Determining amino acid requirements from repeated observations on indicator amino acid oxidation method by mixed-effect change-point regression models

In nutrition studies, it is often of primary interest to determine the critical threshold value of some biological quantities. To determine the amino acid requirement, the tracer approach including the indicator amino acid oxidation method is useful for the investigation of human subjects. In this approach, measurements of amino acids other than the test amino acid are often repeatedly carried out with various intakes of the test amino acid. Change-point regression models have often been applied to determine the amino acid requirement. However, within-subject dependence due to repeated measurements has not been sufficiently taken into account. In this paper, we propose a mixed-effect change-point model to estimate the amino acid requirements when utilizing the tracer approach. Inference based on Akaike Information Criteria is introduced to include selection of the optimal model and construction of a confidence interval. Our method can easily be applied with a standard software package, and we found that appropriate accounting for within-subject dependence may lead to a much narrower confidence interval. We recommend application of a mixed-effect change-point regression model to determine the amino acid requirements in studies utilizing the tracer approach

Dependence of weak interaction rates on the nuclear composition during stellar core collapse

We investigate the influences of the nuclear composition on the weak interaction rates of heavy nuclei during the core collapse of massive stars. The nuclear abundances in nuclear statistical equilibrium (NSE) are calculated by some equation of state (EOS) models including in-medium effects on nuclear masses. We systematically examine the sensitivities of electron capture and neutrino-nucleus scattering on heavy nuclei to the nuclear shell effects and the single-nucleus approximation. We find that the washout of the shell effect at high temperatures brings significant change to weak rates by smoothing the nuclear abundance distribution: the electron capture rate decreases by ∼20% in the early phase and increases by ∼40% in the late phase at most, while the cross section for neutrino-nucleus scattering is reduced by ∼15%. This is because the open-shell nuclei become abundant instead of those with closed neutron shells as the shell effects disappear. We also find that the single-nucleus description based on the average values leads to underestimations of weak rates. Electron captures and neutrino coherent scattering on heavy nuclei are reduced by ∼80% in the early phase and by ∼5% in the late phase, respectively. These results indicate that NSE like EOS accounting for shell washout is indispensable for the reliable estimation of weak interaction rates in simulations of core-collapse supernovae

Five-point Likert scaling on MRI predicts clinically significant prostate carcinoma

Background: To clarify the relationship between the probability of prostate cancer scaled using a 5-point Likert system and the biological characteristics of corresponding tumor foci. Methods: The present study involved 44 patients undergoing 3.0-Tesla multiparametric MRI before laparoscopic radical prostatectomy. Tracing based on pathological and MRI findings was performed. The relationship between the probability of cancer scaled using the 5-point Likert system and the biological characteristics of corresponding tumor foci was evaluated. Results: A total of 102 tumor foci were identified histologically from the 44 specimens. Of the 102 tumors, 55 were assigned a score based on MRI findings (score 1: n = 3; score 2: n = 3; score 3: n = 16; score 4: n = 11 score 5: n = 22), while 47 were not pointed out on MRI. The tracing study revealed that the proportion of >0.5 cm3 tumors increased according to the upgrade of Likert scores (score 1 or 2: 33 %; score 3: 68.8 %; score 4 or 5: 90.9 %, χ2 test, p 7 also increased from scale 2 to scale 5 (scale 2: 0 %; scale 3: 56.3 %; scale 4: 72.7 %; 5: 90.9 %, χ2 test, p = 0.0001). On using score 3 or higher as the threshold of cancer detection on MRI, the detection rate markedly improved if the tumor volume exceeded 0.5 cm3 (<0.2 cm3: 10.3 %; 0.2-0.5 cm3: 25 %; 0.5-1.0 cm3: 66.7 %; 1.0 < cm3: 92.1 %). Conclusions: Each Likert scale favobably reflected the corresponding tumor’s volume and Gleason score. Our observations show that “score 3 or higher” could be a useful threshold to predict clinically significant carcinoma when considering treatment options

Zinc binding of a Cys2His2-type zinc finger protein is enhanced by the interaction with DNA

Zinc finger proteins specifically recognize DNA sequences and, therefore, play a crucial role in living organisms. In this study the Zn(II)-, and DNA-binding of 1MEY#, an artificial zinc finger protein consisting of three finger units was characterized by multiple methods. Fluorimetric, circular dichroism and isothermal calorimetric titrations were applied to determine the accurate stability constant of a zinc finger protein. Assuming that all three zinc finger subunits behave identically, the obtained thermodynamic data for the Zn(II) binding were ΔHbinding site =  − (23.5 − 28.0) kcal/mol (depending on the applied protonation state of the cysteines) and logβ’pH 7.4 = 12.2 ± 0.1, being similar to those of the CP1 consensus zinc finger peptide. The specific DNA binding of the protein can be characterized by logβ’pH 7.4 = 8.20 ± 0.08, which is comparable to the affinity of the natural zinc finger proteins (Sp1, WT1, TFIIIA) toward DNA. This value is ~ 1.9 logβ’ unit higher than those determined for semi- or nonspecific DNA binding. Competitive circular dichroism and electrophoretic mobility shift measurements revealed that the conditional stability constant characteristic for Zn(II) binding of 1MEY# protein increased by 3.4 orders of magnitude in the presence of its target DNA sequence

Involvement of CTCF in transcription regulation of EGR1 at early G1 phase as an architecture factor

Early growth response 1 (EGR1) is a transcription factor and regulates cellular processes such as proliferation, differentiation, and apoptosis. The expression of EGR1 is rapidly induced in response to several stimuli, and it activates the expression of downstream target genes involved in signaling cascades. EGR1 gene is also known to be transcribed in early G1 phase. However, the regulation of EGR1 transcription in early G1 phase is not clarified well. Here we found that CCCTC-binding factor (CTCF), a chromatin binding protein, is required to transcribe EGR1 gene at the onset of early G1 phase. We found that CTCF mediated the formation of higher-order chromatin structures among CTCF binding sites located in the EGR1 locus. Disruption of the CTCF-dependent higher-order chromatin structure using nuclease-dead Cas9 (dCas9)-mediated interference reduced the EGR1 transcription in early G1 phase. Collectively, we propose that CTCF has functional roles for the temporal expression of EGR1 in early G1 phase through regulation of higher-order chromatin structure organization

Vitamin B6 Prevents IL-1β Protein Production by Inhibiting NLRP3 Inflammasome Activation

Vitamin B6 includes six water-soluble vitamers: pyridoxal (PL), pyridoxamine (PM), pyridoxine (PN), and their phosphorylated forms. Pyridoxal 5o\u27-phosphate (PLP) is an important cofactor for many metabolic enzymes. Several lines of evidence demonstrate that blood levels of PLP are significantly lower in patients with inflammation than in control subjects and that vitamin B6 has anti-inflammatory effects, with therapeutic potential for a variety of inflammatory diseases. Although one of our group previously demonstrated that PL inhibits the NF-κB pathway, the molecular mechanism by which vitamin B6 suppresses inflammation is not well understood. Here, we showed that both PL and PLP suppressed the expression of cytokine genes in macrophages by inhibiting Toll-like receptor (TLR)-mediated TAK1 phosphorylation and the subsequent NF-κB and JNK activation. Furthermore, PL and PLP abolished NLRP3-dependent caspase-1 processing and the subsequent secretion of mature IL-1β and IL-18 in LPS-primed macrophages. In contrast, PM and PN had little effect on IL-1β production. PLP, but not PL, markedly reduced the production of mitochondrial reactive oxygen species (ROS) in peritoneal macrophages. Importantly, PL and PLP reduced IL-1β production induced by LPS and ATP, or by LPS alone, in mice. Moreover, PL and PLP protected mice from lethal endotoxic shock. Collectively, these findings reveal novel anti-inflammatory activities for vitamin B6 and suggest its potential for preventing inflammatory diseases driven by the NLRP3 inflammasome. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc

Mitotic phosphorylation of CCCTC-binding factor (CTCF) reduces its DNA binding activity

During mitosis, higher order chromatin structures are disrupted and chromosomes are condensed to achieve accurate chromosome segregation. CCCTC-binding factor (CTCF) is a highly conserved and ubiquitously expressed C2H2-type zinc finger protein which is considered to be involved in epigenetic memory through regulation of higher order chromatin architecture. However, the regulatory mechanism of CTCF in mitosis is still unclear. Here we found that the DNA-binding activity of CTCF is regulated in a phosphorylation-dependent manner during mitosis. The linker domains of the CTCF zinc finger domain were found to be phosphorylated during mitosis. The phosphorylation of linker domains impaired the DNA-binding activity in vitro. Mutation analyses showed that amino acid residues (Thr289, Thr317, Thr346, Thr374, Ser402, Ser461, and Thr518) located in the linker domains were phosphorylated during mitosis. Based on these results, we propose that the mitotic phosphorylation of the linker domains of CTCF is important for the dissociation of CTCF from mitotic chromatin

Histone acetylation-independent transcription stimulation by a histone chaperone

Histone chaperones are thought to be important for maintaining the physiological activity of histones; however, their exact roles are not fully understood. The physiological function of template activating factor (TAF)-I, one of the histone chaperones, also remains unclear; however, its biochemical properties have been well studied. By performing microarray analyses, we found that TAF-I stimulates the transcription of a sub-set of genes. The transcription of endogenous genes that was up-regulated by TAF-I was found to be additively stimulated by histone acetylation. On performing an experiment with a cell line containing a model gene integrated into the chromosome, TAF-I was found to stimulate the model gene transcription in a histone chaperone activity-dependent manner additively with histone acetylation. TAF-I bound to the core histones and remodeled the chromatin structure independent of the N-terminal histone tail and its acetylation level in vitro. These results suggest that TAF-I remodel the chromatin structure through its interaction with the core domain of the histones, including the histone fold, and this mechanism is independent of the histone acetylation status