Essential requirement for JPT2 in NAADP-evoked Ca²⁺ signaling

Nicotinic acid adenine dinucleotide phosphate (NAADP) is a second messenger that releases Ca2+ from acidic organelles through the activation of two-pore channels (TPCs) to regulate endolysosomal trafficking events. NAADP action is mediated by NAADP-binding protein(s) of unknown identity that confer NAADP sensitivity to TPCs. Here, we used a “clickable” NAADP-based photoprobe to isolate human NAADP-binding proteins and identified Jupiter microtubule-associated homolog 2 (JPT2) as a TPC accessory protein required for endogenous NAADP-evoked Ca2+ signaling. JPT2 was also required for the translocation of a severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pseudovirus through the endolysosomal system. Thus, JPT2 is a component of the NAADP receptor complex that is essential for TPC-dependent Ca2+ signaling and control of coronaviral entry

Mitochondrial Uptake of Thiamin Pyrophosphate: Physiological and Cell Biological Aspects (vol 8, e73503, 2013)

[This corrects the article DOI: 10.1371/journal.pone.0073503.]

Supernovae from rotating stars

The present paper discusses the main physical effects produced by stellar rotation on presupernovae, as well as observations which confirm these effects and their consequences for presupernova models. Rotation critically influences the mass of the exploding cores, the mass and chemical composition of the envelopes and the types of supernovae, as well as the properties of the remnants and the chemical yields. In the formation of gamma-ray bursts, rotation and the properties of rotating stars appear as the key factor. In binaries, the interaction between axial rotation and tidal effects often leads to interesting and unexpected results. Rotation plays a key role in shaping the evolution and nucleosynthesis in massive stars with very low metallicities (metallicity below about the Small Magellanic Cloud metallicity down to Population III stars). At solar and higher metallicities, the effects of rotation compete with those of stellar winds. In close binaries, the synchronisation process can lock the star at a high rotation rate despite strong mass loss and thus both effects, rotation and stellar winds, have a strong impact. In conclusion, rotation is a key physical ingredient of the stellar models and of presupernova stages, and the evolution both of single stars and close binaries. Moreover, important effects are expected along the whole cosmic history.Comment: 36 pages, 15 figures, published in Handbook of Supernovae, A.W. Alsabti and P. Murdin (eds), Springe

Identifying Activated T Cells in Reconstituted RAG Deficient Mice Using Retrovirally Transduced Pax5 Deficient Pro-B Cells

Various methods have been used to identify activated T cells such as binding of MHC tetramers and expression of cell surface markers in addition to cytokine-based assays. In contrast to these published methods, we here describe a strategy to identify T cells that respond to any antigen and track the fate of these activated T cells. We constructed a retroviral double-reporter construct with enhanced green fluorescence protein (EGFP) and a far-red fluorescent protein from Heteractis crispa (HcRed). LTR-driven EGFP expression was used to enrich and identify transduced cells, while HcRed expression is driven by the CD40Ligand (CD40L) promoter, which is inducible and enables the identification and cell fate tracing of T cells that have responded to infection/inflammation. Pax5 deficient pro-B cells that can give rise to different hematopoietic cells like T cells, were retrovirally transduced with this double-reporter cassette and were used to reconstitute the T cell pool in RAG1 deifcient mice that lack T and B cells. By using flow cytometry and histology, we identified activated T cells that had developed from Pax5 deficient pro-B cells and responded to infection with the bacterial pathogen Listeria monocytogenes. Microscopic examination of organ sections allowed visual identification of HcRed-expressing cells. To further characterize the immune response to a given stimuli, this strategy can be easily adapted to identify other cells of the hematopoietic system that respond to infection/inflammation. This can be achieved by using an inducible reporter, choosing the appropriate promoter, and reconstituting mice lacking cells of interest by injecting gene-modified Pax5 deficient pro-B cells

Dysregulation of lysosomal morphology by pathogenic LRRK2 is corrected by two-pore channel 2 inhibition

Two-pore channels (TPCs) are endo-lysosomal ion channels implicated in Ca2+ signalling from acidic organelles. The relevance of these ubiquitous proteins for human disease however is unclear. Here we report that lysosomes are enlarged and aggregated in fibroblasts from Parkinson disease patients with the common G2019S mutation in LRRK2. Defects were corrected by molecular silencing of TPC2, pharmacological inhibition of TPC regulators (Rab7, NAADP, PI(3,5)P2) and buffering local Ca2+ increases. NAADP-evoked Ca2+ signals were exaggerated in diseased cells. TPC2 is thus a potential druggable target within a pathogenic LRRK2 cascade that disrupts Ca2+-dependent trafficking in Parkinson disease

Structural insight into SUMO chain recognition and manipulation by the ubiquitin ligase RNF4

The small ubiquitin-like modifier (SUMO) can form polymeric chains that are important signals in cellular processes such as meiosis, genome maintenance and stress response. The SUMO-targeted ubiquitin ligase RNF4 engages with SUMO chains on linked substrates and catalyses their ubiquitination, which targets substrates for proteasomal degradation. Here we use a segmental labelling approach combined with solution nuclear magnetic resonance (NMR) spectroscopy and biochemical characterization to reveal how RNF4 manipulates the conformation of the SUMO chain, thereby facilitating optimal delivery of the distal SUMO domain for ubiquitin transfer

Single Gene Deletions of Orexin, Leptin, Neuropeptide Y, and Ghrelin Do Not Appreciably Alter Food Anticipatory Activity in Mice

Timing activity to match resource availability is a widely conserved ability in nature. Scheduled feeding of a limited amount of food induces increased activity prior to feeding time in animals as diverse as fish and rodents. Typically, food anticipatory activity (FAA) involves temporally restricting unlimited food access (RF) to several hours in the middle of the light cycle, which is a time of day when rodents are not normally active. We compared this model to calorie restriction (CR), giving the mice 60% of their normal daily calorie intake at the same time each day. Measurement of body temperature and home cage behaviors suggests that the RF and CR models are very similar but CR has the advantage of a clearly defined food intake and more stable mean body temperature. Using the CR model, we then attempted to verify the published result that orexin deletion diminishes food anticipatory activity (FAA) but observed little to no diminution in the response to CR and, surprisingly, that orexin KO mice are refractory to body weight loss on a CR diet. Next we tested the orexigenic neuropeptide Y (NPY) and ghrelin and the anorexigenic hormone, leptin, using mouse mutants. NPY deletion did not alter the behavior or physiological response to CR. Leptin deletion impaired FAA in terms of some activity measures, such as walking and rearing, but did not substantially diminish hanging behavior preceding feeding time, suggesting that leptin knockout mice do anticipate daily meal time but do not manifest the full spectrum of activities that typify FAA. Ghrelin knockout mice do not have impaired FAA on a CR diet. Collectively, these results suggest that the individual hormones and neuropepetides tested do not regulate FAA by acting individually but this does not rule out the possibility of their concerted action in mediating FAA

The Mitochondrial Ca(2+) Uniporter: Structure, Function, and Pharmacology.

Mitochondrial Ca(2+) uptake is crucial for an array of cellular functions while an imbalance can elicit cell death. In this chapter, we briefly reviewed the various modes of mitochondrial Ca(2+) uptake and our current understanding of mitochondrial Ca(2+) homeostasis in regards to cell physiology and pathophysiology. Further, this chapter focuses on the molecular identities, intracellular regulators as well as the pharmacology of mitochondrial Ca(2+) uniporter complex