TANGO1 facilitates cargo loading at endoplasmic reticulum exit sites.

A genome-wide screen revealed previously unidentified components required for transport and Golgi organization (TANGO). We now provide evidence that one of these proteins, TANGO1, is an integral membrane protein localized to endoplasmic reticulum (ER) exit sites, with a luminal SH3 domain and a cytoplasmic proline-rich domain (PRD). Knockdown of TANGO1 inhibits export of bulky collagen VII from the ER. The SH3 domain of TANGO1 binds to collagen VII; the PRD binds to the COPII coat subunits, Sec23/24. In this scenario, PRD binding to Sec23/24 subunits could stall COPII carrier biogenesis to permit the luminal domain of TANGO1 to guide SH3-bound cargo into a growing carrier. All cells except those of hematopoietic origin express TANGO1. We propose that TANGO1 exports other cargoes in cells that do not secrete collagen VII. However, TANGO1 does not enter the budding carrier, which represents a unique mechanism to load cargo into COPII carriers.

[Pouchitis showed complete response to ustekinumab].

Pouchitis is the most common long-term complication following ileal pouch-anal anastomosis (IPAA) in patients with ulcerative colitis. Although several agents, including probiotics, steroids, and immunomodulators, have been used, the treatment of pouchitis remains challenging. Owing to the proven efficacy of biological therapy in inflammatory bowel disease, there is now growing evidence suggesting the potential benefits of biological therapy in refractory pouchitis. Here, we report the case of a 64-year-old woman with pouchitis due to ulcerative colitis who was successfully treated with ustekinumab (UST). The patient developed ulcerative pancolitis at the age of 35. Total colectomy and IPAA with J-pouch anastomosis were performed when the patient was 47 years old. Ileotomy closure was performed 6 months later. Postoperatively, the patient developed steroid-dependent pouchitis. Three years later, she developed steroid-induced diabetes. The patient has been taking 3mg of steroid for 20 years;therefore, her lifetime total steroid dose was 21g. The patient had over 20 episodes of bloody diarrhea a day. The last pouchoscopy in 20XX-9 revealed inflammatory stenosis with deep ulcerations of the afferent limb just before the ileoanal pouch junction. In July 20XX, when we took over her treatment, the policy of treatment was to withdraw her from steroids. Pouchoscopy revealed a widened but still tight afferent limb through which the scope could easily pass, and the ileoanal pouch still showed erosive ileitis without ulcers. Thiopurine administration and steroid tapering were initiated. Steroid tapering increased the erythrocyte sedimentation rate (ESR). As ESR increased, her arthritis exacerbated. Six months after the end of steroid administration, the patient consented to UST treatment. On April 20XX+1, the patient received her first 260-mg UST infusion. At this point, she experienced 14-15 episodes of muddy bloody stools. She had no abdominal pain;however, she experienced shoulder pain. Gradually, UST affected both pouchitis and arthritis. UST treatment was continued at 90mg subcutaneously every 12 weeks without abdominal pain recurrence. Eight months after the first UST infusion, nonsteroidal anti-inflammatory drugs were no longer necessary for shoulder pain. Follow-up pouchoscopy performed 14 months after UST optimization revealed a normal afferent limb without ulcerations in either segment. Pouchitis remission was maintained for over 2 years.

Structural Analysis of Intracellular Lipid Radicals by LC/MS/MS Using a BODIPY-Based Profluorescent Nitroxide Probe.

Free radical-mediated lipid peroxidation (LPO) induces the formation of numerous lipid radicals, which contribute to the development of several oxidative diseases. To understand the mechanism of LPO in biological systems and the significance of these radicals, identifying the structures of individual lipid radicals is imperative. In this study, we developed an analytical method based on liquid chromatography coupled with tandem mass spectrometry (LC/MS/MS) and a profluorescent nitroxide probe, N-(1-oxyl-2,2,6-trimethyl-6-pentylpiperidin-4-yl)-3-(5,5-difluoro-1,3-dimethyl-3H,5H-5l4-dipyrrolo[1,2-c:2',1'-f][1,3,2]diazaborinin-7-yl)propanamide (BDP-Pen), for the detailed structural analysis of lipid radicals. The MS/MS spectra of BDP-Pen-lipid radical adducts showed product ions and thus allow the prediction of the lipid radical structures and individual detection of isomeric adducts. Using the developed technology, we separately detected the isomers of arachidonic acid (AA)-derived radicals generated in AA-treated HT1080 cells. This analytical system is a powerful tool for elucidating the mechanism of LPO in biological systems.

Sequence Type Changes Associated with Decreasing Macrolide-Resistant Mycoplasma pneumoniae, Japan.

We compared sequence types (STs) of Mycoplasma pneumoniae isolates from Japan during 2002-2019. ST3 and ST14 dominated during 2002-2016, and ST7 and ST33 dominated during 2018-2019. These STs were associated with a decrease in macrolide-resistant strains after an epidemic of infection with M. pneumoniae during 2011-2012.

Remodeling of ER-exit sites initiates a membrane supply pathway for autophagosome biogenesis.

Autophagosomes are double-membrane vesicles generated during autophagy. Biogenesis of the autophagosome requires membrane acquisition from intracellular compartments, the mechanisms of which are unclear. We previously found that a relocation of COPII machinery to the ER-Golgi intermediate compartment (ERGIC) generates ERGIC-derived COPII vesicles which serve as a membrane precursor for the lipidation of LC3, a key membrane component of the autophagosome. Here we employed super-resolution microscopy to show that starvation induces the enlargement of ER-exit sites (ERES) positive for the COPII activator, SEC12, and the remodeled ERES patches along the ERGIC A SEC12 binding protein, CTAGE5, is required for the enlargement of ERES, SEC12 relocation to the ERGIC, and modulates autophagosome biogenesis. Moreover, FIP200, a subunit of the ULK protein kinase complex, facilitates the starvation-induced enlargement of ERES independent of the other subunits of this complex and associates via its C-terminal domain with SEC12. Our data indicate a pathway wherein FIP200 and CTAGE5 facilitate starvation-induced remodeling of the ERES, a prerequisite for the production of COPII vesicles budded from the ERGIC that contribute to autophagosome formation.

Testing theories of financial decision making.

We describe the observable content of some of the most widely used models of decision under uncertainty: models of translation invariant preferences. In particular, we characterize the models of variational, maxmin, constant absolute risk aversion, and constant relative risk aversion utilities. In each case we present a revealed preference axiom that is satisfied by a dataset if and only if the dataset is consistent with the corresponding utility representation. We test our axioms using data from an experiment on financial decisions.

The binding of TBK1 to STING requires exocytic membrane traffic from the ER.

Stimulator of interferon genes (STING) is essential for the type I interferon and pro-inflammatory responses against DNA pathogens. In response to the presence of cytosolic DNA, STING translocates from the endoplasmic reticulum (ER) to the Golgi, and activates TANK-binding kinase 1 (TBK1), a cytosolic kinase that is essential for the activation of STING-dependent downstream signalling. The organelles where TBK1 binds to STING remain unknown. Here we show that TBK1 binds to STING at the Golgi, not at the ER. Treatment with brefeldin A, an agent to block ER-to-Golgi traffic, or knockdown of Sar1, a small GTPase that regulates coat protein complex II (COP-II)-mediated ER-to-Golgi traffic, inhibited the binding of TBK1 to STING. Endogenous TBK1 was recruited to the Golgi when STING was transported to the Golgi, as shown by immunofluorescence microscopy. STING variants that constitutively induce the type I interferon response were found in patients with autoinflammatory diseases. Even these disease-causative STING variants could not bind to TBK1 when the STING variants were trapped in the ER. These results demonstrate that the Golgi is an organelle at which STING recruits and activates TBK1 for triggering the STING-dependent type I interferon response.

Chemical Events in Oligo(3-methoxythiophene) Coating Solutions and Their Effect on the Goldlike Coating Film Properties.

Metal-free, metal-like lustrous films may find applications in a variety of fields, and a study of the factors affecting their stability is highly desirable. In particular, chemical events occurring in the coating solutions might affect the supramolecular organization of the films and therefore the metal-like luster. Herein, the chemical events occurring in acetonitrile and nitromethane coating solutions of oligo(3-methoxythiophene) and their effect on the optical properties of the films were investigated by X-ray diffraction, UV-vis absorption, and viscosity measurements. In acetonitrile, the oligomers underwent gradual dedoping with time, but only small changes in viscosity were observed. The solution was applied to a glass plate to yield a dark brown film, which turned into a goldlike lustrous film by rubbing. In nitromethane, the supramolecular structure of the oligomers changed with time from the nonaggregated state to π-dimers and then to π-stacks, and the viscosity increased. The properties of the goldlike films prepared from this solution were greatly affected by this chemical event. Remarkably, the π-dimer solution provided the film with the highest specular reflectance, yellowness, greenness, brightness, and crystallinity.

Mechanisms for exporting large-sized cargoes from the endoplasmic reticulum.

Cargo proteins exported from the endoplasmic reticulum to the Golgi apparatus are typically transported in coat protein complex II (COPII)-coated vesicles of 60-90 nm diameter. Several cargo molecules including collagens and chylomicrons form structures that are too large to be accommodated by these vesicles, but their secretion still requires COPII proteins. Here, we first review recent progress on large cargo secretions derived especially from animal models and human diseases, which indicate the importance of COPII proteins. We then discuss the recent isolation of specialized factors that modulate the process of COPII-dependent cargo formation to facilitate the exit of large-sized cargoes from the endoplasmic reticulum. Based on these findings, we propose a model that describes the importance of the GTPase cycle for secretion of oversized cargoes. Next, we summarize reports that describe the structures of COPII proteins and how these results provide insight into the mechanism of assembly of the large cargo carriers. Finally, we discuss what issues remain to be solved in the future.

p24 family Tango(1) at the endoplasmic reticulum exit site to organize cargo exit.

The p24 family of proteins have been regarded as cargo receptors for endoplasmic reticulum (ER) to Golgi transport; however, their precise functions have yet to be revealed. In this issue, Pastor-Pareja and colleagues (https://doi.org/10.1083/jcb.202309045) show that the interaction of these proteins with Tango1 is critical for their localization at the ER exit site (ERES) and efficient transport of secretory proteins in Drosophila.

RhoA activation participates in rearrangement of processing bodies and release of nucleated AU-rich mRNAs.

Cytoplasmic ribonucleoprotein granules, known as processing bodies (P-bodies), contain a common set of conserved RNA-processing enzymes, and mRNAs with AU-rich elements (AREs) are delivered to P-bodies for translational silencing. Although the dynamics of P-bodies is physically linked to cytoskeletal network, it is unclear how small GTPases are involved in the P-body regulation and the ARE-mRNA metabolism. We found here that glucose depletion activates RhoA GTPase and alters the P-body dynamics in HeLa cells. These glucose-depleted effects are reproduced by the overexpression of the RhoA-subfamily GTPases and conversely abolished by the inhibition of RhoA activation. Interestingly, both RhoA activation and glucose depletion inhibit the mRNA accumulation and degradation. These findings indicate that RhoA participates in the stress-induced rearrangement of P-bodies and the release of nucleated ARE-mRNAs for their stabilization.

Evaluation of the enhancement of photosynthetic rate in a komatsuna (Brassica rapa L. var. perviridis) canopy with upward lighting using an optical simulation in a plant factory with artificial light.

In a plant factory with artificial light (PFAL), upward lighting is expected to prevent senescence and decrease in the photosynthetic capacity of the lower leaves in the canopy. Upward lighting may also increase the photosynthetic rate of a canopy by improving its photosynthetic photon flux density (PPFD) distribution. However, the net photosynthetic rate (Pn) of leaves is lower when the abaxial surface is irradiated than that when the adaxial surface is irradiated. The aim of this study was to estimate the PPFD in a PFAL and the Pn of plants using three-dimensional plant models and optical simulation. First, we measured the Pn of komatsuna (Brassica rapa L. var. perviridis) leaves under different conditions of the proportion (pad ) of PPFD on the adaxial surface to total PPFD on both surfaces and developed an equation for the light response curve of photosynthesis considering pad . When PPFD was low, except when it was 30 and 70 µmol m-2 s-1, Pn increased as pad increased, because the absorptance also increased with pad . Under high PPFD conditions, Pn was maximized at 67-83% of pad because the light would be distributed more efficiently for photosynthesis. Next, using optical simulation and the developed equation, we estimated the photosynthetic rate of a komatsuna canopy (CPn) under downward and upward lighting. The CPn increased by 1.08-1.13 times by combining downward and upward lighting due to the increase in the photosynthetic photon flux (PPF) of light incident on the canopy and the decrease in the spatial variation of PPFD on the leaves in the canopy. As the depreciation of lamps for upward lighting accounts for 7.5-9.0% of the production cost in a PFAL, even if the depreciation of lamps for upward lighting increased, enhancement of CPn by upward lighting would be cost-effective. We performed optical simulations under 220 conditions and evaluated them using CPn as an index. Moreover, we provided the proportion of PPF of upward lighting that improved CPn and discussed the reason for this improvement. The result shows that optical simulation is useful for evaluating the lighting design in a PFAL and analyzing the effects of the lighting design on the light environment and photosynthesis.

Functional genomics reveals genes involved in protein secretion and Golgi organization.

Yeast genetics and in vitro biochemical analysis have identified numerous genes involved in protein secretion. As compared with yeast, however, the metazoan secretory pathway is more complex and many mechanisms that regulate organization of the Golgi apparatus remain poorly characterized. We performed a genome-wide RNA-mediated interference screen in a Drosophila cell line to identify genes required for constitutive protein secretion. We then classified the genes on the basis of the effect of their depletion on organization of the Golgi membranes. Here we show that depletion of class A genes redistributes Golgi membranes into the endoplasmic reticulum, depletion of class B genes leads to Golgi fragmentation, depletion of class C genes leads to aggregation of Golgi membranes, and depletion of class D genes causes no obvious change. Of the 20 new gene products characterized so far, several localize to the Golgi membranes and the endoplasmic reticulum.

Cargo receptor Surf4 regulates endoplasmic reticulum export of proinsulin in pancreatic ß-cells.

Insulin is an essential peptide hormone that maintains blood glucose levels. Although the mechanisms underlying insulin exocytosis have been investigated, the mechanism of proinsulin export from the endoplasmic reticulum (ER) remains unclear. Here, we demonstrated that Surf4, a cargo receptor homolog, regulates the ER export of proinsulin via its recruitment to ER exit sites (ERES). Under high-glucose conditions, Surf4 expression was upregulated, and Surf4 proteins mainly localized to the ER at a steady state and accumulated in the ERES, along with proinsulin in rat insulinoma INS-1 cells. Surf4-knockdown resulted in proinsulin retention in the ER and decreased the levels of mature insulin in secretory granules, thereby significantly reducing insulin secretion. Surf4 forms an oligomer and can physically interact with proinsulin and Sec12, essential for COPII vesicle formation. Our findings suggest that Surf4 interacts with proinsulin and delivers it into COPII vesicles for ER export in co-operation with Sec12 and COPII.

Slippage- and load-induced changes in the crystalline orientation of oligo(3-methoxythiophene) powder to develop a gold-tone luster.

The achievement of molecular orientation control by rubbing and pressing poly(3-alkylthiophene)s is a powerful technique to improve the performance of organic electronic devices. We report here that the rubbing and pressing of blackish-brown 3-methoxythiophene oligomer powders yield layer and tablet samples with gold tones, respectively. Specular reflectivity, colorimetric, and X-ray diffraction measurements reveal that this gold tone is caused by an increase in the ratio of edge-on lamellar crystallites to face-on ones, which is promoted by rubbing/pressing. In contrast to the 3-alkylthiophene polymer, which develops a dominant face-on lamellar structure, rubbing of the 3-methoxythiophene oligomer increases the relative amount of edge-on lamellar crystallites to face-on lamellar ones. Furthermore, gold tone development in the tablet samples is limited to the near-surface area, despite the fact that pressure is also applied to the tablet bulk. These specific chemical events are explained by considering the repulsive interactions between the 3-methoxythiophene backbone and the functional groups on the surface of the substrate employed during the rubbing/pressing processes. Despite the lower applied pressure, gold tone development by rubbing is accompanied by a higher reflective property than by pressing because of the formation of larger relative amounts and sizes of edge-on lamellar crystallites, which are responsible for the gold tone.

Effects of maternal smoking during pregnancy on body composition in offspring.

BACKGROUND: The aim of the present cross-sectional study was to use objective methods to assess the association between maternal smoking and body composition in offspring. METHODS: A total of 2508 grade 4 school children were enrolled; all underwent lifestyle disease and passive smoking screening. Children were classified into four groups according to their urinary cotinine level and maternal smoking status during or before pregnancy. Items measured on lifestyle disease screening were compared among the four groups. RESULTS: Only degree of obesity (DO) and body mass index (BMI) were significantly associated with maternal smoking during pregnancy. The prevalence of both DO >20% and DO >30%, and BMI >22% and BMI >25% was highest in children of mothers who smoked during pregnancy. These children had a tendency toward shorter height and increased weight although it was not statistically significant. There were no significant differences between maternal smoking status and lipid profile among groups. Confounders such as food, exercise and sleep were able to be eliminated. CONCLUSION: Maternal smoking during pregnancy may be an independent risk factor of changing body composition in offspring, that is, shorter height and increased weight.

Remimazolam anesthesia for cardiac surgery with cardiopulmonary bypass: a case report.

BACKGROUND: Remimazolam has less cardiovascular depressant effects than propofol in non-cardiac surgical patients. However, the efficacy and safety of remimazolam in cardiac surgery with cardiopulmonary bypass (CPB) have not been reported. We present a case of successful anesthetic management using remimazolam in cardiac surgery with CPB. CASE PRESENTATION: A 76-year-old female was scheduled for mitral valve repair, tricuspid annuloplasty, maze procedure, and left atrial appendage closure. We used remimazolam in induction (6.0 mg/kg/h) and maintenance (0.6-1.0 mg/kg/h) of general anesthesia, and the bispectral index value was maintained in the range of 36 to 48 including the period of CPB. Hemodynamics, mixed venous oxygen saturation, and bilateral regional cerebral oxygen saturation were maintained within acceptable ranges. There was no intraoperative awareness/recall or serious complications associated with remimazolam throughout the perioperative period. CONCLUSIONS: Remimazolam can be used the same as other existing anesthetics in cardiac surgery with CPB.

The LIM protein Ajuba is required for ciliogenesis and left-right axis determination in medaka.

Cilia are microtubule-based organelles that are present on the surfaces of almost all vertebrate cells. Most cilia function as sensory or molecular transport structures. Malfunctions of cilia have been implicated in several diseases of human development. The assembly of cilia is initiated by the centriole (or basal body), and several centrosomal proteins are involved in this process. The mammalian LIM protein Ajuba is a well-studied centrosomal protein that regulates cell division but its role in ciliogenesis is unknown. In this study, we isolated the medaka homolog of Ajuba and showed that Ajuba localizes to basal bodies of cilia in growth-arrested cells. Knockdown of Ajuba resulted in randomized left-right organ asymmetries and altered expression of early genes responsible for left-right body axis determination. At the cellular level, we found that Ajuba function was essential for ciliogenesis in the cells lining Kupffer's vesicle; it is these cells that induce the asymmetric fluid flow required for left-right axis determination. Taken together, our findings identify a novel role for Ajuba in the regulation of vertebrate ciliogenesis and left-right axis determination.

Mitotic ER exit site dynamics: insights into blockade of secretion from the ER during mitosis.

How ER exit sites disassemble during mitosis is not well understood. Transport ANd Golgi Organization 1 (TANGO1, also known as MIA3), a cargo receptor originally identified for collagens, acts as a hub for ER exit site disassembly under the control of Casein Kinase 1 (CK1)-mediated phosphorylation and Protein Phosphatase 1 (PP1)-mediated dephosphorylation. Impaired dephosphorylation during mitosis induces ER exit site disassembly.

Mitotic ER Exit Site Disassembly and Reassembly Are Regulated by the Phosphorylation Status of TANGO1.

Golgi fragmentation and ER exit site disassembly are considered to be the leading causes of the mitotic block of secretion from the ER. Although the mechanisms of Golgi fragmentation have been extensively characterized, ER exit block early in mitosis is not well understood. We previously demonstrated that TANGO1 organizes ER exit sites by directly interacting with Sec16. In this study, we identified TANGO1 as a regulator of ER exit site disassembly during mitosis. TANGO1 phosphorylation was observed to be coordinately regulated by a kinase (CK1) and a phosphatase (PP1). CK1-mediated TANGO1 phosphorylation reduces binding to Sec16, leading to the disassembly of ER exit sites. CK1 constantly phosphorylates TANGO1, whereas PP1-mediated dephosphorylation of TANGO1 decreases during mitosis. Thus, the phosphorylation status of TANGO1, which is controlled by balanced activities of the kinase CK1 and the phosphatase PP1, regulates the organization of ER exit sites during the cell cycle.