Glycosphingolipids in signaling and development: from liposomes to model organisms

Since the launch of the raft hypothesis in 1997, data generated in liposomes and cultured cells have highlighted the role of glycosphingolipids (GSLs) in the dynamic organization of biological membranes and the activity of signaling complexes. In parallel studies, genetic analysis of the GSL synthetic pathway has begun to reveal some of the specific roles of GSLs in vivo. Here, we review the role of GSLs in signaling in the context of a refined raft hypothesis. Recent genetic studies in worms, flies, and mice give us the opportunity to integrate these in vivo data with earlier in vitro liposome studies.     

A taste illusion: taste sensation localized by touch.

Taste sensations appear to come from all over the inner surface of the mouth, yet the taste receptors are restricted to relatively small particular areas of the oral surface. In addition, even if a relatively large (e.g., one half) proportion of the taste field is damaged, subjective taste experience may be unaffected. The touch system contributes to this constancy because taste sensations appear to be localized by touch. If a taste solution is painted from the side of the tongue (an area of low receptor density) past the tip (an area of high receptor density) and on to the second side, the taste sensation begins weak, gets stronger at the tip, and retains much of its intensity. The strong taste from the tip follows the tactile path of the stimulus sweep. This illusion occurs for all four stimuli tested: sucrose, sodium chloride, citric acid, and quinine hydrochloride.     

A symptom network model of misophonia: From heightened sensory sensitivity to clinical comorbidity

Objectives

Misophonia—an unusually strong intolerance of certain sounds—can cause significant distress and disruption to those who have it but is an enigma in terms of our scientific understanding. A key challenge for explaining misophonia is that, as with other disorders, it is likely to emerge from an interaction of traits that also occur in the general population (e.g., sensory sensitivity and anxiety) and that are transdiagnostic in nature (i.e., shared with other disorders).

Methods

In this preregistered study with a large sample of participants (N = 1430), we performed a cluster analysis (based on responses to questions relating to misophonia) and identified two misophonia subgroups differing in severity, as well as a third group without misophonia. A subset of this sample (N = 419) then completed a battery of measures designed to assess sensory sensitivity and clinical comorbidities.

Results

Clinical symptoms were limited to the most severe group of misophonics (including autistic traits, migraine with visual aura, anxiety sensitivity, obsessive-compulsive traits). Both the moderate and severe groups showed elevated attention-to-detail and hypersensitivity (across multiple senses). A novel symptom network model of the data shows the presence of a central hub linking misophonia to sensory sensitivity which, in turn, connects to other symptoms in the network (relating to autism, anxiety, etc.).

Conclusion

The core features of misophonia are sensory-attentional in nature with severity linked strongly to comorbidities.     

ATR signaling in mammalian meiosis: From upstream scaffolds to downstream signaling

In germ cells undergoing meiosis, the induction of double strand breaks (DSBs) is required for the generation of haploid gametes. Defects in the formation, detection, or recombinational repair of DSBs often result in defective chromosome segregation and aneuploidies. Central to the ability of meiotic cells to properly respond to DSBs are DNA damage response (DDR) pathways mediated by DNA damage sensor kinases. DDR signaling coordinates an extensive network of DDR effectors to induce cell cycle arrest and DNA repair, or trigger apoptosis if the damage is extensive. Despite their importance, the functions of DDR kinases and effector proteins during meiosis remain poorly understood and can often be distinct from their known mitotic roles. A key DDR kinase during meiosis is ataxia telangiectasia and Rad3-related (ATR). ATR mediates key signaling events that control DSB repair, cell cycle progression, and meiotic silencing. These meiotic functions of ATR depend on upstream scaffolds and regulators, including the 9-1-1 complex and TOPBP1, and converge on many downstream effectors such as the checkpoint kinase CHK1. Here, we review the meiotic functions of the 9-1-1/TOPBP1/ATR/CHK1 signaling pathway during mammalian meiosis.     

The contribution of light touch sensory cues to corrective reactions during treadmill locomotion

The arms play an important role in balance regulation during walking. In general, perturbations delivered during walking trigger whole-body corrective responses. For instance, holding to stable handles can largely attenuate and even suppress responses in the leg muscles to perturbations during walking. Particular attention has been given to the influence of light touch on postural control. During standing, lightly touching a stable contact greatly reduces body sway and enhances corrective responses to postural perturbations, whereas light touch during walking allows subjects to continue to walk on a treadmill with the eyes closed. We hypothesized that in the absence of mechanical support from the arms, sensory cues from the hands would modulate responses in the legs to balance disturbing perturbations delivered at the torso during walking. To test this, subjects walked on a treadmill while periodically being pulled backwards at the waist while walking. The amplitude of the responses evoked in tibialis anterior to these perturbations was compared across 4 test conditions, in a 2 × 2 design. Subjects either (a) lightly touched or (b) did not touch a stable contact, while the eyes were (c) open or (d) closed. Allowing the subjects to touch a stable contact resulted in a reduction in the amount of fore-aft oscillation of the body on the treadmill, which was accompanied by a reduction in the ongoing electromyographic activity in both tibialis anterior and soleus during undisturbed walking. In contrast, the provision of touch resulted in an increase in the amplitude of the evoked responses in tibialis anterior to the backward perturbations that was more evident when subjects walked with the eyes closed. These results indicate that light touch provides a sensory cue that can be used to assist in stabilizing the body while walking. In addition, the sensory information provided by light touch contributes to the regulation of corrective reactions initiated by balance disturbances encountered during walking.     

Indoleamine 2,3-dioxygenase: From catalyst to signaling function

Control of tryptophan metabolism by indoleamine 2,3-dioxygenase (IDO) in dendritic cells (DCs) is a highly versatile regulator of innate and adaptive immune responses. In acute reactions, the otherwise inflammatory cytokine interferon γ (IFN-γ) acts in a feedback fashion to induce IDO's enzymatic function - and thus prevent potentially harmful, exaggerated responses - through the combined effects of tryptophan starvation and tryptophan catabolites acting via the aryl hydrocarbon receptor of T cells. IDO, however, is also involved in the maintenance of stable tolerance to self in noninflammatory contexts, thus restraining autoimmunity. Exposure, indeed, of mouse plasmacytoid DCs (pDCs) to transforming growth factor β (TGF-β) provides IDO with regulatory effects that are distinct, in nature, from its enzymic activity. Once phosphorylated, IDO mediates signaling events culminating in self-amplification and maintenance of a stably regulatory condition in pDCs. Therefore, IDO has dual immunoregulatory functions driven by distinct cytokines. Firstly, the IFN-γ-IDO axis is crucial in generating and sustaining the function of regulatory T cells. Secondly, a nonenzymic function of IDO - as a signaling molecule - contributes to TGF-β-driven tolerance. The latter function is part of a regulatory circuit in pDCs whereby - in response to TGF-β - the kinase Fyn mediates tyrosine phosphorylation of IDO-associated immunoreceptor tyrosine-based inhibitory motifs, resulting in downstream effects that regulate gene expression and preside over a proper, homeostatic balance between immunity and tolerance. All these aspects are covered in this review.     

Conditioned taste aversions: generalization to taste mixtures

Rats were trained to take their daily water ration within a 30-min session, during which the number of licks per 10-sec presentation of a drinking tube could be recorded. During one of these sessions, one of three stimuli (sucrose, NaCl or HCl) was presented, followed by the administration of cyclophosphamide to produce a conditioned taste aversion. When tested with mixtures of the conditioned stimulus (CS) with the other two stimuli and also with quinine hydrochloride, the animals avoided mixtures containing the CS in proportion to its concentration in the mixture. Although the natural preferences and aversions for these stimuli interacted somewhat with the learned taste aversions, rats responded to the presence of a CS in a mixture and did not generalize to other stimuli not containing the CS. Thus, the generalization of conditioned taste aversions provides a good measure of the behavioral similarities among gustatory stimuli.     

Disturbances in affective touch in hereditary sensory & autonomic neuropathy type III

Hereditary sensory and autonomic neuropathy type III (HSAN III, Riley–Day syndrome, Familial Dysautomia) is characterised by elevated thermal thresholds and an indifference to pain. Using microelectrode recordings we recently showed that these patients possess no functional stretch-sensitive mechanoreceptors in their muscles (muscle spindles), a feature that may explain their lack of stretch reflexes and ataxic gait, yet patients have apparently normal low-threshold cutaneous mechanoreceptors. The density of C-fibres in the skin is markedly reduced in patients with HSAN III, but it is not known whether the C-tactile afferents, a distinct type of low-threshold C fibre present in hairy skin that is sensitive to gentle stroking and has been implicated in the coding of pleasant touch are specifically affected in HSAN III patients. We addressed the relationship between C-tactile afferent function and pleasant touch perception in 15 patients with HSAN III and 15 age-matched control subjects. A soft make-up brush was used to apply stroking stimuli to the forearm and lateral aspect of the leg at five velocities: 0.3, 1, 3, 10 and 30 cm/s. As demonstrated previously, the control subjects rated the slowest and highest velocities as less pleasant than those applied at 1–10 cm/s, which fits with the optimal velocities for exciting C-tactile afferents. Conversely, for the patients, ratings of pleasantness did not fit the profile for C-tactile afferents. Patients either rated the higher velocities as more pleasant than the slow velocities, with the slowest velocities being rated unpleasant, or rated all velocities equally pleasant. We interpret this to reflect absent or reduced C-tactile afferent density in the skin of patients with HSAN III, who are likely using tactile cues (i.e. myelinated afferents) to rate pleasantness of stroking or are attributing pleasantness to this type of stimulus irrespective of velocity.     

Espin cytoskeletal proteins in the sensory cells of rodent taste buds

Espins are multifunctional actin-bundling proteins that are highly enriched in the microvilli of certain chemosensory and mechanosensory cells, where they are believed to regulate the integrity and/or dimensions of the parallel-actin-bundle cytoskeletal scaffold. We have determined that, in rats and mice, affinity purified espin antibody intensely labels the lingual and palatal taste buds of the oral cavity and taste buds in the pharyngo-laryngeal region. Intense immunolabeling was observed in the apical, microvillar region of taste buds, while the level of cytoplasmic labeling in taste bud cells was considerably lower. Taste buds contain tightly packed collections of sensory cells (light, or type II plus type III) and supporting cells (dark, or type I), which can be distinguished by microscopic features and cell type-specific markers. On the basis of results obtained using an antigen-retrieval method in conjunction with double immunofluorescence for espin and sensory taste cell-specific markers, we propose that espins are expressed predominantly in the sensory cells of taste buds. In confocal images of rat circumvallate taste buds, we counted 21.5 ± 0.3 espin-positive cells/taste bud, in agreement with a previous report showing 20.7 ± 1.3 light cells/taste bud when counted at the ultrastructural level. The espin antibody labeled spindle-shaped cells with round nuclei and showed 100% colocalization with cell-specific markers recognizing all type II [inositol 1,4,5-trisphosphate receptor type III (IP₃R₃)_, α-gustducin, protein-specific gene product 9.5 (PGP9.5)] and a subpopulation of type III (IP₃R₃, PGP9.5) taste cells. On average, 72%, 50%, and 32% of the espin-positive taste cells were labeled with antibodies to IP₃R₃, α-gustducin, and PGP9.5, respectively. Upon sectional analysis, the taste buds of rat circumvallate papillae commonly revealed a multi-tiered, espin-positive apical cytoskeletal apparatus. One espin-positive zone, a collection of ∼3 μm-long microvilli occupying the taste pore, was separated by an espin-depleted zone from a second espin-positive zone situated lower within the taste pit. This latter zone included espin-positive rod-like structures that occasionally extended basally to a depth of 10–12 μm into the cytoplasm of taste cells. We propose that the espin-positive zone in the taste pit coincides with actin bundles in association with the microvilli of type II taste cells, whereas the espin-positive microvilli in the taste pore are the single microvilli of type III taste cells.     

DiGeorge syndrome: the use of model organisms to dissect complex genetics

The research interest in DiGeorge syndrome (DGS) is partly due to its clinical importance. However, fundamental questions of genetics and developmental biology related to DGS are inspiring investigators to experiment with model systems. Most DGS cases are caused by a heterozygous chromosomal deletion del22q11, and the search for haploinsufficient genes has been successful in mice and led to the discovery of Tbx1 as a major player in the development of the pharyngeal arches and pouches. Whether TBX1 is haploinsufficient in humans, as several other T-box genes are, is yet to be proven. The puzzling clinical variability in patients with del22q11 is also being addressed in model organisms. Consistent with clinical data, experiments in mice indicate that genetics can only explain part of the phenotypic variability. The recent identification of phenotypic modifiers further underscores the complex genetics of this syndrome.     

Parkinson's disease: Insights from non-traditional model organisms

Parkinson's disease (PD) was one of the first neurological disorders to have aspects of the disease modeled faithfully in non-human animal species. A key feature of the disease is a diminished control over voluntary movement and progressive depletion of brain dopamine (DA) levels that stems from the large-scale loss of DA-producing neurons. Despite their inherent limitations, rodent and non-human primate models of PD have helped unravel several aspects of PD pathogenesis. Thus, we now have neurotransmitter replacement therapy for PD, and a number of neuroprotective compounds that can be assessed in clinical trials. However, no treatment is currently available that can halt or retard the progressive loss of DA neurons, which underlies PD pathology. Moreover, no therapies can permanently alleviate the clinical features of the disease. The lack of a cure or long-term effective treatment is paralled by our incomplete understanding of the underlying pathomechanisms of the disease. A range of robust, flexible, and complementary animal models will be an invaluable tool with which to unravel the pathogenesis of PD. Here we review the most important contributions made by non-mammalian model organisms. These include zebrafish (Danio rerio), flies (Drosophila melanogaster), anurans (frogs and toads) and nematodes (Caenorhabditis elegans). While it is not anticipated that they will replace rodent and primate-based ones, they offer convenient systems with which to explore the relative contribution made by genetic and environmental factors to PD pathology. In addition, they offer an economic and rapid alternative for testing compounds that target PD. Most importantly, the combined use of these models allow for ongoing research to uncover the basic mechanisms underlying PD pathogenesis.     

A forebrain undivided: Unleashing model organisms to solve the mysteries of holoprosencephaly

Evolutionary conservation and experimental tractability have made animal model systems invaluable tools in our quest to understand human embryogenesis, both normal and abnormal. Standard genetic approaches, particularly useful in understanding monogenic diseases, are no longer sufficient as research attention shifts toward multifactorial outcomes. Here, we examine this progression through the lens of holoprosencephaly (HPE), a common human malformation involving incomplete forebrain division, and a classic example of an etiologically complex outcome. We relate the basic underpinning of HPE pathogenesis to critical cell-cell interactions and signaling molecules discovered through embryological and genetic approaches in multiple model organisms, and discuss the role of the mouse model in functional examination of HPE-linked genes. We then outline the most critical remaining gaps to understanding human HPE, including the conundrum of incomplete penetrance/expressivity and the role of gene-environment interactions. To tackle these challenges, we outline a strategy that leverages new and emerging technologies in multiple model systems to solve the puzzle of HPE.     

Developmental patterning genes and their conserved functions: from model organisms to humans

Molecular and genetic evidence accumulated during the past 20 years in the field of developmental biology indicates that different animals possess many common genetic systems for embryonic patterning. In this review we describe the conserved functions of such developmental patterning genes and their relevance for human pathological conditions. Special attention is given to the Hox genetic system, involved in establishing cell identities along the anterior-posterior axis of all higher metazoans. We also describe other conserved genetic systems, such as the involvement of Pax6 genes in eye development and the role of Nkx2.5-type proteins in heart development. Finally, we outline some fascinating problems at the forefront of the studies of developmental patterning genes and show how knowledge obtained from model genetic organisms such as Drosophila helps to explain normal human morphogenesis and the genetic basis of some birth defects.     

Autophagy and ageing: Insights from invertebrate model organisms

Ageing in diverse species ranging from yeast to humans is associated with the gradual, lifelong accumulation of molecular and cellular damage. Autophagy, a conserved lysosomal, self-destructive process involved in protein and organelle degradation, plays an essential role in both cellular and whole-animal homeostasis. Accumulating evidence now indicates that autophagic degradation declines with age and this gradual reduction of autophagy might have a causative role in the functional deterioration of biological systems during ageing. Indeed, loss of autophagy gene function significantly influences longevity. Moreover, genetic or pharmacological manipulations that extend lifespan in model organisms often activate autophagy. Interestingly, conserved signalling pathways and environmental factors that regulate ageing, such as the insulin/IGF-1 signalling pathway and oxidative stress response pathways converge on autophagy. In this article, we survey recent findings in invertebrates that contribute to advance our understanding of the molecular links between autophagy and the regulation of ageing. In addition, we consider related mechanisms in other organisms and discuss their similarities and idiosyncratic features in a comparative manner.