1 can be recruited to the plasma membrane in both a CAAX-dependent and CAAX-independent manner but that PRKL-1 can only inhibit neurite formation in a CAAX-dependent manner

DIP-2 suppresses ectopic neurite sprouting and axonal regeneration in mature neurons.

Neuronal morphology and circuitry established during early development must often be maintained over the entirety of animal lifespans. Compared with neuronal development, the mechanisms that maintain mature neuronal structures and architecture are little understood. The conserved disco-interacting protein 2 (DIP2) consists of a DMAP1-binding domain and two adenylate-forming domains (AFDs). We show that the Caenorhabditis elegans DIP-2 maintains morphology of mature neurons. dip-2 loss-of-function mutants display a progressive increase in ectopic neurite sprouting and branching during late larval and adult life. In adults, dip-2 also inhibits initial stages of axon regeneration cell autonomously and acts in parallel to DLK-1 MAP kinase and EFA-6 pathways. The function of DIP-2 in maintenance of neuron morphology and in axon regrowth requires its AFD domains and is independent of its DMAP1-binding domain. Our findings reveal a new conserved regulator of neuronal morphology maintenance and axon regrowth after injury

A genetic screen for VC4 and VC5 neurite outgrowth defective (Nde) mutants.

<p>(A) Worm schematic showing the position of VC1–6 (ventral view). VC4 and VC5 neurons extend neurites along a LR axis and VC1–3 and VC6 extend neurites along the AP body axis. (A) Wild-type VC4 and VC5. (B) In <i>prkl-1</i> and <i>vang-1</i> (not shown) mutants, VC4 and VC5 display ectopic neurites (arrows) directed along the AP axis. (D) Schematic outline of a forward genetic screen for VC4 and VC5 neurite outgrowth defective (<i>nde</i>) mutants. (E) Quantification of ectopic neurite defects in <i>nde</i> mutants. Graph shows the percentage of VC4 and VC5 neurons (pooled) displaying at least one ectopic neurite. Error bars represent the 95% confidence interval of the proportion. For each genotype, n>160. (F and G) Representative images of VC4 and VC5 in <i>nde-4</i> (F) and <i>nde-5</i> (G) mutants. Ectopic neurites are marked with arrows. All lines contain the <i>cyIs4[Pcat-1</i>::<i>GFP]</i> reporter background. All images are ventral views. Scale bars, 10μm.</p

FNTB-1 missense mutations mapped to human FTase.

<p>(A) Crystal structure of the human protein FTase (PDB ID: 1S63) heterodimer (α subunit; red and β subunit; blue). Yellow circles mark the location of residues C104, G250, S301 and Q336 of the worm FTase. Lipid substrate farnesyl diphosphate, (FPP: isoprenoid moiety, green; diphosphate, red) and zinc ion (grey sphere) mark the site of catalysis. (B) Enlarged view of the FTase active site. G250 (yellow surface representation) makes van der Waals contact with FPP (grey surface). Any mutation at position 250 would result in a larger side chain that would affect FPP binding. (C) Enlarged view around residue C92 in worm FTase (C104 human FTase; green stick). The four major rotamers for the C104Y mutation and their corresponding van der Waals surfaces are shown in yellow. A van der Waals surface calculated around the alpha subunit (red) and beta subunit (blue) is shown. Any tyrosine rotamer results in a steric class with the protein. This mutation is predicted to compromise the packing and the stability of the enzyme.</p

VC4 and VC5 neurite outgrowth defective (<i>nde</i>) mutants.

<p>VC4 and VC5 neurite outgrowth defective (<i>nde</i>) mutants.</p

The PRKL-1 CAAX motif is important for membrane localization.

<p>(A) Transgene expression of a GFP::PRKL-1 fusion shows punctate localization on the plasma membrane of VC4 and VC5 (many puncta) at the early L4 stage. Representative images showing the localization of a CAAX-deleted PRKL-1::GFP fusion in a <i>wild-type</i> (<i>wt</i>) background (B) and full length GFP::PRKL-1 in a <i>vang-1</i> mutant background (C). In both panels B and C, a localization pattern resembling full length PRKL-1 (many puncta) or diminished membrane localization (few or no puncta, arrows) are observed. (D) Expression of a CAAX-deleted PRKL-1 construct in a <i>vang-1</i> mutant background shows loss of plasma membrane localization in VC4 and VC5 (few/no puncta). (E) Quantification of full length GFP::PRKL-1 and GFP::PRKL-1 ΔCAAX membrane distribution in <i>wt</i> and <i>vang-1</i> mutants in early L4 stage VC4 and VC5.</p

Sequence changes in <i>nde</i> mutants.

<p>Sequence changes in <i>nde</i> mutants.</p