Transcription factor Nrf2 is essential for the antioxidant responsive element (ARE)-mediated induction of phase II detoxifying and oxidative stress enzyme genes. Detailed analysis of differential Nrf2 activity displayed in transfected cell lines ultimately led to the identification of a new protein, which we named Keap1, that suppresses Nrf2 transcriptional activity by specific binding to its evolutionarily conserved amino-terminal regulatory domain. The closest homolog of Keap1 is a Drosophila actin-binding protein called Kelch, implying that Keap1 might be a Nrf2 cytoplasmic effector. We then showed that electrophilic agents antagonize Keap1 inhibition of Nrf2 activity in vivo, allowing Nrf2 to traverse from the cytoplasm to the nucleus and potentiate the ARE response. We postulate that Keap1 and Nrf2 constitute a crucial cellular sensor for oxidative stress, and together mediate a key step in the signaling pathway that leads to transcriptional activation by this novel Nrf2 nuclear shuttling mechanism. The activation of Nrf2 leads in turn to the induction of phase II enzyme and antioxidative stress genes in response to electrophiles and reactive oxygen species.

/pdf/keap1-represses-nuclear-activation-of-antioxidant-responsive-4szbzl5y9i.pdf

Keap1 represses nuclear activation of antioxidant responsive elements by Nrf2 through binding to the amino-terminal Neh2 domain

The primary cilium has recently stepped into the spotlight, as a flood of data show that this organelle has crucial roles in vertebrate development and human genetic diseases. Cilia are required for the response to developmental signals, and evidence is accumulating that the primary cilium is specialized for hedgehog signal transduction. The formation of cilia, in turn, is regulated by other signalling pathways, possibly including the planar cell polarity pathway. The cilium therefore represents a nexus for signalling pathways during development. The connections between cilia and developmental signalling have begun to clarify the basis of human diseases associated with ciliary dysfunction.

/pdf/the-primary-cilium-a-signalling-centre-during-vertebrate-58mv3re75c.pdf

The primary cilium: a signalling centre during vertebrate development

Intraflagellar transport (IFT) proteins were first identified as essential factors for the growth and maintenance of flagella in the single-celled alga Chlamydomonas reinhardtii. In a screen for embryonic patterning mutations induced by ethylnitrosourea, here we identify two mouse mutants, wimple (wim) and flexo (fxo), that lack ventral neural cell types and show other phenotypes characteristic of defects in Sonic hedgehog signalling. Both mutations disrupt IFT proteins: the wim mutation is an allele of the previously uncharacterized mouse homologue of IFT172; and fxo is a new hypomorphic allele of polaris, the mouse homologue of IFT88. Genetic analysis shows that Wim, Polaris and the IFT motor protein Kif3a are required for Hedgehog signalling at a step downstream of Patched1 (the Hedgehog receptor) and upstream of direct targets of Hedgehog signalling. Our data show that IFT machinery has an essential and vertebrate-specific role in Hedgehog signal transduction.

Hedgehog signalling in the mouse requires intraflagellar transport proteins

The present invention is directed to secreted and transmembrane polypeptides and to nucleic acid molecules encoding those polypeptides. Also provided herein are vectors and host cells comprising those nucleic acid sequences, chimeric polypeptide molecules comprising the polypeptides of the present invention fused to heterologous polypeptide sequences, antibodies which bind to the polypeptides of the present invention and to methods for producing the polypeptides of the present invention.

Secreted and Transmembrane Polypeptides and Nucleic Acids Encoding the Same

An elaborate network of highly inducible proteins protects aerobic cells against the cumulative damaging effects of reactive oxygen intermediates and toxic electrophiles, which are the major causes of neoplastic and chronic degenerative diseases. These cytoprotective proteins share common transcriptional regulation, through the Keap1–Nrf2 pathway, which can be activated by various exogenous and endogenous small molecules (inducers). Inducers chemically react with critical cysteine residues of the sensor protein Keap1, leading to stabilisation and nuclear translocation of transcription factor Nrf2, and ultimately to coordinate enhanced expression of genes coding for cytoprotective proteins. In addition, inducers inhibit pro-inflammatory responses, and there is a linear correlation spanning more than six orders of magnitude of concentrations between inducer and anti-inflammatory activity. Genetic deletion of transcription factor Nrf2 renders cells and animals much more sensitive to the damaging effects of electrophiles, oxidants and inflammatory agents in comparison with their wild-type counterparts. Conversely, activation of the Keap1–Nrf2 pathway allows survival and adaptation under various conditions of stress and has protective effects in many animal models. Cross-talks with other signalling pathways broadens the role of the Keap1–Nrf2 pathway in determining the fate of the cell, impacting fundamental biological processes such as proliferation, apoptosis, angiogenesis and metastasis.

The cytoprotective role of the Keap1–Nrf2 pathway

Costal2, a Novel Kinesin-Related Protein in the Hedgehog Signaling Pathway

Sonic hedgehog in the nervous system: functions, modifications and mechanisms

Caffeine is one of the most widely consumed stimulants in the world and has been proposed to promote wakefulness by antagonizing function of the adenosine A2A receptor. Here, we show that chronic administration of caffeine reduces and fragments sleep in Drosophila and also lengthens circadian period. To identify the mechanisms underlying these effects of caffeine, we first generated mutants of the only known adenosine receptor in flies (dAdoR), which by sequence is most similar to the mammalian A2A receptor. Mutants lacking dAdoR have normal amounts of baseline sleep, as well as normal homeostatic responses to sleep deprivation. Surprisingly, these mutants respond normally to caffeine. On the other hand, the effects of caffeine on sleep and circadian rhythms are mimicked by a potent phosphodiesterase inhibitor, IBMX (3-isobutyl-1-methylxanthine). Using in vivo fluorescence resonance energy transfer imaging, we find that caffeine induces widespread increase in cAMP levels throughout the brain. Finally, the effects of caffeine on sleep are blocked in flies that have reduced neuronal PKA activity. We suggest that chronic administration of caffeine promotes wakefulness in Drosophila, at least in part, by inhibiting cAMP phosphodiesterase activity.

https://www.jneurosci.org/content/jneuro/29/35/11029.full.pdf

The Effects of Caffeine on Sleep in Drosophila Require PKA Activity, But Not the Adenosine Receptor

In 2000, Drosophila melanogaster joined the ranks of vertebrates and invertebrates with a defined behavioral sleep state. The characterization of this sleep state revealed striking similarities to sleep in humans: sleep in flies has both circadian and homeostatic components, it is influenced by sex and age, and it is affected by pharmacological agents such as caffeine and antihistamines. As in mammals, arousal thresholds in flies increase with sleep deprivation. Furthermore, changes in brain electrical activity accompany the change from wake to sleep states. Not only do flies and vertebrates share these behavioral and physiological traits of sleep, but they are likely to share at least some genetic mechanisms underlying the regulation of sleep as well. This article reviews the methods currently used to identify and characterize the Drosophila sleep state. As these methods become more refined and our understanding of Drosophila sleep more detailed, the powerful techniques afforded by this organism are likely to unveil deep insights into the function(s) and regulatory mechanisms of sleep.

Drosophila melanogaster: an insect model for fundamental studies of sleep.

Copy-number variations (CNVs) are strong risk factors for neurodevelopmental and psychiatric disorders. The 15q13.3 microdeletion syndrome region contains up to ten genes and is associated with numerous conditions, including autism spectrum disorder (ASD), epilepsy, schizophrenia, and intellectual disability; however, the mechanisms underlying the pathogenesis of 15q13.3 microdeletion syndrome remain unknown. We combined whole-genome sequencing, human brain gene expression (proteome and transcriptome), and a mouse model with a syntenic heterozygous deletion (Df(h15q13)/+ mice) and determined that the microdeletion results in abnormal development of cortical dendritic spines and dendrite outgrowth. Analysis of large-scale genomic, transcriptomic, and proteomic data identified OTUD7A as a critical gene for brain function. OTUD7A was found to localize to dendritic and spine compartments in cortical neurons, and its reduced levels in Df(h15q13)/+ cortical neurons contributed to the dendritic spine and dendrite outgrowth deficits. Our results reveal OTUD7A as a major regulatory gene for 15q13.3 microdeletion syndrome phenotypes that contribute to the disease mechanism through abnormal cortical neuron morphological development.

/pdf/otud7a-regulates-neurodevelopmental-phenotypes-in-the-15q13-cn9sphx3fl.pdf

Karen S. Ho

Papers

Costal2, a Novel Kinesin-Related Protein in the Hedgehog Signaling Pathway

Sonic hedgehog in the nervous system: functions, modifications and mechanisms

The Effects of Caffeine on Sleep in Drosophila Require PKA Activity, But Not the Adenosine Receptor

Drosophila melanogaster: an insect model for fundamental studies of sleep.

OTUD7A Regulates Neurodevelopmental Phenotypes in the 15q13.3 Microdeletion Syndrome