The large and small SPEN family proteins stimulate axon outgrowth during neurosecretory cell remodeling in Drosophila.

Split ends (SPEN) is the founding member of a well conserved family of nuclear proteins with critical functions in transcriptional regulation and the post-transcriptional processing and nuclear export of transcripts. In animals, the SPEN proteins fall into two size classes that perform either complementary or antagonistic functions in different cellular contexts. Here, we show that the two Drosophila representatives of this family, SPEN and Spenito (NITO), regulate metamorphic remodeling of the CCAP/bursicon neurosecretory cells.

Regulatory Mechanisms of Metamorphic Neuronal Remodeling Revealed Through a Genome-Wide Modifier Screen in .

During development, neuronal remodeling shapes neuronal connections to establish fully mature and functional nervous systems. Our previous studies have shown that the RNA-binding factor () is an important regulator of neuronal remodeling during metamorphosis in , and loss of leads to smaller soma size and fewer neurites in a stage-dependent manner. To shed light on the mechanisms by which regulates neuronal remodeling, we conducted a genetic modifier screen for suppressors of -dependent wing expansion defects and cellular morphological defects in a set of peptidergic neurons, the bursicon neurons, that promote posteclosion wing expansion.

Neuronal remodeling during metamorphosis is regulated by the alan shepard (shep) gene in Drosophila melanogaster.

Peptidergic neurons are a group of neuronal cells that synthesize and secrete peptides to regulate a variety of biological processes. To identify genes controlling the development and function of peptidergic neurons, we conducted a screen of 545 splice-trap lines and identified 28 loci that drove expression in peptidergic neurons when crossed to a GFP reporter transgene. Among these lines, an insertion in the alan shepard (shep) gene drove expression specifically in most peptidergic neurons.

Vesicle capture, not delivery, scales up neuropeptide storage in neuroendocrine terminals.

Neurons vary in their capacity to produce, store, and release neuropeptides packaged in dense-core vesicles (DCVs). Specifically, neurons used for cotransmission have terminals that contain few DCVs and many small synaptic vesicles, whereas neuroendocrine neuron terminals contain many DCVs. Although the mechanistic basis for presynaptic variation is unknown, past research demonstrated transcriptional control of neuropeptide synthesis suggesting that supply from the soma limits presynaptic neuropeptide accumulation.

Insulin signaling regulates neurite growth during metamorphic neuronal remodeling.

Although the growth capacity of mature neurons is often limited, some neurons can shift through largely unknown mechanisms from stable maintenance growth to dynamic, organizational growth (e.g. to repair injury, or during development transitions).

Cryptocephal, the Drosophila melanogaster ATF4, is a specific coactivator for ecdysone receptor isoform B2.

The ecdysone receptor is a heterodimer of two nuclear receptors, the Ecdysone receptor (EcR) and Ultraspiracle (USP). In Drosophila melanogaster, three EcR isoforms share common DNA and ligand-binding domains, but these proteins differ in their most N-terminal regions and, consequently, in the activation domains (AF1s) contained therein. The transcriptional coactivators for these domains, which impart unique transcriptional regulatory properties to the EcR isoforms, are unknown.

Synaptic neuropeptide release induced by octopamine without Ca2+ entry into the nerve terminal.

Synaptic release of neurotransmitters is evoked by activity-dependent Ca(2+) entry into the nerve terminal. However, here it is shown that robust synaptic neuropeptide release from Drosophila motoneurons is evoked in the absence of extracellular Ca(2+) by octopamine, the arthropod homolog to norepinephrine. Genetic and pharmacology experiments demonstrate that this surprising peptidergic transmission requires cAMP-dependent protein kinase, with only a minor contribution of exchange protein activated by cAMP (epac).

The buzz on fly neuronal remodeling.

Hormone-dependent rewiring of axons and dendrites is a conserved feature of nervous system development and plasticity. During metamorphosis in insects, steroid hormones (the ecdysteroids) and terpenoid hormones (the juvenile hormones) regulate extensive remodeling of the nervous system. These changes retool the nervous system for new behavioral and physiological functions that are required for the adult stage of the life cycle.

A Drosophila gain-of-function screen for candidate genes involved in steroid-dependent neuroendocrine cell remodeling.

The normal functioning of neuroendocrine systems requires that many neuropeptidergic cells change, to alter transmitter identity and concentration, electrical properties, and cellular morphology in response to hormonal cues. During insect metamorphosis, a pulse of circulating steroids, ecdysteroids, governs the dramatic remodeling of larval neurons to serve adult-specific functions. To identify molecular mechanisms underlying metamorphic remodeling, we conducted a neuropeptidergic cell-targeted, gain-of-function genetic screen.

Presynaptic ryanodine receptor-activated calmodulin kinase II increases vesicle mobility and potentiates neuropeptide release.

Although it has been postulated that vesicle mobility is increased to enhance release of transmitters and neuropeptides, the mechanism responsible for increasing vesicle motion in nerve terminals and the effect of perturbing this mobilization on synaptic plasticity are unknown. Here, green fluorescent protein-tagged dense-core vesicles (DCVs) are imaged in Drosophila motor neuron terminals, where DCV mobility is increased for minutes after seconds of activity. Ca2+-induced Ca2+ release from presynaptic endoplasmic reticulum (ER) is shown to be necessary and sufficient for sustained DCV mobilization.

Regulation of secretory protein expression in mature cells by DIMM, a basic helix-loop-helix neuroendocrine differentiation factor.

During differentiation, neuroendocrine cells acquire highly amplified capacities to synthesize neuropeptides to overcome dilution of these signals in the general circulation. Once mature, the normal functioning of integrated physiological systems requires that neuroendocrine cells remain plastic to dramatically alter neuropeptide expression for long periods in response to hormonal and electrical cues. The mechanisms underlying the long-term regulation of neuroendocrine systems are poorly understood.

Transcriptional regulation of neuropeptide and peptide hormone expression by the Drosophila dimmed and cryptocephal genes.

The regulation of neuropeptide and peptide hormone gene expression is essential for the development and function of neuroendocrine cells in integrated physiological networks. In insects, a decline in circulating ecdysteroids triggers the activation of a neuroendocrine system to stimulate ecdysis, the behaviors used to shed the old cuticle at the culmination of each molt. Here we show that two evolutionarily conserved transcription factor genes, the basic helix-loop-helix (bHLH) gene dimmed (dimm) and the basic-leucine zipper (bZIP) gene cryptocephal (crc), control expression of diverse neuropeptides and peptide hormones in Drosophila.

Nearly neutral secretory vesicles in Drosophila nerve terminals.

The acidity of mammalian secretory vesicles drives concentration and processing of their contents. Here, pH-sensitive green fluorescent protein (GFP) variants show that the > or =30-fold (H+) difference between secretory vesicles (pH < or = 5.7) and the cytoplasm (pH = 7.

Activity-dependent liberation of synaptic neuropeptide vesicles.

Despite the importance of neuropeptide release, which is evoked by long bouts of action potential activity and which regulates behavior, peptidergic vesicle movement has not been examined in living nerve terminals. Previous in vitro studies have found that secretory vesicle motion at many sites of release is constitutive: Ca(2+) does not affect the movement of small synaptic vesicles in nerve terminals or the movement of large dense core vesicles in growth cones and endocrine cells. However, in vivo imaging of a neuropeptide, atrial natriuretic factor, tagged with green fluorescent protein in larval Drosophila melanogaster neuromuscular junctions shows that peptidergic vesicle behavior in nerve terminals is sensitive to activity-induced Ca(2+) influx.

The bHLH protein Dimmed controls neuroendocrine cell differentiation in Drosophila.

Neuroendocrine cells are specialized to produce, maintain and release large stores of secretory peptides. We show that the Drosophila dimmed/Mist1 bHLH gene confers such a pro-secretory phenotype on neuroendocrine cells. dimmed is expressed selectively in central and peripheral neuroendocrine cells.