Golgi Cisternal Unstacking Stimulates COPI Vesicle Budding and Protein Transport

The Golgi apparatus in mammalian cells is composed of flattened cisternae that are densely packed to form stacks. We have used the Golgi stacking protein GRASP65 as a tool to modify the stacking state of Golgi cisternae. We established an assay to measure protein transport to the cell surface in post-mitotic cells in which the Golgi was unstacked. Cells with an unstacked Golgi showed a higher transport rate compared to cells with stacked Golgi membranes. Vesicle budding from unstacked cisternae in vitro was significantly increased compared to stacked membranes. These results suggest that Golgi cisternal stacking can directly regulate vesicle formation and thus the rate of protein transport through the Golgi. The results further suggest that at the onset of mitosis, unstacking of cisternae allows extensive and rapid vesiculation of the Golgi in preparation for its subsequent partitioning

Simulation of intracardiac electrograms around acute ablation lesions

Radiofrequency ablation (RFA) is a widely used clinical treatment for many types of cardiac arrhythmias. However, nontransmural lesions and gaps between linear lesions often lead to recurrence of the arrhythmia. Intracardiac electrograms (IEGMs) provide real-time information regarding the state of the cardiac tissue surrounding the catheter tip. Nevertheless, the formation and interpretation of IEGMs during the RFA procedure is complex and yet not fully understood. In this in-silico study, we propose a computational model for acute ablation lesions. Our model consists of a necrotic scar core and a border zone, describing irreversible and reversible temperature induced electrophysiological phenomena. These phenomena are modeled by varying the intra- and extracellular conductivity of the tissue as well as a regulating zone factor. The computational model is evaluated regarding its feasibility and validity. Therefore, this model was compared to an existing one and to clinical measurements of five patients undergoing RFA. The results show that the model can indeed be used to recreate IEGMs. We computed IEGMs arising from complex ablation scars, such as scars with gaps or two overlapping ellipsoid scars. For orthogonal catheter orientation, the presence of a second necrotic core in the near-field of a punctiform acute ablation lesion had minor impact on the resulting signal morphology. The presented model can serve as a base for further research on the formation and interpretation of IEGMs

ERK regulates Golgi and centrosome orientation towards the leading edge through GRASP65

Directed cell migration requires the orientation of the Golgi and centrosome toward the leading edge. We show that stimulation of interphase cells with the mitogens epidermal growth factor or lysophosphatidic acid activates the extracellular signal–regulated kinase (ERK), which phosphorylates the Golgi structural protein GRASP65 at serine 277. Expression of a GRASP65 Ser277 to alanine mutant or a GRASP65 1–201 truncation mutant, neither of which can be phosphorylated by ERK, prevents Golgi orientation to the leading edge in a wound assay. We show that phosphorylation of GRASP65 with recombinant ERK leads to the loss of GRASP65 oligomerization and causes Golgi cisternal unstacking. Furthermore, preventing Golgi polarization by expressing mutated GRASP65 inhibits centrosome orientation, which is rescued upon disassembly of the Golgi structure by brefeldin A. We conclude that Golgi remodeling, mediated by phosphorylation of GRASP65 by ERK, is critical for the establishment of cell polarity in migrating cells

Selenium Deficiency Is Associated with Mortality Risk from COVID-19

SARS-CoV-2 infections underlie the current coronavirus disease (COVID-19) pandemic and are causative for a high death toll particularly among elderly subjects and those with comorbidities. Selenium (Se) is an essential trace element of high importance for human health and particularly for a well-balanced immune response. The mortality risk from a severe disease like sepsis or polytrauma is inversely related to Se status. We hypothesized that this relation also applies to COVID-19. Serum samples (n = 166) from COVID-19 patients (n = 33) were collected consecutively and analyzed for total Se by X-ray fluorescence and selenoprotein P (SELENOP) by a validated ELISA. Both biomarkers showed the expected strong correlation (r = 0.7758, p < 0.001), pointing to an insufficient Se availability for optimal selenoprotein expression. In comparison with reference data from a European cross-sectional analysis (EPIC, n = 1915), the patients showed a pronounced deficit in total serum Se (mean ± SD, 50.8 ± 15.7 vs. 84.4 ± 23.4 µg/L) and SELENOP (3.0 ± 1.4 vs. 4.3 ± 1.0 mg/L) concentrations. A Se status below the 2.5th percentile of the reference population, i.e., [Se] < 45.7 µg/L and [SELENOP] < 2.56 mg/L, was present in 43.4% and 39.2% of COVID samples, respectively. The Se status was significantly higher in samples from surviving COVID patients as compared with non-survivors (Se; 53.3 ± 16.2 vs. 40.8 ± 8.1 µg/L, SELENOP; 3.3 ± 1.3 vs. 2.1 ± 0.9 mg/L), recovering with time in survivors while remaining low or even declining in non-survivors. We conclude that Se status analysis in COVID patients provides diagnostic information. However, causality remains unknown due to the observational nature of this study. Nevertheless, the findings strengthen the notion of a relevant role of Se for COVID convalescence and support the discussion on adjuvant Se supplementation in severely diseased and Se-deficient patients

The mitotic spindle mediates inheritance of the Golgi ribbon structure

The mammalian Golgi ribbon disassembles during mitosis and reforms in both daughter cells after division. Mitotic Golgi membranes concentrate around the spindle poles, suggesting that the spindle may control Golgi partitioning. To test this, cells were induced to divide asymmetrically with the entire spindle segregated into only one daughter cell. A ribbon reforms in the nucleated karyoplasts, whereas the Golgi stacks in the cytoplasts are scattered. However, the scattered Golgi stacks are polarized and transport cargo. Microinjection of Golgi extract together with tubulin or incorporation of spindle materials rescues Golgi ribbon formation. Therefore, the factors required for postmitotic Golgi ribbon assembly are transferred by the spindle, but the constituents of functional stacks are partitioned independently, suggesting that Golgi inheritance is regulated by two distinct mechanisms

Clinical Efficacy of Blue Light Full Body Irradiation as Treatment Option for Severe Atopic Dermatitis

BACKGROUND: Therapy of atopic dermatitis (AD) relies on immunosuppression and/or UV irradiation. Here, we assessed clinical efficacy and histopathological alterations induced by blue light-treatment of AD within an observational, non-interventional study. METHODOLOGY/PRINCIPAL FINDINGS: 36 patients with severe, chronic AD resisting long term disease control with local corticosteroids were included. Treatment consisted of one cycle of 5 consecutive blue light-irradiations (28.9 J/cm(2)). Patients were instructed to ask for treatment upon disease exacerbation despite interval therapy with topical corticosteroids. The majority of patients noted first improvements after 2-3 cycles. The EASI score was improved by 41% and 54% after 3 and 6 months, respectively (p≤0.005, and p≤0.002). Significant improvement of pruritus, sleep and life quality was noted especially after 6 months. Also, frequency and intensity of disease exacerbations and the usage of topical corticosteroids was reduced. Finally, immunohistochemistry of skin biopsies obtained at baseline and after 5 and 15 days revealed that, unlike UV light, blue light-treatment did not induce Langerhans cell or T cell depletion from skin. CONCLUSIONS/SIGNIFICANCE: Blue light-irradiation may represent a suitable treatment option for AD providing long term control of disease. Future studies with larger patient cohorts within a randomized, placebo-controlled clinical trial are required to confirm this observation

Mutations in GDF5 Reveal a Key Residue Mediating BMP Inhibition by NOGGIN

Signaling output of bone morphogenetic proteins (BMPs) is determined by two sets of opposing interactions, one with heterotetrameric complexes of cell surface receptors, the other with secreted antagonists that act as ligand traps. We identified two mutations (N445K,T) in patients with multiple synostosis syndrome (SYM1) in the BMP–related ligand GDF5. Functional studies of both mutants in chicken micromass culture demonstrated a gain of function caused by a resistance to the BMP–inhibitor NOGGIN and an altered signaling effect. Residue N445, situated within overlapping receptor and antagonist interfaces, is highly conserved among the BMP family with the exception of BMP9 and BMP10, in which it is substituted with lysine. Like the mutant GDF5, both BMPs are insensitive to NOGGIN and show a high chondrogenic activity. Ectopic expression of BMP9 or the GDF5 mutants resulted in massive induction of cartilage in an in vivo chick model presumably by bypassing the feedback inhibition imposed by endogenous NOGGIN. Swapping residues at the mutation site alone was not sufficient to render Bmp9 NOG-sensitive; however, successive introduction of two additional substitutions imparted high to total sensitivity on customized variants of Bmp9. In conclusion, we show a new mechanism for abnormal joint development that interferes with a naturally occurring regulatory mechanism of BMP signaling

Heteromeric clusters of ubiquitinated ER-shaping proteins drive ER-phagy

Membrane-shaping proteins characterized by reticulon homology domains play an important part in the dynamic remodelling of the endoplasmic reticulum (ER). An example of such a protein is FAM134B, which can bind LC3 proteins and mediate the degradation of ER sheets through selective autophagy (ER-phagy)1. Mutations in FAM134B result in a neurodegenerative disorder in humans that mainly affects sensory and autonomic neurons2. Here we report that ARL6IP1, another ER-shaping protein that contains a reticulon homology domain and is associated with sensory loss3, interacts with FAM134B and participates in the formation of heteromeric multi-protein clusters required for ER-phagy. Moreover, ubiquitination of ARL6IP1 promotes this process. Accordingly, disruption of Arl6ip1 in mice causes an expansion of ER sheets in sensory neurons that degenerate over time. Primary cells obtained from Arl6ip1-deficient mice or from patients display incomplete budding of ER membranes and severe impairment of ER-phagy flux. Therefore, we propose that the clustering of ubiquitinated ER-shaping proteins facilitates the dynamic remodelling of the ER during ER-phagy and is important for neuronal maintenance.</p

A922 Sequential measurement of 1 hour creatinine clearance (1-CRCL) in critically ill patients at risk of acute kidney injury (AKI)

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