Molecular Remodeling of the Presynaptic Active Zone of Drosophila Photoreceptors via Activity-Dependent Feedback

SummaryNeural activity contributes to the regulation of the properties of synapses in sensory systems, allowing for adjustment to a changing environment. Little is known about how synaptic molecular components are regulated to achieve activity-dependent plasticity at central synapses. Here, we found that after prolonged exposure to natural ambient light the presynaptic active zone in Drosophila photoreceptors undergoes reversible remodeling, including loss of Bruchpilot, DLiprin-α, and DRBP, but not of DSyd-1 or Cacophony. The level of depolarization of the postsynaptic neurons is critical for the light-induced changes in active zone composition in the photoreceptors, indicating the existence of a feedback signal. In search of this signal, we have identified a crucial role of microtubule meshwork organization downstream of the divergent canonical Wnt pathway, potentially via Kinesin-3 Imac. These data reveal that active zone composition can be regulated in vivo and identify the underlying molecular machinery

Modular proteins from the Drosophila sallimus (sls) gene and their expression in muscles with different extensibility

The passive elasticity of the sarcomere in striated muscle is determined by large modular proteins, such as titin in vertebrates. In insects, the function of titin is divided between two shorter proteins, projectin and sallimus (Sls), which are the products of different genes. The Drosophila sallimus (sls) gene codes for a protein of 2 MDa. The N-terminal half of the protein is largely made up of immunoglobulin domains and unique sequence; the C-terminal half has two stretches of sequence similar to the elastic PEVK region of titin, and at the end of the molecule there is a region of tandem Ig and fibronectin domains. We have investigated splicing pathways of the sls gene and identified isoforms expressed in different muscle types, and at different stages of Drosophila development. The 5’ half of sls codes for zormin and kettin; both proteins contain Ig domains and can be expressed as separate isoforms, or as larger proteins linked to sequence downstream. There are multiple splicing pathways between the kettin region of sls and sequence coding for the two PEVK regions. All the resulting protein isoforms have sequence derived from the 3’ end of the sls gene. Splicing of exons varies at different stages of development. Kettin RNA is predominant in the embryo, and longer transcripts are expressed in larva, pupa and adult. Sls isoforms in the indirect flight muscle (IFM) are zormin, kettin and Sls(700), in which sequence derived from the end of the gene is spliced to kettin RNA. Zormin is in both M-line and Z-disc. Kettin and Sls(700) extend from the Z-disc to the ends of the thick filaments, though, Sls(700) is only in the myofibril core. These shorter isoforms would contribute to the high stiffness of IFM. Other muscles in the thorax and legs have longer Sls isoforms with varying amounts of PEVK sequence; all span the I-band to the ends of the thick filaments. In muscles with longer Ibands, the proportion of PEVK sequence would determine the extensibility of the sarcomere. Alternative Sls isoforms could regulate the stiffness of the many fibre types in Droso phila muscles

Prostaglandin E2 Signals Through PTGER2 to Regulate Sclerostin Expression

The Wnt signaling pathway is a robust regulator of skeletal homeostasis. Gain-of-function mutations promote high bone mass, whereas loss of Lrp5 or Lrp6 co-receptors decrease bone mass. Similarly, mutations in antagonists of Wnt signaling influence skeletal integrity, in an inverse relation to Lrp receptor mutations. Loss of the Wnt antagonist Sclerostin (Sost) produces the generalized skeletal hyperostotic condition of sclerosteosis, which is characterized by increased bone mass and density due to hyperactive osteoblast function. Here we demonstrate that prostaglandin E2 (PGE2), a paracrine factor with pleiotropic effects on osteoblasts and osteoclasts, decreases Sclerostin expression in osteoblastic UMR106.01 cells. Decreased Sost expression correlates with increased expression of Wnt/TCF target genes Axin2 and Tcf3. We also show that the suppressive effect of PGE2 is mediated through a cyclic AMP/PKA pathway. Furthermore, selective agonists for the PGE2 receptor EP2 mimic the effect of PGE2 upon Sost, and siRNA reduction in Ptger2 prevents PGE2-induced Sost repression. These results indicate a functional relationship between prostaglandins and the Wnt/β-catenin signaling pathway in bone

Control of Dendritic Morphogenesis by Trio in Drosophila melanogaster

Abl tyrosine kinase and its effectors among the Rho family of GTPases each act to control dendritic morphogenesis in Drosophila. It has not been established, however, which of the many GTPase regulators in the cell link these signaling molecules in the dendrite. In axons, the bifunctional guanine exchange factor, Trio, is an essential link between the Abl tyrosine kinase signaling pathway and Rho GTPases, particularly Rac, allowing these systems to act coordinately to control actin organization. In dendritic morphogenesis, however, Abl and Rac have contrary rather than reinforcing effects, raising the question of whether Trio is involved, and if so, whether it acts through Rac, Rho or both. We now find that Trio is expressed in sensory neurons of the Drosophila embryo and regulates their dendritic arborization. trio mutants display a reduction in dendritic branching and increase in average branch length, whereas over-expression of trio has the opposite effect. We further show that it is the Rac GEF domain of Trio, and not its Rho GEF domain that is primarily responsible for the dendritic function of Trio. Thus, Trio shapes the complexity of dendritic arbors and does so in a way that mimics the effects of its target, Rac

Sarcomere Formation Occurs by the Assembly of Multiple Latent Protein Complexes

The stereotyped striation of myofibrils is a conserved feature of muscle organization that is critical to its function. Although most components that constitute the basic myofibrils are well-characterized biochemically and are conserved across the animal kingdom, the mechanisms leading to the precise assembly of sarcomeres, the basic units of myofibrils, are poorly understood. To gain insights into this process, we investigated the functional relationships of sarcomeric protein complexes. Specifically, we systematically analyzed, using either RNAi in primary muscle cells or available genetic mutations, the organization of myofibrils in Drosophila muscles that lack one or more sarcomeric proteins. Our study reveals that the thin and thick filaments are mutually dependent on each other for striation. Further, the tension sensor complex comprised of zipper/Zasp/α-actinin is involved in stabilizing the sarcomere but not in its initial formation. Finally, integrins appear essential for the interdigitation of thin and thick filaments that occurs prior to striation. Thus, sarcomere formation occurs by the coordinated assembly of multiple latent protein complexes, as opposed to sequential assembly