The role of bystin in embryo implantation and in ribosomal biogenesis

Human bystin was identified as a cytoplasmic protein directly binding to trophinin, a cell adhesion molecule potentially involved in human embryo implantation. Although the trophinin gene is unique to mammals, the bystin gene (BYSL) is conserved across eukaryotes. Recent studies show that bystin plays a key role during the transition from silent trophectoderm to an active trophoblast upon trophinin-mediated cell adhesion. Bystin gene knockout and knockdown experiments demonstrate that bystin is essential for embryonic stem cell survival and trophectoderm development in the mouse. Furthermore, biochemical analysis of bystin in human cancer cells and mouse embryos indicates a function in ribosomal biogenesis, specifically in processing of 18S RNA in the 40S subunit. Strong evidence that BYSL is a target of c-MYC is consistent with a role for bystin in rapid protein synthesis, which is required for actively growing cells

1-(Trifluoromethyl)vinyllithium

InChI = 1S/C3H2F3.Li/c1-2-3(4,5)6;/h1H2; InChIKey = YDBNXCHLLXTABG-UHFFFAOYSA-N (reagent used as a versatile vinylation component) Physical Data: thermally unstable; decomposes quickly at temperatures above −78 °C. Solubility: soluble in diethyl ether, pentane, and THF. Form Supplied in: not available commercially. Preparative Method: prepared from the reaction of commercially available 2-bromo-3,3,3-trifluoropropene with n-, sec-, or tert-butyllithium in diethyl ether at −100 to −110 °C. Handling, Storage, and Precautions: prepared and transferred at low temperature (≤ −100 °C) under inert atmosphere (Ar); air and moisture sensitive; handle in fume hood

The molecular basis for genetic polymorphism of human deoxyribonuclease I: identification of the nucleotide substitution that generates the fourth allele

AbstractIn addition to the three alleles commonly responsible for the protein polymorphism of human deoxyribonuclease I, a mutation encoded by a fourth allele, DNASEI*4, was detected by isoelectric focusing. All 8 exons covering the entire open reading frame of the human DNase I gene were amplified by the polymerase chain reaction and subjected to direct sequencing. Only one nucleotide substitution, a C-to-G transition (CAG → GAG), in the codon for amino acid 9 of the mature enzyme was found. This substitution resulted in the replacement of Gln with Glu (Q9E)

Utility of Contrast-Enhanced FDG-PET/CT in the Clinical Management of Pancreatic Cancer Impact on Diagnosis, Staging, Evaluation of Treatment Response, and Detection of Recurrence

Objectives: Fluorodeoxyglucose (FDG)-positron emission tomography/contrast-enhanced computed tomography (PET/CE-CT) involving whole-body scanning first by non-CE-CT and FDG-PET followed by CE-CT has been used for detailed examination of pancreatic lesions. We evaluated PET/CE-CT images with regard to differential diagnosis, staging, treatment response, and postoperative recurrence in pancreatic cancer. Methods: Positron emission tomography/CE-CT was conducted in 108 patients with pancreatic cancer and in 41 patients with other pancreatic tumor diseases. Results: The maximum standardized uptake value (SUVmax) overlapped in benign and malignant cases, suggesting that differential diagnosis of pancreatic tumors based on the SUVmax is difficult. In the evaluation of staging in 31 resectable pancreatic cancer by PET/CE-CT, the diagnostic accuracy rate was more than 80% for most factors concerning local invasion and 94% for distant metastasis but only 42% for lymph node metastasis. Significant positive correlations were found between the SUVmax and tumor size/markers, suggesting that SUVmax may be a useful indicator for the treatment response. Regarding the diagnosis of the postoperative recurrence, PET/CE-CT correctly detected local recurrence in all the 11 cases of recurrence, whereas abdominal CE-CT detected only 7 of 11 cases, suggesting that PET/CE-CT is superior in this context. Conclusions: Positron emission tomography/CE-CT is useful for the clinical management of pancreatic cancer

Efficacy and Safety of Three-dimensional Conformal Radiotherapy for Macroscopic Vascular Invasion of Hepatocellular Carcinoma

Chemotherapy is insufficient to treat macroscopic vascular invasion (MVI) of hepatocellular carcinoma (HCC). We retrospectively investigated the treatment outcomes of patients who underwent three-dimensional conformal radiotherapy (3D-CRT) for HCC MVI and analyzed prognostic factors by multivariate analysis using a Cox proportional hazard model. Sixty-five patients were studied. MVI sites were the portal vein (n=48 patients), portal and hepatic veins (n=8), and hepatic vein (n=9). The median irradiation dose was 50 Gy. The median survival time (MST) was 7.5 months. Performance status 2 or 3, modified albumin-bilirubin grade 2b or 3, and massive/diffuse type were poor prognostic factors. Nineteen patients (29%) with a treatment effect of 3 or 4 (≥ 50% of tumor necrosis or regression) at the irradiation sites according to the Response Evaluation Criteria in Cancer of the Liver showed longer survival than those with an effect of 1 or 2 (MST 18.7 vs. 5.9 months, p<0.001). No treatment-related death occurred. The hepatic function reserve was preserved in more than 70% of patients. 3D-CRT controlled HCC MVI safely and was suggested to be a good treatment option

Structural change of ribosomes during apoptosis: Degradation and externalization of ribosomal proteins in doxorubicin-treated Jurkat cells

金沢大学医薬保健研究域薬学系Changes in the amount and localization of human ribosomal proteins during apoptosis were determined. When total lysates of Jurkat cells undergoing apoptosis induced by doxorubicin were analyzed by Western blotting, degradation of three ribosomal proteins, S18, L5, and L14, was detected at 48 h after the induction of apoptosis. Decreases in the amounts of these three ribosomal proteins were also observed in ribosome-enriched fractions. These changes were partly abolished by the addition of the pan-caspase inhibitor z-VAD-fmk. Moreover, formation of the 80S ribosome complex appeared to be inhibited at 48 h after apoptosis induction. On the other hand, the rate of protein synthesis, assessed by measuring the incorporation of [35S]Met into bulk proteins, decreased as early as 12 h after the addition of doxorubicin. These results indicate that changes in the amount of ribosomal proteins and the overall structure of ribosomes in apoptosing cells occur after protein synthesis declines. Finally, analyses by flow cytometry, immunofluorescence, and Western blotting showed that six ribosomal proteins, S15, PO, L5, L6, L36a, and L41, were relocalized and expressed at the cell surface during apoptosis. The above results collectively indicate that ribosomes are structurally altered in apoptotic cells following inactivation of protein synthesis

Analysis of two human pre-ribosomal factors, bystin and hTsr1, highlights differences in evolution of ribosome biogenesis between yeast and mammals

Recent studies reveal that maturation of the 40S ribosomal subunit precursors in mammals includes an additional step during processing of the internal transcribed spacer 1 (ITS1), when compared with yeast Saccharomyces cerevisiae, even though the protein content of the pre-40S particle appears to be the same. Here, we examine by depletion with siRNA treatment the function of human orthologs of two essential yeast pre-ribosomal factors, hEnp1/bystin and hTsr1. Like their yeast orthologs, bystin is required for efficient cleavage of the ITS1 and further processing of this domain within the pre-40S particles, whereas hTsr1 is necessary for the final maturation steps. However, bystin depletion leads to accumulation of an unusual 18S rRNA precursor, revealing a new step in ITS1 processing that potentially involves an exonuclease. In addition, pre-40S particles lacking hTsr1 are partially retained in the nucleus, whereas depletion of Tsr1p in yeast results in strong cytoplasmic accumulation of pre-40S particles. These data indicate that ITS1 processing in human cells may be more complex than currently envisioned and that coordination between maturation and nuclear export of pre-40S particles has evolved differently in yeast and mammalian cells

Mutational and biochemical analysis of the DNA-entry nuclease EndA from Streptococcus pneumoniae

EndA is a membrane-attached surface-exposed DNA-entry nuclease previously known to be required for genetic transformation of Streptococcus pneumoniae. More recent studies have shown that the enzyme also plays an important role during the establishment of invasive infections by degrading extracellular chromatin in the form of neutrophil extracellular traps (NETs), enabling streptococci to overcome the innate immune system in mammals. As a virulence factor, EndA has become an interesting target for future drug design. Here we present the first mutational and biochemical analysis of recombinant forms of EndA produced either in a cell-free expression system or in Escherichia coli. We identify His160 and Asn191 to be essential for catalysis and Asn182 to be required for stability of EndA. The role of His160 as the putative general base in the catalytic mechanism is supported by chemical rescue of the H160A variant of EndA with imidazole added in excess. Our study paves the way for the identification and development of protein or low-molecular-weight inhibitors for EndA in future high-throughput screening assays