Combinatorial Activation and Repression by Seven Transcription Factors Specify Drosophila Odorant Receptor Expression

A systematic analysis reveals a regulatory network controlling selective odorant receptor expression and neuronal diversity in Drosophila

Mechanisms of sensory neuron diversification during development and in the adult Drosophila : How to make a difference

The nervous system contains a vast number of neurons and displays a great diversity in cell types and classes. Even though this has been known for a long time, the exact mechanism of cell specification is still poorly understood. How does a cell know what type of neuron to which it should be specified? It is important to understand cellular specification, not only for our general understanding of biological processes, but also to allow us to develop treatments for patients with destructive diseases, such as Alzheimer’s, Parkinson, cancer or stroke. To address how neuronal specification and thereby diversification is evolved, we have chosen to study a complex but defined set of neurons, the Drosophila olfactory system. Olfactory sensory neurons (OSNs) detect an enormous variety of small volatile molecules with extremely high specificity and sensitivity. The adult Drosophila olfactory system contains 34 OSN classes each defined by their expression of a specific odorant receptor (OR). In both insects and vertebrates, each OSN expresses only one OR. In mouse there are approximately 1200 and in Drosophila 60 different OR genes. Despite the range of mechanisms known to determine cell identity and that the olfactory system is remarkably conserved across the phyla, it is still unclear how an OSN chooses to express a particular OR from a large genomic repertoire. In this thesis, the specification and diversification of the final steps establishing an OSN identity is addressed. We find seven transcription factors that are continuously required in different combinations for the expression of all ORs. The TFs can in different gene context both activate and repress OR expression, making the regulation more economical and indicating that repression is crucial for correct gene expression. We further identified a repressor complex that is able to segregate OR expression between OSN classes and propose a mechanism on how one single co-repressor can specify a large number of neuron classes.Exploring the OSN we found the developmental Hh signalling pathway is expressed in the postmitotic neuron. We show several fundamental similarities between the canonical Drosophila Hh pathway and the cilia mediated Hh transduction in component function. Further investigation revealed a function of cilia mediated Hh signalling in sensory neuron modulator. The results generated here will create a greater in vivo understanding of how postmitotic processes generate neurons with different fates and contribute to the maintaining of neuron function

Cilia-Mediated Hedgehog Signaling in Drosophila

Cilia mediate Hedgehog (Hh) signaling in vertebrates and Hh deregulation results in several clinical manifestations, such as obesity, cognitive disabilities, developmental malformations, and various cancers. Drosophila cells are nonciliated during development, which has led to the assumption that cilia-mediated Hh signaling is restricted to vertebrates. Here, we identify and characterize a cilia-mediated Hh pathway in Drosophila olfactory sensory neurons. We demonstrate that several fundamental key aspects of the vertebrate cilia pathway, such as ciliary localization of Smoothened and the requirement of the intraflagellar transport system, are present in Drosophila. We show that Cos2 and Fused are required for the ciliary transport of Smoothened and that cilia mediate the expression of the Hh pathway target genes. Taken together, our data demonstrate that Hh signaling in Drosophila can be mediated by two pathways and that the ciliary Hh pathway is conserved from Drosophila to vertebrates.At the time for thesis presentation publication was in status: Manuscript</p

Cilia-Mediated Hedgehog Signaling in Drosophila

Cilia mediate Hedgehog (Hh) signaling in vertebrates and Hh deregulation results in several clinical manifestations, such as obesity, cognitive disabilities, developmental malformations, and various cancers. Drosophila cells are nonciliated during development, which has led to the assumption that cilia-mediated Hh signaling is restricted to vertebrates. Here, we identify and characterize a cilia-mediated Hh pathway in Drosophila olfactory sensory neurons. We demonstrate that several fundamental key aspects of the vertebrate cilia pathway, such as ciliary localization of Smoothened and the requirement of the intraflagellar transport system, are present in Drosophila. We show that Cos2 and Fused are required for the ciliary transport of Smoothened and that cilia mediate the expression of the Hh pathway target genes. Taken together, our data demonstrate that Hh signaling in Drosophila can be mediated by two pathways and that the ciliary Hh pathway is conserved from Drosophila to vertebrates.At the time for thesis presentation publication was in status: Manuscript</p

Long-term treatment with antidepressants, but not environmental stimulation, induces expression of NP2 mRNA in hippocampus and medial habenula.

In experimental settings, antidepressant treatment as well as a stimulating environment has a positive influence on cognition and hippocampal plasticity. One putative mediator of this process is Neuronal Pentraxin 2 (NP2, Narp), known to mediate clustering of glutamatergic AMPA receptors at synapses, and demonstrated to play a role in activity-dependent synaptogenesis and synaptic plasticity. This study demonstrates that NP2 mRNA is robustly expressed in all hippocampal subregions and the medial habenula (MHb), both regions implicated in cognitive functions. Furthermore, NP2 mRNA expression is upregulated in the hippocampal subregions as well as in the MHb after long-term treatment with different antidepressant drugs regardless of monoaminergic profile, suggesting NP2 as a common mode of action of different antidepressant drugs. This effect occurs at the time frame where clinical response is normally achieved. In contrast, neither environmental enrichment nor deprivation has any influence on long-term NP2 mRNA expression. These findings support an involvement of NP2 in the pathway of antidepressant-induced plasticity, but not EE-induced plasticity; that NP2 might constitute a common link for the action of different types of antidepressant drugs and that the MHb could be a putative region for further studies of NP2

Hedgehog Signaling Regulates the Ciliary Transport of Odorant Receptors in Drosophila

Hedgehog (Hh) signaling is a key regulatory pathway during development and also has a functional role in mature neurons. Here, we show that Hh signaling regulates the odor response in adult Drosophila olfactory sensory neurons (OSNs). We demonstrate that this is achieved by regulating odorant receptor (OR) transport to and within the primary cilium in OSN neurons. Regulation relies on ciliary localization of the Hh signal transducer Smoothened (Smo). We further demonstrate that the Hh- and Smo-dependent regulation of the kinesin-like protein Cos2 acts in parallel to the intraflagellar transport system (IFT) to localize ORs within the cilium compartment. These findings expand our knowledge of Hh signaling to encompass chemosensory modulation and receptor trafficking.Funding Agencies|Swedish Foundation for Strategic Research [F06-0013]</p

The location of the binding site upstream of the OR dictates Xbp1 function.

<p>(A) Motif density plot, showing motifs found upstream of OR genes that did not require the matching TF (see <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1001280#pbio.1001280.s007" target="_blank">Table S3</a> for statistics). (B) Bar graph depicting the total number of motifs located upstream or downstream the TATA box for ORs that either require the TF (“essential”) or not (“nonessential”) for expression. (C) Double in situ labeling of <i>Or98a</i> and <i>Or56a</i> in wild type (Wt) and <i>xbp1-IR</i> antennae revealed ectopic <i>Or98a</i> expression next to <i>Or56a</i>. The RNAi phenotypes are summarized as a matrix (grey, wild-type expression; red, ectopic; and black, loss of expression). (D) One Xbp1 motif (purple) was found next to the TATA box (green) of <i>Or98a</i>. The <i>Or98a</i> promoter construct produced expression in a single domain (light blue oval, black expression). Whereas, the same <i>Or98a</i> promoter construct with a mutated Xbp1 motif (red) produced a distal expansion of the expression.</p

Hedgehog Signaling Regulates the Ciliary Transport of Odorant Receptors in Drosophila

Hedgehog (Hh) signaling is a key regulatory pathway during development and also has a functional role in mature neurons. Here, we show that Hh signaling regulates the odor response in adult Drosophila olfactory sensory neurons (OSNs). We demonstrate that this is achieved by regulating odorant receptor (OR) transport to and within the primary cilium in OSN neurons. Regulation relies on ciliary localization of the Hh signal transducer Smoothened (Smo). We further demonstrate that the Hh- and Smo-dependent regulation of the kinesin-like protein Cos2 acts in parallel to the intraflagellar transport system (IFT) to localize ORs within the cilium compartment. These findings expand our knowledge of Hh signaling to encompass chemosensory modulation and receptor trafficking.Funding Agencies|Swedish Foundation for Strategic Research [F06-0013]</p

Transcriptional activation and repression are required for correct expression of each OR gene to one OSN class.

<p>(A) Double in situ labeling of <i>Or23a</i> and <i>Or43b</i> in wild type (Wt) and <i>E93-IR</i> antennae, the <i>Or43ba</i> expression phenotypes are further depicted schematically and summarized as a matrix (grey, wild-type expression; red, ectopic; and black, loss of expression). (B) Double in situ hybridization labeling of <i>Or67a</i> and <i>Or67b</i> expression in wild type (Wt) and <i>acj6-IR</i> antenna. The resultant phenotypes are further summarized as a schematic and a matrix summary. Note the new pair of <i>Or43b</i> and <i>Or23a</i> when <i>E93</i> is knocked down (A), and OR coexpression generated in <i>acj6</i> knock-downs (B). (C) Model depicting how activation and repression of OR expression can specify an OSN class. Activation of OR gene expression (left box); different combinations of a limited set of TFs bind a proximal upstream region and produce OR expression in a broad antenna region. Repression of OR gene expression (right box), distal located repressors together with the dual function of the TFs determined by binding site location or possibly cofactor use, restrict OR expression. The combined sum of OR gene activation and repression produce expression to one single OSN class.</p