Schizosaccharomyces pombe nup97, which Genetically Interacts with mex67, is essential for growth and involved in mRNA export

We have isolated previously three synthetic lethal mutants in Schizosaccharomyces pombe, which genetically interact with mex67, in order to identify the genes involved in mRNA export. A novel nup97 gene was isolated by complementation of the growth defect in one of the synthetic lethal mutants, SLMex3. The nup97 gene contains one intron and encodes an 851 amino-acid protein that is similar to nucleoporins, Npp106p in S. pombe and Nic96p in Saccharomyces cerevisiae. The nup97 gene is essential for vegetative growth, and nup97 null mutant harboring pREP41X-Nup97 showed poly(A)+ RNA export defect when expression of nup97 is repressed in the presence of thiamine. These results suggest that nup97 is involved in mRNA export from the nucleus to cytoplasm.     

Protocol for the recovery of in vivo matured canine oocytes based on once daily measurement of serum progesterone

The collection of in vivo matured canine oocytes relies on the accurate prediction of ovulation. The present study was designed to develop a protocol for the recovery of in vivo matured canine oocytes based on once daily measurements of serum progesterone (P(4)) concentrations. Blood samples (2 mL) were collected every day at 0900 h, and P(4) concentrations were analyzed using a DSL-3900 ACTIVE((R)) Progesterone Coated-Tube Radioimmunoassay Kit. The average number of oocytes at the metaphase II (M II) stage was significantly higher at or after 72 h (6.7 to 7.5) compared to 56 h (1.7) following ovulation. The highest numbers of corpora lutea, and therefore the highest numbers of oocytes, were recovered from bitches with initial ovulatory P(4) concentrations ranging from 6.0 to 8.0 ng/ mL (12.2 and 11.4, respectively) compared to from 4.0 to 4.9 ng/ mL (9.6 and 8.8, respectively; p < 0.05). The average number of M II oocytes recovered at 84 h from bitches with initial ovulatory P(4) levels of 5.0 to 5.9 ng/mL (7.7) was higher compared to bitches with P(4) levels of 4.0 to 4.9 ng/ mL (3.5) and 6.0 to 8.0 ng/ mL (4.8; p < 0.05). When oocyte recovery time was adjusted for initial ovulatory P(4) concentration, no significant difference in recovery rates or oocyte quality were observed. In conclusion, once daily measurements of P(4) can be used to predict ovulation in bitches, and oocyte recovery time should be adjusted for initial ovulatory serum P(4) concentrations.     

Colorimetric biosensor using dual-amplification of enzyme-free reaction through universal hybridization chain reaction system

On-site genetic detection needs to develop a sensitive and straightforward biosensor without special equipment, which can detect various genetic biomarkers. Hybridization chain reaction (HCR) amplifying signal isothermally could be considered as a good candidate for on-site detection. Here, we developed a novel genetic biosensor on the basis of enzyme-free dual-amplification of universal hybridization chain reaction (uHCR) and hemin/G-quadruplex horseradish peroxidase (HRP)-mimicking DNAzyme. The uHCR is the strategy which enables simple design for multiple target detection by the introduction of target-specific trigger hairpin without changing the whole system according to a target change. Also, HRP-mimicking DNAzyme could produce a sensitive and quantitative colorimetric signal with increased stability with a limit of detection (LOD) of 5.67 nM. The universality of the uHCR biosensor was proven by the detection of four different targets (miR-21, miR-125b, KRAS-Q61K, and BRAF-V600E) for cancer diagnosis. The uHCR biosensor showed specificity that could discriminate single-nucleotide polymorphism. Moreover, the uHCR biosensor could detect targets in the diluted serum sample. Overall, the uHCR biosensor demonstrated the potential for field testing with a simple redesign without complicated steps or special equipment using a universal hairpin system and enzyme-free amplification. This strategy could enable stable and sensitive detection of a variety of targets. Therefore, it could be applied to urgent detection of various pathogens, remote diagnosis, and self-screening of diseases.     

Ancient-to-modern secular changes in Korean stature

Statural growth in human populations is a sensitive indicator of socio-economic well-being, and improvements in socio-economic status are reflected in secular increases in adult height. In the present study, we investigated the statures of historical Korean societies to show how stature changed over time. Applying Fujii's equation, derived from modern Japanese, to the measurement of femora removed from 15th- to 19th-century Joseon tombs, the average heights of Korean adults during the Joseon dynasty were estimated to be 161.1 ± 5.6 cm and 148.9 ± 4.6 cm for males and females, respectively. Plotting statures for successive historical societies against time revealed that Korean heights remained relatively unchanged through to the end of the 19th century, a pattern that differs from that seen in many Western countries in which stature transiently decreases after the Middle Ages. In contrast, a sharp increase in Korean stature was observed at the beginning of the 20th century, similar to trends seen in other nations (although exact timing varies in different countries). There were no accompanying changes of stature sexual dimorphism. The data reported in this study reflect the unique historical experience of Korea; the relative isolation of Joseon society, the late onset of modernization (at the end of the 19th century), and the later occurrence of industrialization (during the 1960s).     

Covalent NEDD8 Conjugation Increases RCAN1 Protein Stability and Potentiates Its Inhibitory Action on Calcineurin

Similar to ubiquitin, regulatory roles for NEDD8 (neural precursor cell-expressed developmentally down-regulated 8) are being clarified during cell growth, signal transduction, immune response, and development. However, NEDD8 targets and their functional alterations are not well known. Regulator of calcineurin 1 (RCAN1/DSCR1P1) is located near the Down syndrome critical region on the distal part of chromosome 21, and its gene product is an endogenous inhibitor of calcineurin signaling. RCAN1 is modified by ubiquitin and consequently undergoes proteasomal degradation. Here we report that NEDD8 is conjugated to RCAN1 (RCAN1-1S) via three lysine residues, K96, K104, and K107. Neddylation enhances RCAN1 protein stability without affecting its cellular location. In addition, we found that neddylation significantly inhibits proteasomal degradation of RCAN1, which may underlie the ability of NEDD8 to enhance RCAN1 stability. Furthermore, neddylation increases RCAN1 binding to calcineurin, which potentiates its inhibitory activity toward downstream NFAT signaling. The present study provides a new regulatory mechanism of RCAN1 function and highlights an important role for diverse RCAN1-involved cellular physiology.     

Late chronotypes are associated with neoadjuvant chemotherapy-induced nausea and vomiting in women with breast cancer

Neoadjuvant chemotherapy, that is, the administration of chemotherapy before surgery, has been commonly used for locally advanced breast cancer to improve the surgical outcomes and increase the opportunity for breast-conserving therapy. Women with breast cancer often receive an anthracycline-based regimen as the neoadjuvant chemotherapy, which is associated with a high risk of emesis. Despite the development of novel antiemetics, chemotherapy-induced nausea and vomiting (CINV) has been commonly reported as a major adverse effect, affecting the quality of life of the patients. However, the factors predicting CINV in women with breast cancer undergoing neoadjuvant chemotherapy remain unclear. In this single-institution, prospective, observational study conducted at an outpatient cancer centre in the Republic of Korea from November 2013 to March 2016, we analysed women with breast cancer who planned to be treated with neoadjuvant chemotherapy before surgery. Candidate factors associated with CINV were assessed before neoadjuvant chemotherapy using the Munich Chronotype Questionnaire, Pittsburgh Sleep Quality Index and Hospital Anxiety and Depression Scale. CINV was assessed after chemotherapy by using the Multinational Association of Supportive Care in Cancer Antiemesis Tool. Of a total of 143 participants, 7 patients were lost to follow-up and 2 patients were excluded due to changes in their treatment plan; thus, 134 patients were finally included in the analyses. Overall, 48.5% of the participants experienced CINV, with delayed CINV prevalence (42.5%) being more common than acute (39.6%). In the univariate analyses, overall CINV was significantly associated with late chronotypes (odds ratio [OR], 3.49; 95% confidence interval [CI], 1.37–8.87; p = 0.009), a history of nausea/vomiting (OR, 2.19; 95% CI, 1.10–4.37; p = 0.026) and anxiety (OR, 2.25; 95% CI, 1.05–4.81; p = 0.036). In the multivariate analyses, late chronotypes (OR, 3.53; 95% CI, 1.27–9.79; p = 0.015) and a history of nausea/vomiting (OR, 2.83; 95% CI, 1.31–6.13; p = 0.008) remained significantly associated with CINV. In conclusion, in women with breast cancer undergoing neoadjuvant chemotherapy before surgery, late chronotypes were found to have an increased risk of CINV; these data suggest that clinicians need to assess and consider the chronotype in the management of CINV.     

The identification of a domain in Escherichia coli elongation factor Tu that interacts with elongation factor Ts.

A method has been developed to search for the elongation factor Tu (EF-Tu) domain(s) that interact with elongation factor Ts (EF-Ts). This method is based on the suppression of Escherichia coli EF-Tu-dominant negative mutation K136E, a mutation that exerts its effect by sequestering EF-Ts. We have identified nine single-amino acid- substituted suppression mutations in the region 146-199 of EF-Tu. These mutations are R154C, P168L, A174V, K176E, D181G, E190K, D196G, S197F, and I199V. All suppression mutations but one (R154C) significantly affect EF-Tu's ability to interact with EF-Ts under equilibrium conditions. Moreover, with the exception of mutation A174V, the GDP affinity of EF-Tu appears to be relatively unaffected by these mutations. These results suggest that the domain of residues 154 to 199 on EF-Tu is involved in interacting with EF-Ts. These suppression mutations are also capable of suppressing dominant negative mutants N135D and N135I to various degrees. This suggests that dominant negative mutants N135D and N135I are likely to have the same molecular basis as the K136E mutation. The method we have developed in this study is versatile and can be readily adapted to map other regions of EF-Tu. A model of EF-Ts-catalyzed guanine-nucleotide exchange is discussed.     

Characterization of in-frame proteins encoded by cvaA, an essential gene in the colicin V secretion system: CvaA* stabilizes CvaA to enhance secretion.

Colicin V (ColV), an antibacterial peptide toxin, uses a dedicated signal sequence-independent export system for its extracellular secretion in Escherichia coli. The products of at least three genes (a chromosomal tolC gene and two plasmid-born cvaA and cvaB genes) are involved in this process. To characterize the gene products, the cvaA gene was subcloned and expressed under the control of T7 RNA polymerase promoter. Two in-frame proteins, CvaA and CvaA*, were expressed and identified. DNA sequences predicted that both proteins have two potential translational initiation sites. N-terminal peptide sequencing showed that the translation of CvaA starts from a TTG, 11 amino acids upstream of the previously proposed ATG initiation site. CvaA* is translated from an upstream ATG. Expression of both CvaA and CvaA* was induced by the iron chelator 2,2'-dipyridyl, indicating that cvaA is negatively regulated at least partially by Fur. CvaA*-depleted cells were found to secrete less ColV, based on reduced activity in the supernatant, than did wild type, which was recovered by the addition of a plasmid producing CvaA*. Interestingly, CvaA*-depleted and wild-type cells had similar levels of intracellular ColV activity. Translational fusions showed that the syntheses of ColV and CvaA are not affected by CvaA* depletion. However, CvaA in CvaA*-depleted cells was less stable than that in wild-type cells, indicating that CvaA* may directly or indirectly affect the stability of CvaA. We conclude that CvaA* is not essential for ColV secretion but that it enhances the ColV secretion by stabilizing the CvaA protein.