Interferon signaling and hypercytokinemia-related gene expression in the blood of antidepressant non-responders

Only 50% of patients with depression respond to the first antidepressant drug administered. Thus, biomarkers for prediction of antidepressant responses are needed, as predicting which patients will not respond to antidepressants can optimize selection of alternative therapies. We aimed to identify biomarkers that could predict antidepressant responsiveness using a novel data-driven approach based on statistical pattern recognition. We retrospectively divided patients with major depressive disorder into antidepressant responder and non-responder groups. Comprehensive gene expression analysis was performed using peripheral blood without narrowing the genes. We designed a classifier according to our own discrete Bayes decision rule that can handle categorical data. Nineteen genes showed differential expression in the antidepressant non-responder group (n = 15) compared to the antidepressant responder group (n = 15). In the training sample of 30 individuals, eight candidate genes had significantly altered expression according to quantitative real-time polymerase chain reaction. The expression of these genes was examined in an independent test sample of antidepressant responders (n = 22) and non-responders (n = 12). Using the discrete Bayes classifier with the HERC5, IFI6, and IFI44 genes identified in the training set yielded 85% discrimination accuracy for antidepressant responsiveness in the 34 test samples. Pathway analysis of the RNA sequencing data for antidepressant responsiveness identified that hypercytokinemia- and interferon-related genes were increased in non-responders. Disease and biofunction analysis identified changes in genes related to inflammatory and infectious diseases, including coronavirus disease. These results strongly suggest an association between antidepressant responsiveness and inflammation, which may be useful for future treatment strategies for depression

Engineering of cyclodextrin glucanotransferases and the impact for biotechnological applications

Cyclodextrin glucanotransferases (CGTases) are industrially important enzymes that produce cyclic α-(1,4)-linked oligosaccharides (cyclodextrins) from starch. Cyclodextrin glucanotransferases are also applied as catalysts in the synthesis of glycosylated molecules and can act as antistaling agents in the baking industry. To improve the performance of CGTases in these various applications, protein engineers are screening for CGTase variants with higher product yields, improved CD size specificity, etc. In this review, we focus on the strategies employed in obtaining CGTases with new or enhanced enzymatic capabilities by searching for new enzymes and improving existing enzymatic activities via protein engineering

Regulation of transketolase like 1 gene expression in the murine one-cell stage embryos.

In mice, transcription from the zygotic genome starts at the mid-one-cell stage after fertilization. Previous studies showed that an enhancer is not required for transcription at this stage, and that the enhancer-dependent mechanism of transcription is established during the two-cell stage. However, these results were obtained using reporter gene assays with promoters derived from viruses, rather than from endogenous genes. We conducted a reporter-gene assay using the promoter of Tktl1, which is transcribed after fertilization, to investigate the mechanism regulating gene expression at the one-cell stage. When a plasmid containing the 2467 bp upstream and 25 bp downstream of the Tktl1 transcription start site (TSS) was microinjected into the nuclei of growing oocytes, and one-cell stage and early and late two-cell-stage embryos, transcriptional activity was detected in the one-cell- and two-cell-stage embryos, but not in the oocytes. It was highest at the early two-cell stage and was reduced at the late two-cell stage. The decrease in activity at the late two-cell stage was prevented by inhibiting the second round of DNA replication, suggesting that the transcriptionally repressive state is established during the two-cell stage by a mechanism coupled to DNA replication. When the Tktl1 promoter was deleted to leave 56 bp upstream of the TSS which includes GC and TATA boxes, transcriptional activity was still detected in one-cell-stage embryos, but not early or late two-cell-stage embryos. The core promoter of Tktl1 alone seems to be able to induce basal transcription at the one-cell stage. These results suggest that repressive chromatin is established after fertilization in two steps, which occur during the transition from the one- to two-cell stage and during DNA replication at the two-cell stage

The elements required for transcriptional activity of the core promoter region of <i>Tktl1</i>.

<p>(A) Transcription-factor-binding sites in the 56 bp upstream of the TSS of <i>Tktl1</i>. (B) Effects of mutation of the GC box and TATA box-like elements on the transcriptional activity of p<i>Tktl1-56</i>. The sequences GGGCGG (GC box) and TTTTAA (TATA box like) were mutated to GGAAAG and TTGCGA, respectively. Plasmids (500 ng/µl) were microinjected into the male pronuclei of one-cell-stage embryos. Experiments were performed three times and the results are presented as means ± SEM. Asterisks indicate significant differences (Student's <i>t</i>-test; <i>P</i><0.05).</p

Effect of DNA synthesis inhibition on <i>Tktl1</i> transcriptional activity in two-cell-stage embryos.

<p>Transcription of p<i>Tktl1-2467+25</i> was measured in embryos in which DNA synthesis was inhibited by treatment with aphidicolin. Experiments were performed three times and the results are presented as means ± SEM. Asterisks indicate significant differences (Student's <i>t</i>-test; <i>P</i><0.05).</p

Transcriptional activity of the 56<i>Tktl1</i> in one- and two-cell-stage embryos.

<p>Plasmids (200 ng/µl) were microinjected into the male pronuclei of one-cell-stage embryos and the nuclei of early and late two-cell-stage embryos. Microinjection was performed at 9, 18 and 27 h after insemination in one-cell and early and late two-cell-stage embryos, respectively. Experiments were performed four times and the results are presented as means ± SEM. Asterisks indicate significant differences (Student's <i>t</i>-test; <i>P</i><0.05).</p

PCR primers.

<p>PCR primers.</p

Transcriptional activity of <i>Tktl1</i>-promoter deletion mutants in one-cell-stage embryos.

<p>(A) The upstream region of the <i>Tktl1</i> promoter (<i>Tktl1-2467+25</i>) was deleted to leave 227 bp (<i>Tktl1-227+25</i>) and 56 bp (<i>Tktl1-56+25</i>), and transcriptional activity was then evaluated. (B) The 25 bp downstream of the TSS was deleted from p<i>Tktl1-56+25</i> and transcriptional activity was measured. Plasmids (500 ng/µl) were microinjected into the male pronuclei of one-cell-stage embryos. The experiments were performed at least 11 times and the results are presented as means ± SEM. Asterisks indicate significant differences (Student's <i>t</i>-test; <i>P</i><0.05).</p

Expression of Tktl1 mRNA during preimplantation development.

<p>The expression of <i>Tktl1</i> in MII stage oocytes and preimplantation embryos was assessed by semi-quantitative RT-PCR. Embryos were collected at the following time points after insemination: one-cell stage, 13 h; two-cell stage, 28 h; 4-cell stage, 45 h; morula, 60 h; and blastocyst, 96 h. The experiments were performed four times and the data are presented as means ± SEM. The value for two-cell-stage embryos was set to 1.0 and relative values were calculated for the other stages.</p

Transcriptional activity of the <i>Tktl1</i> promoter in oocytes and preimplantation embryos.

<p>Plasmids containing the promoter regions of the <i>Tktl1</i> (p<i>Tktl1-2467+25</i>) and <i>Zp3</i> genes were microinjected into growing oocytes (A), one-cell-stage embryos (B), and early and late two-cell-stage embryos (C and D), and their transcriptional activities were then evaluated by luciferase reporter gene assay. The transcriptional activity of a plasmid without a promoter (None) was used as a control to determine the background level. Plasmids (200 ng/µl) were microinjected into the nucleus. Experiments were performed at least three times and the results are presented as means ± SEM. Asterisks indicate significant differences (Student's <i>t</i>-test; <i>P</i><0.05).</p