Spastin mutations impair coordination between lipid droplet dispersion and reticulum.

Lipid droplets (LD) are affected in multiple human disorders. These highly dynamic organelles are involved in many cellular roles. While their intracellular dispersion is crucial to ensure their function and other organelles-contact, underlying mechanisms are still unclear. Here we show that Spastin, one of the major proteins involved in Hereditary Spastic Paraplegia (HSP), controls LD dispersion. Spastin depletion in zebrafish affects metabolic properties and organelle dynamics. These functions are ensured by a conserved complex set of splice variants. M1 isoforms determine LD dispersion in the cell by orchestrating endoplasmic reticulum (ER) shape along microtubules (MTs). To further impact LD fate, Spastin modulates transcripts levels and subcellular location of other HSP key players, notably Seipin and REEP1. In pathological conditions, mutations in human Spastin M1 disrupt this mechanism and impacts LD network. Spastin depletion influences not only other key proteins but also modulates specific neutral lipids and phospholipids, revealing an impact on membrane and organelle components. Altogether our results show that Spastin and its partners converge in a common machinery that coordinates LD dispersion and ER shape along MTs. Any alteration of this system results in HSP clinical features and impacts lipids profile, thus opening new avenues for novel biomarkers of HSP

Molecular codes and in vitro generation of hypocretin and melanin concentrating hormone neurons.

Hypocretin/orexin (HCRT) and melanin concentrating hormone (MCH) neuropeptides are exclusively produced by the lateral hypothalamus and play important roles in sleep, metabolism, reward, and motivation. Loss of HCRT (ligands or receptors) causes the sleep disorder narcolepsy with cataplexy in humans and in animal models. How these neuropeptides are produced and involved in diverse functions remain unknown. Here, we developed methods to sort and purify HCRT and MCH neurons from the mouse late embryonic hypothalamus. RNA sequencing revealed key factors of fate determination for HCRT (Peg3, Ahr1, Six6, Nr2f2, and Prrx1) and MCH (Lmx1, Gbx2, and Peg3) neurons. Loss of Peg3 in mice significantly reduces HCRT and MCH cell numbers, while knock-down of a Peg3 ortholog in zebrafish completely abolishes their expression, resulting in a 2-fold increase in sleep amount. We also found that loss of HCRT neurons in Hcrt-ataxin-3 mice results in a specific 50% decrease in another orexigenic neuropeptide, QRFP, that might explain the metabolic syndrome in narcolepsy. The transcriptome results were used to develop protocols for the production of HCRT and MCH neurons from induced pluripotent stem cells and ascorbic acid was found necessary for HCRT and BMP7 for MCH cell differentiation. Our results provide a platform to understand the development and expression of HCRT and MCH and their multiple functions in health and disease

Distinct patterns of skeletal muscle mitochondria fusion, fission and mitophagy upon duration of exercise training.

Healthy ageing interventions encompass regular exercise to prevent mitochondrial dysfunction, key player in sarcopenia pathogenesis. Mitochondrial biogenesis has been well documented, but mitochondrial remodelling in response to exercise training is poorly understood. Here we investigated fusion, fission and mitophagy before and after an exercise intervention in older adults. Skeletal muscle biopsies were collected from 22 healthy sedentary men and women before and after 4 months of supervised training. Eight lifelong trained age- and gender-matched volunteers served as positive controls. Transmission electron microscopy was used to estimate mitochondrial content. Western blotting and qRT-PCR were used to detect changes in specific proteins and transcripts. After intervention, mitochondrial content increased to levels of controls. While enhancement of fusion was prevalent after intervention, inhibition of fission and increased mitophagy were dominant in controls. Similarly to PARKIN, BCL2L13 content was higher in controls. The observed molecular adaptations paralleled long-term effects of training on physical fitness, exercise efficiency and oxidative capacity. This study describes distinct patterns of molecular adaptations in human skeletal muscle under chronic exercise training. After 16 weeks of exercise, the pattern was dominated by fusion to increase mitochondrial content to the metabolic demands of exercise. In lifelong exercise, the pattern was dominated by mitophagy synchronized with increased fusion and decreased fission, indicating an increased mitochondrial turnover. In addition to these temporally distinct adaptive mechanisms, this study suggests for the first time a specific role of BCL2L13 in chronic exercise that requires constant maintenance of mitochondrial quality

Fluorescent amino acids as versatile building blocks for chemical biology

Fluorophores have transformed the way we study biological systems, enabling non-invasive studies in cells and intact organisms, which increase our understanding of complex processes at the molecular level. Fluorescent amino acids have become an essential chemical tool because they can be used to construct fluorescent macromolecules, such as peptides and proteins, without disrupting their native biomolecular properties. Fluorescent and fluorogenic amino acids with unique photophysical properties have been designed for tracking protein–protein interactions in situ or imaging nanoscopic events in real time with high spatial resolution. In this Review, we discuss advances in the design and synthesis of fluorescent amino acids and how they have contributed to the field of chemical biology in the past 10 years. Important areas of research that we review include novel methodologies to synthesize building blocks with tunable spectral properties, their integration into peptide and protein scaffolds using site-specific genetic encoding and bioorthogonal approaches, and their application to design novel artificial proteins, as well as to investigate biological processes in cells by means of optical imaging. [Figure not available: see fulltext.]

Vesicular Axonal Transport is Modified In Vivo by Tau Deletion or Overexpression in Drosophila.

Structural microtubule associated protein Tau is found in high amount in axons and is involved in several neurodegenerative diseases. Although many studies have highlighted the toxicity of an excess of Tau in neurons, the in vivo understanding of the endogenous role of Tau in axon morphology and physiology is poor. Indeed, knock-out mice display no strong cytoskeleton or axonal transport phenotype, probably because of some important functional redundancy with other microtubule-associated proteins (MAPs). Here, we took advantage of the model organism <i>Drosophila</i> , which genome contains only one homologue of the Tau/MAP2/MAP4 family to decipher (endogenous) Tau functions. We found that Tau depletion leads to a decrease in microtubule number and microtubule density within axons, while Tau excess leads to the opposite phenotypes. Analysis of vesicular transport in <i>tau</i> mutants showed altered mobility of vesicles, but no change in the total amount of putatively mobile vesicles, whereas both aspects were affected when Tau was overexpressed. In conclusion, we show that loss of Tau in <i>tau</i> mutants not only leads to a decrease in axonal microtubule density, but also impairs axonal vesicular transport, albeit to a lesser extent compared to the effects of an excess of Tau

Processen att skapa en marknadsplan : Ett framhållande av teoretiska och praktiska roller

Recent advances in the understanding of basic pathological mechanisms in various neurological diseases depend directly on the development of novel bioanalytical technologies that allow sensitive and specific chemical imaging at high resolution in cells and tissues. Mass spectrometry-based molecular imaging (IMS) has gained increasing popularity in biomedical research for mapping the spatial distribution of molecular species in\ua0situ. The technology allows for comprehensive, untargeted delineation of in\ua0situ distribution profiles of metabolites, lipids, peptides and proteins. A major advantage of IMS over conventional histochemical techniques is its superior molecular specificity. Imaging mass spectrometry has therefore great potential for probing molecular regulations in CNS-derived tissues and cells for understanding neurodegenerative disease mechanism. The goal of this review is to familiarize the reader with the experimental workflow, instrumental developments and methodological challenges as well as to give a concise overview of the major advances and recent developments and applications of IMS-based protein and peptide profiling with particular focus on neurodegenerative diseases. This article is part of the Special Issue “Proteomics”. (Figure presented.)

Tanycytes Regulate Lipid Homeostasis by Sensing Free Fatty Acids and Signaling to Key Hypothalamic Neuronal Populations via FGF21 Secretion.

The hypothalamus plays a key role in the detection of energy substrates to regulate energy homeostasis. Tanycytes, the hypothalamic ependymo-glia, are located at a privileged position to integrate multiple peripheral inputs. We observed that tanycytes produce and secrete Fgf21 and are located close to Fgf21-sensitive neurons. Fasting, likely via the increase in circulating fatty acids, regulates this central Fgf21 production. Tanycytes store palmitate in lipid droplets and oxidize it, leading to the activation of a reactive oxygen species (ROS)/p38-MAPK signaling pathway, which is essential for tanycytic Fgf21 expression upon palmitate exposure. Tanycytic Fgf21 deletion triggers an increase in lipolysis, likely due to impaired inhibition of key neurons during fasting. Mice deleted for tanycytic Fgf21 exhibit increased energy expenditure and a reduction in fat mass gain, reminiscent of a browning phenotype. Our results suggest that tanycytes sense free fatty acids to maintain body lipid homeostasis through Fgf21 signaling within the hypothalamus