CaV2.2 (N-type) voltage-gated calcium channels are activated by SUMOylation pathways

SUMOylation is an important post-translational modification process involving covalent attachment of SUMO (Small Ubiquitin-like MOdifier) protein to target proteins. Here, we investigated the potential for SUMO-1 protein to modulate the function of the CaV2.2 (N-type) voltage-gated calcium channel (VGCC), a protein vital for presynaptic neurotransmitter release. Co-expression of SUMO-1, but not the conjugation-deficient mutant SUMO-1ΔGG, increased heterologously-expressed CaV2.2 Ca2+ current density, an effect potentiated by the conjugating enzyme Ubc9. Expression of sentrin-specific protease (SENP)-1 or Ubc9 alone, had no effect on recombinant CaV2.2 channels. Co-expression of SUMO-1 and Ubc9 caused an increase in whole-cell maximal conductance (Gmax) and a hyperpolarizing shift in the midpoint of activation (V1/2). Mutation of all five CaV2.2 lysine residues to arginine within the five highest probability (>65 %) SUMOylation consensus motifs (SCMs) (construct CaV2.2-Δ5KR), produced a loss-of-function mutant. Mutagenesis of selected individual lysine residues identified K394, but not K951, as a key residue for SUMO-1-mediated increase in CaV2.2 Ca2+ current density. In synaptically-coupled superior cervical ganglion (SCG) neurons, SUMO-1 protein was distributed throughout the cell body, axons and dendrites and presumptive presynaptic terminals, whilst SUMO-1ΔGG protein was largely confined to the cell body, in particular, the nucleus. SUMO-1 expression caused increases in paired excitatory postsynaptic potential (EPSP) ratio at short (20− 120 ms) inter-stimuli intervals in comparison to SUMO-1ΔGG, consistent with an increase in residual presynaptic Ca2+ current and an increase in release probability of synaptic vesicles. Together, these data provide evidence for CaV2.2 VGCCs as novel targets for SUMOylation pathways

Experimental Investigation and Numerical Simulation of a Self-Wiping Corotating Parallel Octa-Screw Extruder

An octa-screw extruder (OSE) is equipment for pelletizing, blending, and mixing polymers and composites. In this study, the degree of resin filling, residence time distribution (RTD) of molten resin, and temperature profile in the octa-screw extruder were evaluated both experimentally and numerically. An intermeshing corotating parallel octa-screw kneading extruder was used for the experiments. For the comparison study, the results obtained from this extruder were compared with the twin-screw extruder. High-density polyethylene was selected as the material for extrusion. Meanwhile, a numerical code, based on a 2.5 D finite element method derived from the Hele–Shaw flow model, was developed to simulate the octa-screw extrusion process. The empirical outcomes suggest that octa-screw extrusion exhibited a narrower RTD of the molten resin compared with the twin-screw extrusion, suggesting better extrudate quality. The octa-screw extrusion also showed a lower temperature profile than twin-screw extrusion. The results of the simulation were also found to be in good agreement with experimental measurements. Experimental and numerical investigations of an OSE enable detailed comprehension and visualization of resin distribution in the entire length of the OSE, thus providing advantages in terms of process optimization

Transcriptome Analysis Reveals Differential Gene Expression between the Closing Ductus Arteriosus and the Patent Ductus Arteriosus in Humans

The ductus arteriosus (DA) immediately starts closing after birth. This dynamic process involves DA-specific properties, including highly differentiated smooth muscle, sparse elastic fibers, and intimal thickening (IT). Although several studies have demonstrated DA-specific gene expressions using animal tissues and human fetuses, the transcriptional profiles of the closing DA and the patent DA remain largely unknown. We performed transcriptome analysis using four human DA samples. The three closing DA samples exhibited typical DA morphology, but the patent DA exhibited aorta-like elastic lamellae and poorly formed IT. A cluster analysis revealed that samples were clearly divided into two major clusters, the closing DA and patent DA clusters, and showed distinct gene expression profiles in IT and the tunica media of the closing DA samples. Cardiac neural crest-related genes such as JAG1 were highly expressed in the tunica media and IT of the closing DA samples compared to the patent DA sample. Abundant protein expressions of jagged 1 and the differentiated smooth muscle marker calponin were observed in the closing DA samples but not in the patent DA sample. Second heart field-related genes such as ISL1 were enriched in the patent DA sample. These data indicate that the patent DA may have different cell lineages compared to the closing DA

SAD-B Phosphorylation of CAST Controls Active Zone Vesicle Recycling for Synaptic Depression

SummaryShort-term synaptic depression (STD) is a common form of activity-dependent plasticity observed widely in the nervous system. Few molecular pathways that control STD have been described, but the active zone (AZ) release apparatus provides a possible link between neuronal activity and plasticity. Here, we show that an AZ cytomatrix protein CAST and an AZ-associated protein kinase SAD-B coordinately regulate STD by controlling reloading of the AZ with release-ready synaptic vesicles. SAD-B phosphorylates the N-terminal serine (S45) of CAST, and S45 phosphorylation increases with higher firing rate. A phosphomimetic CAST (S45D) mimics CAST deletion, which enhances STD by delaying reloading of the readily releasable pool (RRP), resulting in a pool size decrease. A phosphonegative CAST (S45A) inhibits STD and accelerates RRP reloading. Our results suggest that the CAST/SAD-B reaction serves as a brake on synaptic transmission by temporal calibration of activity and synaptic depression via RRP size regulation