Proximity-Directed Labeling Reveals a New Rapamycin-Induced Heterodimer of FKBP25 and FRB in Live Cells

Mammalian target of rapamycin (mTOR) signaling is a core pathway in cellular metabolism, and control of the mTOR pathway by rapamycin shows potential for the treatment of metabolic diseases. In this study, we employed a new proximity biotin-labeling method using promiscuous biotin ligase (pBirA) to identify unknown elements in the rapamycin-induced interactome on the FK506-rapamycin binding (FRB) domain in living cells. FKBP25 showed the strongest biotin labeling by FRB-pBirA in the presence of rapamycin. Immunoprecipitation and immunofluorescence experiments confirmed that endogenous FKBP25 has a rapamycin-induced physical interaction with the FRB domain. Furthermore, the crystal structure of the ternary complex of FRB-rapamycin-FKBP25 was determined at 1.67-angstrom resolution. In this crystal structure we found that the conformational changes of FRB generate a hole where there is a methionine-rich space, and covalent metalloid coordination was observed at C2085 of FRB located at the bottom of the hole. Our results imply that FKBP25 might have a unique physiological role related to metallomics in mTOR signaling.ope

ArmA and RmtB Were the Predominant 16S RMTase Genes Responsible for Aminoglycoside-resistant Isolates in Korea

Pathogenic gram-negatives that produce 16S ribosomal RNA methyltransferases (16S RMTases) have already been distributed all over the world. To investigate the predominance of aminoglycoside resistance associated with 16S RMTases in Korea, we collected a total of 222 amikacin resistant Gram-negative clinical isolates from patient specimens between 1999 and 2015 from three hospital banks across Korea. ArmA and nntB were the predominant 16S RMTase genes responsible for aminoglycoside-resistant isolates circulating in Korean community settings although only one rmtA-producing isolate was detected in 2006.1

Anesthetic experience of a patient with tracheomegaly -A case report-

Tracheomegaly or tracheobronchomegaly is a rare syndrome that consists of marked dilatation of the trachea and the major bronchi, and this is usually due to a congenital defect of the elastic and muscle fibers of the tracheobroncheal tree. Physicians have had only limited experience with performing anesthesia in patients with this type of syndrome. This syndorme is diagnosed by roentenological investigation and this condition is frequently associated with chronic respiratory infection and partial airway obstruction. In this report, we present a case of performing tracheostomy for a patient with tracheomegaly, and this was probably secondary to mechanical ventilator therapy. The regular tracheostomy tube did not provided sufficient length to allow the cuff to lie properly in the trachea in this patient. Because of the peri-cuff air leakgae and hypercapnea after tracheostomy, we needed a longer tracheostomy tube. But we didn't have such a tube and we didn't know any other method, so we couldn't perform tracheostomy. Therefore, we introduced a method of reducing the length of the endotracheal tube to a suitable size until a longer tracheostomy tube can be obtained for those patients having tracheomegaly

Trib2 regulates the pluripotency of embryonic stem cells and enhances reprogramming efficiency

Embryonic stem (ES) cells are pluripotent cells characterized by self-renewability and differentiation potential. Induced pluripotent stem (iPS) cells are ES cell-equivalent cells derived from somatic cells by the introduction of core reprogramming factors. ES and iPS cells are important sources for understanding basic biology and for generating therapeutic cells for clinical applications. Tribbles homolog 2 (Trib2) functions as a scaffold in signaling pathways. However, the relevance of Trib2 to the pluripotency of ES and iPS cells is unknown. In the present study, we elucidated the importance of Trib2 in maintaining pluripotency in mouse ES cells and in generating iPS cells from somatic cells through the reprogramming process. Trib2 expression decreased as ES cells differentiated, and Trib2 knockdown in ES cells changed their colony morphology while reducing the activity of alkaline phosphatase and the expression of the pluripotency marker genes Oct4, Sox2, Nanog and Klf4. Trib2 directly interacted with Oct4 and elevated Oct4 promoter activity. During the generation of iPS cells, Trib2 knockdown decreased the reprogramming efficiency of mouse embryonic fibroblasts, whereas Trib2 overexpression significantly increased their reprogramming efficiency. In summary, our results suggest that Trib2 is important for maintaining self-renewal in ES cells and for pluripotency induction during the reprogramming process

Helicobacter pylori infection combined with DENA revealed altered expression of p53 and 14-3-3 isoforms in Gulo−/− mice

AbstractUnlike most other mammals, human bodies do not have the ability to synthesize vitamin C inside of their own bodies. Therefore, humans must obtain vitamin C through daily diet. Gulo−/− mice strain is known with deficiency, in which vitamin C intake can be controlled by diet like human, and would be valuable for investigating the molecular mechanism of various diseases. In the present study, we established Gulo−/− mice model and investigated the differentially expressed proteins in stomach tissue of Gulo−/− mice after Helicobacter pylori-infected, and followed by DENA, using immunohistochemistry and proteomic approach. The results of immunohistochemistry analysis of stomach tissue showed that the tumor suppressor, p53 protein, expression was significantly decreased (p<0.05) but not messenger RNA (mRNA) transcriptional level, and 14-3-3ε, 14-3-3δ, Ki-67 and cleaved caspase 3 expressions were significantly increased (p<0.05) by H. Pylori infection, and followed by DENA treatment in Gulo−/− mice. Moreover, knockdown of 14-3-3 isoforms (14-3-3ε, 14-3-3σ, 14-3-3ζ and 14-3-3η) were significantly increased sub-G1 phase (characteristics of apoptosis) in AGS cells and, phenotypic changes like cell shrinkage, density and cleaved nuclei were also observed. Proteome analyses showed that 14-3-3σ, 14-3-3η, and tropomyosin alpha-1 chain were down-regulated, and Hspd1 protein and HSC70 were up-regulated after H. Pylori-infection, and followed by DENA. The combined results of immunohistochemistry and proteomic analysis suggest that H. pylori altered the p53 and 14-3-3 isoforms expression and DENA further enhanced the H. pylori effect, which might be involved in carcinogenesis and metastasis of gastric cancer on Gulo−/− mice

A remnant choledochal cyst after choledochal cyst excision treated with a lumen-apposing metal stent: a case report

A lumen-apposing metal stent (LAMS) is a saddle-shaped stent with large flanges at both ends, thereby preventing stent migration and helping with approximation of the adjacent structures. We report the case of a 25-year-old female with remnant choledochal cyst which was successfully treated with LAMS after initial treatment failure with a plastic stent. Although complete excision of the cyst is the definite treatment of choledochal cysts, endoscopic ultrasonography-guided cystoduodenostomy can be considered in cases wherein surgery is not feasible and dysplasia is not present. LAMS may be preferred to plastic stents for effective resolution of remnant choledochal cyst and prevention of ascending infection

Association between glycemic status and the risk of acute pancreatitis: a nationwide population-based study

Background Although diabetes is reportedly associated with the occurrence of acute pancreatitis (AP), the risk of AP according to the duration and severity of diabetes is not yet clear. We aimed to investigate the risk of AP based on glycemic status and the presence of comorbidities using a nationwide population-based study. Methods We enrolled 3,912,496 adults who underwent health examinations under the National Health Insurance Service in 2009. All participants were categorized by glycemic status as normoglycemic, impaired fasting glucose (IFG), or diabetes. Baseline characteristics and the presence of comorbidities at the time of health check-up were investigated, and the occurrence of AP was followed up until 31 December 2018. We estimated the adjusted hazard ratios (aHRs) for AP occurrence according to the glycemic status, duration of diabetes (new-onset, duration < 5 years, or ≥ 5 years), type and number of anti-diabetic medications, and presence of comorbidities. Results During the observation period of 32,116,716.93 person-years, 8,933 cases of AP occurred. Compared with normoglycemia, the aHRs (95% confidence interval) were 1.153 (1.097–1.212) in IFG, 1.389 (1.260–1.531) in new-onset diabetes, 1.634 (1.496–1.785) in known diabetes < 5 years, and 1.656 (1.513–1.813) in patients with known diabetes aged ≥ 5 years. The presence of comorbidities associated with diabetes severity had a synergistic effect on the relationship between diabetes and AP occurrence. Conclusion As glycemic status worsens, the risk of AP increases, and there is a synergistic effect when comorbidities coexist. To reduce the risk of AP, active control of factors that can cause AP should be considered in patients with long-standing diabetes and comorbidities

LONP1 and ClpP cooperatively regulate mitochondrial proteostasis for cancer cell survival

Mitochondrial proteases are key components in mitochondrial stress responses that maintain proteostasis and mitochondrial integrity in harsh environmental conditions, which leads to the acquisition of aggressive phenotypes, including chemoresistance and metastasis. However, the molecular mechanisms and exact role of mitochondrial proteases in cancer remain largely unexplored. Here, we identified functional crosstalk between LONP1 and ClpP, which are two mitochondrial matrix proteases that cooperate to attenuate proteotoxic stress and protect mitochondrial functions for cancer cell survival. LONP1 and ClpP genes closely localized on chromosome 19 and were co-expressed at high levels in most human cancers. Depletion of both genes synergistically attenuated cancer cell growth and induced cell death due to impaired mitochondrial functions and increased oxidative stress. Using mitochondrial matrix proteomic analysis with an engineered peroxidase (APEX)-mediated proximity biotinylation method, we identified the specific target substrates of these proteases, which were crucial components of mitochondrial functions, including oxidative phosphorylation, the TCA cycle, and amino acid and lipid metabolism. Furthermore, we found that LONP1 and ClpP shared many substrates, including serine hydroxymethyltransferase 2 (SHMT2). Inhibition of both LONP1 and ClpP additively increased the amount of unfolded SHMT2 protein and enhanced sensitivity to SHMT2 inhibitor, resulting in significantly reduced cell growth and increased cell death under metabolic stress. Additionally, prostate cancer patients with higher LONP1 and ClpP expression exhibited poorer survival. These results suggest that interventions targeting the mitochondrial proteostasis network via LONP1 and ClpP could be potential therapeutic strategies for cancer