[A Case of Multiple Renal Carcinomas with Three Different Subtypes].

We report a case of dialysis kidney with multiple renal carcinomas in three locations. A 74-year-old man who had a 20-year history of dialysis, was admitted to our hospital complaining of sudden right lateral area pain. Computed tomography (CT) scan revealed hemorrhage from the right dialyzed renal subcapsule. He underwent immediate transcatheter arterial embolization (TAE), but after 6 days the CT scan showed new active bleeding. A second TAE was performed, but, the CT scan several days later showed hemorrhage from the same site. So we decided to perform a right radical nephrectomy. Pathological results revealed three different renal carcinomas at sites different from the bleeding site. The presence of triple carcinomas in the same organ of the kidney is quite rare. Because of the high incidence of renal carcinoma associated with polycystic kidney disease in long-term hemodialysis patients and the possibility of multiple carcinomas as in this case, radical nephrectomy is recommended even in the case of spontaneous rupture.

Crystallographic snapshots of the EF-hand protein MCFD2 complexed with the intracellular lectin ERGIC-53 involved in glycoprotein transport.

The transmembrane intracellular lectin ER-Golgi intermediate compartment protein 53 (ERGIC-53) and the soluble EF-hand multiple coagulation factor deficiency protein 2 (MCFD2) form a complex that functions as a cargo receptor, trafficking various glycoproteins between the endoplasmic reticulum (ER) and the Golgi apparatus. It has been demonstrated that the carbohydrate-recognition domain (CRD) of ERGIC-53 (ERGIC-53CRD) interacts with N-linked glycans on cargo glycoproteins, whereas MCFD2 recognizes polypeptide segments of cargo glycoproteins. Crystal structures of ERGIC-53CRD complexed with MCFD2 and mannosyl oligosaccharides have revealed protein-protein and protein-sugar binding modes. In contrast, the polypeptide-recognition mechanism of MCFD2 remains largely unknown. Here, a 1.60 Šresolution crystal structure of the ERGIC-53CRD-MCFD2 complex is reported, along with three other crystal forms. Comparison of these structures with those previously reported reveal that MCFD2, but not ERGIC-53-CRD, exhibits significant conformational plasticity that may be relevant to its accommodation of various polypeptide ligands.

Improved secretion of glycoproteins using an N-glycan-restricted passport sequence tag recognized by cargo receptor.

MCFD2 and ERGIC-53, which are the products of causative genes of combined factor V and factor VIII deficiency, form a cargo receptor complex responsible for intracellular transport of these coagulation factors in the early secretory pathway. In this study, using an NMR technique, we successfully identified an MCFD2-binding segment from factor VIII composed of a 10 amino acid sequence that enhances its secretion. This prompted us to examine possible effects of attaching this sequence to recombinant glycoproteins on their secretion. We found that the secretion level of recombinant erythropoietin was significantly increased simply by tagging it with the passport sequence. Our findings not only provide molecular basis for the intracellular trafficking of coagulation factors and their genetic deficiency but also offer a potentially useful tool for increasing the production yields of recombinant glycoproteins of biopharmaceutical interest.

Rapid and reliable species identification of wild mushrooms by matrix assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS).

Mushrooms are a favourite natural food in many countries. However, some wild species cause food poisoning, sometimes lethal, due to misidentification caused by confusing fruiting bodies similar to those of edible species. The morphological inspection of mycelia, spores and fruiting bodies have been traditionally used for the identification of mushrooms. More recently, DNA sequencing analysis has been successfully applied to mushrooms and to many other species. This study focuses on a simpler and more rapid methodology for the identification of wild mushrooms via protein profiling based on matrix-assisted laser desorption/ionization mass spectrometry (MALDI-TOF MS). A preliminary study using 6 commercially available cultivated mushrooms suggested that a more reproducible spectrum was obtained from a portion of the cap than from the stem of a fruiting body by the extraction of proteins with a formic acid-acetonitrile mixture (1 + 1). We used 157 wild mushroom-fruiting bodies collected in the centre of Hokkaido from June to November 2014. Sequencing analysis of a portion of the ribosomal RNA gene provided 134 identifications of mushrooms by genus or species, however 23 samples containing 10 unknown species that had lower concordance rate of the nucleotide sequences in a BLAST search (less than 97%) and 13 samples that had unidentifiable poor or mixed sequencing signals remained unknown. MALDI-TOF MS analysis yielded a reproducible spectrum (frequency of matching score ≥ 2.0 was ≥6 spectra from 12 spectra measurements) for 114 of 157 samples. Profiling scores that matched each other within the database gave correct species identification (with scores of ≥2.0) for 110 samples (96%). An in-house prepared database was constructed from 106 independent species, except for overlapping identifications. We used 48 wild mushrooms that were collected in autumn 2015 to validate the in-house database. As a result, 21 mushrooms were identified at the species level with scores ≥2.0 and 5 mushrooms at the genus level with scores ≥1.7, although the signals of 2 mushrooms were insufficient for analysis. The remaining 20 samples were recognized as "unreliable identification" with scores <1.7. Subsequent DNA analysis confirmed that the correct species or genus identifications were achieved by MALDI-TOF MS for the 26 former samples, whereas the 18 mushrooms with poorly matched scores were species that were not included in the database. Thus, the proposed MALDI-TOF MS coupled with our database could be a powerful tool for the rapid and reliable identification of mushrooms; however, continuous updating of the database is necessary to enrich it with more abundant species.

Structural basis for disparate sugar-binding specificities in the homologous cargo receptors ERGIC-53 and VIP36.

ERGIC-53 and VIP36 are categorized as leguminous type (L-type) lectins, and they function as cargo receptors for trafficking certain N-linked glycoproteins in the secretory pathway in animal cells. They share structural similarities in their carbohydrate recognition domains (CRDs) but exhibit distinct sugar-binding specificities and affinities. VIP36 specifically interacts with the α1,2-linked D1 mannosyl arm without terminal glucosylation, while ERGIC-53 shows a broader specificity and lower binding affinity to the high-mannose-type oligosaccharides, irrespective of the presence or absence of the non-reducing terminal glucose residue at the D1 arm. In this study, we determined the crystal structure of ERGIC-53-CRD in complex with their binding partner, MCFD2 and the α1,2 mannotriose which corresponds to the trisaccharide of the D1 arm of high-mannose-type glycans. ERGIC-53 can interact with the D1 trimannosyl arm in two alternative modes, one of which is similar but distinct from that previously observed for VIP36. ERGIC-53 has a shallower sugar-binding pocket than VIP36 because of the single amino acid substitution, Asp-to-Gly. This enables ERGIC-53 to accommodate the non-reducing terminal glucose of the D1 arm in its CRD. In the other interaction mode, the 3-OH group of the terminal mannose was situated outward with respect to the sugar binding pocket, also enabling the Glcα1-3 linkage formation without steric hindrance. Our findings thus provide a structural basis for the broad sugar-binding specificity of the ERGIC-53/MCFD2 cargo receptor complex.

[Catamenial pneumothorax; report of a case].

A 40-year-old female had right-sided pneumothorax. She was treated with a chest tube. On the 2nd day, her menstruation started. Because of a persistent air leak, she underwent a thoracoscopic operation on the 15th day. During the operation, we noted a pulmonary bulla at the apex and multiple blue berry spots both on the visceral pleura and the tendinous part of the diaphragm. We performed a resection of the pulmonary bulla, partial diaphragmatic excision plus suture. Section of visceral pleura and diaphragm shows ductal structures, which is insistent with endometriosis. The serum CA125 level decreased to 39 U/ml. Following surgery, oral contraceptives was started and no recurrence was encountered thereafter.