Tatsuya Nishikawa

RNA Binding Proteins and Genome Integrity

Genome integrity can be threatened by various endogenous or exogenous events. To counteract these stressors, the DNA damage response network contributes to the prevention and/or repair of genomic DNA damage and serves an essential function in cellular survival. DNA binding proteins are involved in this network. Recently, several RNA-binding proteins (RBPs) that are recruited to DNA damage sites have been shown to be direct players in the prevention or repair of DNA damage. In addition, non-coding RNAs, themselves, are involved in the RNA-mediated DNA repair system. Furthermore, RNA modification such as m6A methylation might also contribute to the ultraviolet-responsive DNA damage response. Accumulating evidence suggests that RNA metabolism is more deeply involved in diverse cellular functions than previously expected, and is also intricately associated with the maintenance of genome integrity. In this review, we highlight the roles of RBPs in the maintenance of genome integrity.

Homeodomain-Interacting Protein Kinase-2: A Critical Regulator of the DNA Damage Response and the Epigenome

Homeodomain-interacting protein kinase 2 (HIPK2) is a serine/threonine kinase that phosphorylates and activates the apoptotic program through interaction with diverse downstream targets including tumor suppressor p53. HIPK2 is activated by genotoxic stimuli and modulates cell fate following DNA damage. The DNA damage response (DDR) is triggered by DNA lesions or chromatin alterations. The DDR regulates DNA repair, cell cycle checkpoint activation, and apoptosis to restore genome integrity and cellular homeostasis. Maintenance of the DDR is essential to prevent development of diseases caused by genomic instability, including cancer, defects of development, and neurodegenerative disorders. Recent studies reveal a novel HIPK2-mediated pathway for DDR through interaction with chromatin remodeling factor homeodomain protein 1γ. In this review, we will highlight the molecular mechanisms of HIPK2 and show its functions as a crucial DDR regulator.

Prognostic impact of chronic total coronary occlusion on long-term outcomes in implantable cardioverter-defibrillator recipients with ischaemic heart disease

Aims The prognostic impact of chronic total coronary occlusion (CTO) on implantable cardioverter-defibrillator (ICD) recipients remains unclear. Methods and results Eighty-four consecutive patients with ischaemic heart disease who received ICD therapy for primary or secondary prevention were analysed. We investigated all-cause mortality and major adverse cardiac events (MACEs) including cardiac death, appropriate device therapy, hospitalization for heart failure, and ventricular assist device implantation. Of the study patients (mean age 70 ± 8 years; 86% men), 34 (40%) had CTO. There were no significant differences in age, left ventricular ejection fraction (LVEF), New York Heart Association functional class III or IV status, and proportion who underwent secondary prevention between patients with CTO (CTO group) and without CTO (non-CTO group). During a median follow-up of 3.8 years (interquartile range 2.7-5.4 years), the CTO group tended to have a higher MACE rate (log-rank P = 0.054) than the non-CTO group. Within the CTO group, there was no difference in the MACE rate between patients with and without viable myocardium. In patients with ICD for secondary prevention (n = 47), 16 patients (34%) with CTO had a higher MACE rate than patients without CTO (log-rank P < 0.01). Cox proportional hazards regression analysis showed that the presence of CTO, but not LVEF, was associated with a higher MACE rate. Multivariate analysis showed that the presence of CTO was a predictor of MACE (P < 0.05). Conclusion In patients with ischaemic heart disease receiving ICD implantation, the presence of CTO has an adverse impact on long-term prognosis, especially as secondary prevention.

Negative perception of socioeconomic status with depressive mood down-regulates expression of PPBP and SLC1A7 genes in peripheral blood leukocytes

Abstract Inequality in socioeconomic status (SES) is associated with an increased risk for the development of mental health problems. Here, we examined the association between socioeconomic status (SSS) and psychological distress, and measured gene expression signatures in peripheral blood leukocytes responsible for this association, in 129 healthy adults (27 males and 102 females, aged 44.0 ± 13.0 years) working in a private hospital in Japan. Depressive mood was assessed by Zung Self-rating Depression Scale (SDS). A multiple regression analysis adjusted for gender and age showed that subjective SSS was independently and negatively associated with SDS score. We next focused on 9 subjects who exhibited low SSS scores and 11 subjects with high SSS scores. Microarray analysis revealed that levels of 522 mRNAs were differentially expressed in periheral leukocytes between low and high-SSS groups. The differentially expressed genes were preferentially involved in cellular movement or inflammatory responses. Among them, mRNA levels of pro-platelet basic protein (PPBP) and solute carrier family 1 (glutamine transporter), member 7 (SLC1A7) were negatively correlated with SSS scores. Our results re-confirmed the association between negative perception of SES and depressive mood in healthy adults, and suggest a possible involvement of PPBP and SLC1A7 in the association.

Serine/arginine-rich splicing factor 7 regulates p21-dependent growth arrest in colon cancer cells

Serine/arginine-rich splicing factors (SRSFs) play wide-ranging roles in gene expression through post-transcriptional regulation as well as pre-mRNA splicing. SRSF7 was highly expressed in colon cancer tissues, and its knockdown inhibited cell growth in colon cancer cells (HCT116) in association with altered expression of 4,499 genes. The Ingenuity Pathway Analysis revealed that cell cycle-related canonical pathways were ranked as the highly enriched category in the affected genes. Western blotting confirmed that p21, a master regulator in cell cycle, was increased without any induction of p53 in SRSF7 knockdown cells. Furthermore, cyclin-dependent kinase 2 and retinoblastoma protein were remained in the hypophosphorylated state. In addition, the SRSF7 knockdown-induced cell growth inhibition was observed in p53-null HCT116 cells, suggesting that p53-independent pathways were involved in the SRSF7 knockdown-induced cell growth inhibition. The reduction of SRSF7 stabilized cyclin-dependent kinase inhibitor 1A (CDKN1A) mRNA without any activation of the CDKN1A promoter. Interestingly, SRSF7 knockdown also blocked p21 degradation. These results suggest that the reduction of SRSF7 post-transcriptionally regulates p21 induction at the multistep processes. Thus, the present findings disclose a novel, important role of SRSF7 in cell proliferation through regulating p21 levels. J. Med. Invest. 63: 219-226, August, 2016.

Geranylgeranylacetone prevents stress‐induced decline of leptin secretion in mice

Geranylgeranylacetone (GGA) is a chaperon inducer that protects various types of cell and tissue against stress. We examined whether GGA modulated energy intake and expenditure under stressful conditions. After mice were untreated or treated orally with GGA (0.16 g per kg body weight per day) for 10 days, they were subjected to 2-h restraint stress once or once a day for 5 consecutive days. GGA administration did not affect corticosterone response to the stress. Restraint stress rapidly decreased plasma leptin levels in control mice. GGA significantly increased circulating leptin levels without changing food intake and prevented the stress-induced decline of circulating leptin. However GGA-treated mice significantly reduced food intake during the repeated stress, compared with control mice. GGA prevented the stress-induced decline of leptin mRNA and its protein levels in epidydimal adipose tissues. We also found that GGA decreased ghrelin mRNA expression in gastric mucosa before the stress, whereas GGA-treated mice recovered the ghrelin mRNA expression to the baseline level after the repeated stress. Leptin and ghrelin are now recognized as regulators of anxiety and depressive mood. Our results suggest that GGA may regulate food intake and relief stress-induced mood disturbance through regulating leptin and ghrelin secretions. J. Med. Invest. 65:103-109, February, 2018.

Nucleolin facilitates nuclear retention of an ultraconserved region containing TRA2β4 and accelerates colon cancer cell growth

Transcribed-ultraconserved regions (T-UCRs), which contain conserved sequences with 100% identity across human, rat and mouse species, are a novel category of functional RNAs. The human transformer 2β gene (TRA2B) encodes a UCR that spans exon 2 (276 bp) and its neighboring introns. Among five spliced RNA variants (TRA2β1-5) transcribed from the TRA2B gene, only TRA2β4 contains the conserved exon 2. TRA2β4 is overexpressed in colon cancer cells and accelerates cell growth by blocking the transcription of CDKN1A. However, the mechanisms underlying the overexpression of TRA2β4 in colon cancer cells are unknown. Using biotinylated RNA pull-down assays followed by liquid chromatography-mass spectrometric analysis, we identified nucleolin as a TRA2β4-binding protein. Knockdown of nucleolin reduced the nuclear retention of TRA2β4 and accelerated its degradation in the cytoplasm, whereas nucleolin overexpression increased TRA2β4 levels and its mitogenic activity. Nucleolin directly bound to TRA2β4 exon 2 via the glycine/arginine-rich (GAR) domain. Overexpression of GAR-deficient nucleolin failed to increase TRA2β4 expression and growth of colon cancer cells. RNA fluorescence in situ hybridization showed that TRA2β4 co-localized with nucleolin in nuclei but not with the mutant lacking GAR. Our results suggest that specific interactions between nucleolin and UCR-containing TRA2β4 may be associated with abnormal growth of colon cancer cells.

Adverse parenting is associated with blunted salivary cortisol awakening response and altered expression of glucocorticoid receptor β and β2-adrenergic receptor mRNAs in leukocytes in Japanese medical students

Abstract Adverse parenting is associated with an increased risk for the development of mood and behavioral disorders. In this study, we assessed the perceived parental bonding of 232 medical students using the parental bonding instrument (PBI) and extracted 22 students who reported their parents’ rearing attitudes as affectionless control (LOW; low care, high overprotection). Using the 28-item general health questionnaire, the Zung self-rating depression scale (Zung-SDS), the hospital anxiety and depression scale (HADS), and the Spielberger state-trait-anxiety-inventory (STAI), physical and mental state of the LOW students were compared with those of 30 students who reported their parental bonding as optimal (OPT; high care and low overprotection). These questionnaire measurements demonstrated significantly higher anxiety and depressive mood in the LOW students versus the OPT students. Compared with the OPT students, the LOW students also exhibited a significantly reduced salivary cortisol awakening response (CAR) without changes across the rest of the diurnal salivary cortisol profile. Among glucocorticoid-related genes examined (GR, ADRB2, IκBα, IL10, IL1R2, IL1RN, MR, MC2R, TGFB1, TGFB2 and FASLG), real-time reverse transcription-PCR showed that the LOW students significantly increased expression of a dominant negative glucocorticoid receptor β (GRβ) mRNA and decreased β2-adrenergic receptor (ADRB2) mRNA levels in circulating leukocytes. These results suggest that negative perception of parents’ child-rearing attitudes may be associated with anxiety and depressive mood and altered glucocorticoid signaling even in healthy young adults.