A Graphical Model to Diagnose Product Defects with Partially Shuffled Equipment Data

The diagnosis of product defects is an important task in manufacturing, and machine learning-based approaches have attracted interest from both the industry and academia. A high-quality dataset is necessary to develop a machine learning model, but the manufacturing industry faces several data-collection issues including partially shuffled data, which arises when a product ID is not perfectly inferred and yields an unstable machine learning model. This paper introduces latent variables to formulate a supervised learning model that addresses the problem of partially shuffled data. The experimental results show that our graphical model deals with the shuffling of product order and can detect a defective product far more effectively than a model that ignores shuffling.This work has supported by the National Research Foundation of Korea (NRF) grant funded by the Korea government (MSIT) (2019R1A2C1088255)

The mammalian Sec6/8 complex interacts with Ca2+ signaling complexes and regulates their activity

The localization of various Ca(2+) transport and signaling proteins in secretory cells is highly restricted, resulting in polarized agonist-stimulated Ca(2+) waves. In the present work, we examined the possible roles of the Sec6/8 complex or the exocyst in polarized Ca(2+) signaling in pancreatic acinar cells. Immunolocalization by confocal microscopy showed that the Sec6/8 complex is excluded from tight junctions and secretory granules in these cells. The Sec6/8 complex was found in at least two cellular compartments, part of the complex showed similar, but not identical, localization with the Golgi apparatus and part of the complex associated with Ca(2+) signaling proteins next to the plasma membrane at the apical pole. Accordingly, immunoprecipitation (IP) of Sec8 did not coimmunoprecipitate betaCOP, Golgi 58K protein, or mannosidase II, all Golgi-resident proteins. By contrast, IP of Sec8 coimmunoprecipitates Sec6, type 3 inositol 1,4,5-trisphosphate receptors (IP(3)R3), and the Gbetagamma subunit of G proteins from pancreatic acinar cell extracts. Furthermore, the anti-Sec8 antibodies coimmunoprecipitate actin, Sec6, the plasma membrane Ca(2+) pump, the G protein subunits Galphaq and Gbetagamma, the beta1 isoform of phospholipase C, and the ER resident IP(3)R1 from brain microsomal extracts. Antibodies against the various signaling and Ca(2+) transport proteins coimmunoprecipitate Sec8 and the other signaling proteins. Dissociation of actin filaments in the immunoprecipitate had no effect on the interaction between Sec6 and Sec8, but released the actin and dissociated the interaction between the Sec6/8 complex and Ca(2+) signaling proteins. Hence, the interaction between the Sec6/8 and Ca(2+) signaling complexes is likely mediated by the actin cytoskeleton. The anti-Sec6 and anti-Sec8 antibodies inhibited Ca(2+) signaling at a step upstream of Ca(2+) release by IP(3). Disruption of the actin cytoskeleton with latrunculin B in intact cells resulted in partial translocation of Sec6 and Sec8 from membranes to the cytosol and interfered with propagation of agonist-evoked Ca(2+) waves. Our results suggest that the Sec6/8 complex has multiple roles in secretory cells including governing the polarized expression of Ca(2+) signaling complexes and regulation of their activity.ope

Homer Binds TRPC Family Channels and Is Required for Gating of TRPC1 by IP3 Receptors

Receptor signaling at the plasma membrane often releases calcium from intracellular stores. For example, inositol triphosphate (IP3) produced by receptor-coupled phospholipase C activates an intracellular store calcium channel, the IP3R. Conversely, stores can induce extracellular calcium to enter the cell through plasma membrane channels, too. How this “reverse” coupling works was unclear, but store IP3Rs were proposed to bind and regulate plasma membrane TRP cation channels. Here, we demonstrate that the adaptor protein, termed Homer, facilitates a physical association between TRPC1 and the IP3R that is required for the TRP channel to respond to signals. The TRPC1-Homer-IP3R complex is dynamic and its disassembly parallels TRPC1 channel activation. Homer's action depends on its ability to crosslink and is blocked by the dominant-negative immediate early gene form, H1a. Since H1a is transcriptionally regulated by cellular activity, this mechanism can affect both short and long-term regulation of TRPC1 function.ope

Carbonic anhydrase 12 mutation modulates membrane stability and volume regulation of aquaporin 5

Patients carrying the carbonic anhydrase12 E143K mutation showed the dry mouth phenotype. The mechanism underlying the modulation of aquaporin 5 and function in the salivary glands by carbonic anhydrase12 remains unknown. In this study, we identified the mislocalised aquaporin 5 in the salivary glands carrying the E143K. The intracellular pH of E143K cells was more acidic than that of the cells carrying wild type. To evaluate the role of carbonic anhydrase12 on the volume regulation of aquaporin 5, the submandibular gland cells were subjected to hypotonic stimuli. E143K enhanced the extent of swelling of cells on hypotonicity. Aquaporin 5 modulates water influx through ion transporters to prevent osmotic imbalance. These results suggest that the carbonic anhydrase12 E143K, including acidification or inflammation, mediates volume dysregulation by the loss of aquaporin 5. Thus, carbonic anhydrase12 may determine sensible effects on the cellular osmotic regulation by modulating aquaporin 5.ope

Genetic and pharmacologic inhibition of the Ca2+ influx channel TRPC3 protects secretory epithelia from Ca2+-dependent toxicity

BACKGROUND & AIMS: Excessive Ca2+ influx mediates many cytotoxic processes, including those associated with autoimmune inflammatory diseases such as acute pancreatitis and Sjögren syndrome. Transient receptor potential (canonical) channel (TRPC) 3 is a major Ca2+ influx channel in pancreatic and salivary gland cells. We investigated whether genetic or pharmacologic inhibition of TRPC3 protects pancreas and salivary glands from Ca2+-dependent damage. METHODS: We developed a Ca2+-dependent model of cell damage for salivary gland acini. Acute pancreatitis was induced by injection of cerulein into wild-type and Trpc3-/- mice. Mice were also given the Trpc3-selective inhibitor pyrazole 3 (Pyr3). RESULTS: Salivary glands and pancreas of Trpc3-/- mice were protected from Ca2+-mediated cell toxicity. Analysis of Ca2+ signaling in wild-type and Trpc3-/- acini showed that Pyr3 is a highly specific inhibitor of Tprc3; it protected salivary glands and pancreas cells from Ca2+-mediated toxicity by inhibiting the Trpc3-mediated component of Ca2+ influx. CONCLUSIONS: TRPC3-mediated Ca2+ influx mediates damage to pancreas and salivary glands. Pharmacologic inhibition of TRPC3 with the highly selective TRPC3 inhibitor Pyr3 might be developed for treatment of patients with acute pancreatitis and Sjögren syndromeope

Gating of store-operated channels by conformational coupling to ryanodine receptors

We report here that RyRs interact with and gate the store-operated hTrp3 and Icrac channels. This gating contributes to activation of hTrp3 and Icrac by agonists. Coupling of hTrp3 to IP3Rs or RyRs in the same cells was found to be mutually exclusive. Biochemical and functional evidence suggest that mutually exclusive coupling reflects clustering and segregation of hTrp3-IP3R and hTrp3-RyR complexes in plasma membrane microdomains. Gating of CCE by RyRs indicates that gating by conformational coupling is not unique to skeletal muscle but is a general mechanism for communication between events in the plasma and endoplasmic reticulum membranes.ope

Convergence of IRBIT, phosphatidylinositol (4,5) bisphosphate, and WNK/SPAK kinases in regulation of the Na+-HCO3− cotransporters family

Fluid and HCO3− secretion is a vital function of secretory epithelia, involving basolateral HCO3− entry through the Na+-HCO3− cotransporter (NBC) NBCe1-B, and luminal HCO3− exit mediated by cystic fibrosis transmembrane conductance regulator (CFTR) and solute carrier family 26 (SLC26) Cl−/HCO3− exchangers. HCO3− secretion is highly regulated, with the WNK/SPAK kinase pathway setting the resting state and the IRBIT/PP1 pathway setting the stimulated state. However, we know little about the relationships between the WNK/SPAK and IRBIT/PP1 sites in the regulation of the transporters. The first 85 N-terminal amino acids of NBCe1-B function as an autoinhibitory domain. Here we have identified a positively charged module within NBCe1-B(37-65) that is conserved in NBCn1-A and all 20 members of the NBC superfamily except NBCe1-A. This module is required for the interaction and activation of NBCe1-B and NBCn1-A by IRBIT and their regulation by phosphatidylinositol 4,5-bisphosphate (PIP2). Activation of the transporters by IRBIT and PIP2 is nonadditive but complementary. Phosphorylation of Ser65 mediates regulation of NBCe1-B by SPAK, and phosphorylation of Thr49 is required for regulation by IRBIT and SPAK. Sequence searches using the NBCe1-B regulatory module as a template identified a homologous sequence in the CFTR R domain and Slc26a6 sulfat transporter and antisigma factor antagonist (STAS) domain. Accordingly, the R and STAS domains bind IRBIT, and the R domain is required for activation of CFTR by IRBIT. These findings reveal convergence of regulatory modalities in a conserved domain of the NBC that may be present in other HCO3− transporters and thus in the regulation of epithelial fluid and HCO3− secretion.ope

Chloride Influx of Anion Exchanger 2 Was Modulated by Calcium-Dependent Spinophilin in Submandibular Glands

Secretory glands including salivary glands by many hormonal inputs produce and secrete biological fluids determined by variety of ion transporters. Spinophilin is a multifunctional scaffolding protein, which involved in receptor signaling and regulation of anion exchangers AE2 activity. We found that spinophilin expressed in salivary glands. The role of salivary spinophilin on the modulation of chloride/bicarbonate exchange remains unknown. The spinophilin enhanced AE2 activity and associated with a STE20/SPS1-related kinase and showed an additive effect on the modulation of the activity of AE2. The cholinergic stimulation and subsequent intracellular Ca2+ increase was required for the interaction with AE2 and spinophilin and abrogated the enhanced effect of spinophilin on Cl- transporting activity. Ductal chloride/bicarbonate exchange activity was increased in pretreatment with carbachol. The CaMKII inhibitor KN-93 suppressed the chloride/bicarbonate exchange activity of ducts, suggesting that CaMKII was required for ductal chloride/bicarbonate exchange activity. Additionally, microtubule destabilization by nocodazole attenuated the interaction of AE2 and spinophilin and almost abolished the ductal chloride/bicarbonate exchange activity. The treatment of siRNA-spinophilin on the isolated salivary ducts also reduced the ductal chloride/bicarbonate exchange activity. Therefore, role of salivary spinophilin on AE2 may facilitate the Cl- influx from basolateral in salivary glands in response to cholinergic inputs.ope

Intracellular Ca 2+-Mediated AE2 Is Involved in the Vectorial Movement of HaCaT Keratinocyte

Keratinocyte migration is initiated toward the wound skin barrier as a crucial process in wound healing. However, the migratory machinery used by keratinocytes is relatively unknown. Histamine signaling, including an increase in the Ca2+ signal, mediated the enhanced protein expression and chloride/bicarbonate exchange activity of anion exchanger AE2 in keratinocytes. In this study, we applied an agarose spot assay to induce a vectorial motion. The vectorial stimulation of the histamine-containing agarose spot enhanced the HaCaT keratinocyte migration, compared to non-directional stimulation. AE2 is associated with the vectorial movement of HaCaT keratinocytes. Enhanced expression of AE2 was mainly associated with an increase in Ca2+ and was abolished by the treatment with the Ca2+ chelating agent BAPTA-AM. These findings revealed that the directionality of Ca2+-exerted stimulation can play a prominent role in facilitating migration through the involvement of AE2 as a migratory machinery in HaCaT keratinocytes.ope

Regulation of Ca2+-release-activated Ca2+ current (Icrac) by ryanodine receptors in inositol 1,4,5-trisphosphate-receptor-deficient DT40 cells

Persistence of capacitative Ca(2+) influx in inositol 1,4,5-trisphosphate (IP(3)) receptor (IP(3)R)-deficient DT40 cells (DT40(IP(3)R-/-)) raises the question of whether gating of Ca(2+)-release activated Ca(2+) current (I(crac)) by conformational coupling to Ca(2+)-release channels is a general mechanism of gating of these channels. In the present work we examined the properties and mechanism of activation of I(crac) Ca(2+) current in wild-type and DT40(IP(3)R-/-) cells. In both cell types passive depletion of internal Ca(2+) stores by infusion of EGTA activated a Ca(2+) current with similar characteristics and time course. The current was highly Ca(2+)-selective and showed strong inward rectification, all typical of I(crac). The activator of ryanodine receptor (RyR), cADP-ribose (cADPR), facilitated activation of I(crac), and the inhibitors of the RyRs, 8-N-cADPR, ryanodine and Ruthenium Red, all inhibited I(crac) activation in DT40(IP(3)R-/-) cells, even after complete depletion of intracellular Ca(2+) stores by ionomycin. Wild-type and DT40(IP(3)R-/-) cells express RyR isoforms 1 and 3. RyR levels were adapted in DT40(IP(3)R-/-) cells to a lower RyR3/RyR1 ratio than in wild-type cells. These results suggest that IP(3)Rs and RyRs can efficiently gate I(crac) in DT40 cells and explain the persistence of I(crac) gating by internal stores in the absence of IP(3)Rs.ope