Control of homologous recombination by the HROB-MCM8-MCM9 pathway.

DNA repair by homologous recombination (HR) is essential for genomic integrity, tumor suppression, and the formation of gametes. HR uses DNA synthesis to repair lesions such as DNA double-strand breaks and stalled DNA replication forks, but despite having a good understanding of the steps leading to homology search and strand invasion, we know much less of the mechanisms that establish recombination-associated DNA polymerization. Here, we report that C17orf53/HROB is an OB-fold-containing factor involved in HR that acts by recruiting the MCM8-MCM9 helicase to sites of DNA damage to promote DNA synthesis. Mice with targeted mutations in Hrob are infertile due to depletion of germ cells and display phenotypes consistent with a prophase I meiotic arrest. The HROB-MCM8-MCM9 pathway acts redundantly with the HELQ helicase, and cells lacking both HROB and HELQ have severely impaired HR, suggesting that they underpin two major routes for the completion of HR downstream from RAD51. The function of HROB in HR is reminiscent of that of gp59, which acts as the replicative helicase loader during bacteriophage T4 recombination-dependent DNA replication. We therefore propose that the loading of MCM8-MCM9 by HROB may similarly be a key step in the establishment of mammalian recombination-associated DNA synthesis.

Genetic profile characterization and population study of 21 autosomal STR in Chinese Kazak ethnic minority group.

Short tandem repeat loci have been recognized as useful tools in the routine forensic application and in recent decades, more and more new short tandem repeat (STR) loci have been constantly discovered, studied, and applied in forensic caseworks. In this study, we investigated the genetic polymorphisms of 21 STR loci in the Kazak ethnic minority as well as the genetic relationships between the Kazak ethnic minority and other populations. Allelic frequencies of 21 STR loci were obtained from 114 unrelated healthy Kazak individuals in the Ili Kazak Autonomous Prefecture, Xinjiang Uigur Autonomous Region of China. We observed a total of 159 alleles in the group with the allelic diversity values ranging from 0.0044 to 0.5088. The highest polymorphism was found at D19S433 locus and the lowest was found at D1S1627. Statistical analysis of the generated data indicated no deviation from Hardy-Weinberg equilibriums at all 21 STR loci. In order to estimate the population differentiation, allelic frequencies of all STR loci of the Kazak were compared with those of other neighboring populations using analysis of molecular variance method. Statistically significant differences were found between the studied population and other populations at 2-7 STR loci. A neighbor-joining tree was constructed based on allelic frequencies of the 21 STR loci and phylogenetic analysis indicates that the Kazak has a close genetic relationship with the Uigur ethnic group. The present results may provide useful information for forensic sciences and population genetics studies, and can also increase our understanding of the genetic background of this group. The present findings showed that all the 21 STR loci are highly genetically polymorphic in the Kazak group, which provided valuable population genetic data for the genetic information study, forensic human individual identification, and paternity tests.

Eye movement verification and evaluation of initial sandplay picture system for internet addiction symptoms in Chinese adolescents.

BACKGROUND: Sandplay therapy is a psychotherapeutic technique, based on the psychoanalytic theory of the unconscious. Nearly a century after it was developed, sandplay can now be applied for the initial diagnosis tools for sand players. The goal of the current research is to demonstrate the role of sandplay in identifying internet-addicted adolescents in China. The study aims to evaluate the reliability and validity for sandplay as a diagnosis and evaluation tool for internet addiction symptoms, and to verify the consistency that exists between results based on sandplay pictures and those based on the Pathological Internet Usage Scale for Adolescents (APIUS). METHODS: The research was conducted with a 2 × 2 mixed factorial design - two types of participants (addicts and non-addicts) and two types of sandplay pictures (pictures for addicts and pictures for non-addicts). An absolute recognition-judgment paradigm was used along with eye movement evaluations to evaluate the existing initial sandplay picture system for internet addiction symptoms (22 sandplay pictures, 11 related to addicts and 11 related to non-addicts, respectively). Sixty Chinese adolescents were selected as the participants (30 as addicts and 30 as non-addicts) according to the APIUS. RESULTS: (1) The initial sandplay pictures for internet addicts are clearly preferred by Chinese internet-addicted adolescents, which are more familiar and easier to process; (2) Such pictures have a higher level of emotional arousal and cognitive resonance for the addicts; (3) Track and heat maps indicate that young internet addicts mainly fixate on the initial sandplay pictures for internet addicts. CONCLUSION: This initial sandplay picture system can be used to screen and identify young Chinese internet addicts based on symptoms, and the evaluation results are consistent with those based on the APIUS.

Pubertal recalibration of cortisol reactivity following early life parent-child separation.

BACKGROUND: The hypothalamic-pituitary-adrenocortical (HPA) axis had been proved to calibrate to early-life adversity and puberty may reverse the calibration. This study examines the consequences of prolonged parent-child separation on HPA axis reactivity and the pubertal recalibration hypothesis. METHODS: Totally of 144 participants aged 8.75 to 15.25 (mean age 12.50 years, SD: 1.32) were enrolled from rural areas of Chizhou city, Anhui Province of China in 2019. Data on parent-child separation was collected from parents. Self-reported Peterson Pubertal Development Scale was used to assess pubertal maturation and HPA axis stress reactivity was measured using the Trier Social Stress Test for Children. RESULTS: For children at early stage of puberty, childhood parent-child separation experiences were associated with blunted HPA axis reactivity (B = -1.888, p = 0.034); while for those at later stage of puberty, HPA axis reactivity was similar between children experienced early childhood separation and those without separation (AUCi: B = -0.426, p = 0.878). In contrast, for children experienced persistent parent-child separation, blunted HPA axis reactivity was observed (all p < 0.05). LIMITATIONS: Due to the cross-sectional nature of this study, conclusions about causality remain speculative. CONCLUSIONS: The effect of parent-child separation on dysregulation of HPA axis acts in a time-dependent manner. This finding provides support for the pubertal recalibration hypothesis suggesting that a focus of improving environment in adolescence would help those individuals reared initially in non-supportive conditions.

A consensus set of genetic vulnerabilities to ATR inhibition.

The response to DNA replication stress in eukaryotes is under the control of the ataxia-telangiectasia and Rad3-related (ATR) kinase. ATR responds to single-stranded (ss) DNA to stabilize distressed DNA replication forks, modulate DNA replication firing and prevent cells with damaged DNA or incomplete DNA replication from entering into mitosis. Furthermore, inhibitors of ATR are currently in clinical development either as monotherapies or in combination with agents that perturb DNA replication. To gain a genetic view of the cellular pathways requiring ATR kinase function, we mapped genes whose mutation causes hypersensitivity to ATR inhibitors with genome-scale CRISPR/Cas9 screens. We delineate a consensus set of 117 genes enriched in DNA replication, DNA repair and cell cycle regulators that promote survival when ATR kinase activity is suppressed. We validate 14 genes from this set and report genes not previously described to modulate response to ATR inhibitors. In particular we found that the loss of the POLE3/POLE4 proteins, which are DNA polymerase ε accessory subunits, results in marked hypersensitivity to ATR inhibition. We anticipate that this 117-gene set will be useful for the identification of genes involved in the regulation of genome integrity and the characterization of new biological processes involving ATR, and may reveal biomarkers of ATR inhibitor response in the clinic.

[Receptor-related mechanism of proliferation inhibilion and apoptosis induetion of human tongue squamous cell line Tca8113 by retinoids].

OBJECTIVE: To investigate the receptor-related mechanism of retinoids inhibiting proliferation and inducing apoptosis of human oral squamous cell carcinoma cell line Tca8113. METHODS: The effects of 3 retinoids (namely 9-cis-RA, at-RA and 13-cis-RA), TTNPB (RAR agonist) and methoprene acid (Ma, RXR agonist) on proliferation and cell cycle of Tca8113 cells were analyzed by MTT assay and flow cytometry. The roles of these agents in inducing apoptosis of Tca8113 cells were also evaluated by detecting the expression of Bcl-2/Bax, TUNEL and active caspase-3 analysis. RESULTS: Both retinoids and TTNPB could inhibit the proliferation of Tca8113 cells, and the effect of TTNPB was the most powerful in all the reagents, but MA had no such effect. At the concentration of 1 x 10(-5) mol/L, all the agents except for Ma could increase the percentage of G(1)/G(0)-stage cells after incubation of the cells for 24 h and 48 h. Retinoids and TTNPB could up-regulate the expression of Bax and down-regulate Bcl-2 expression. The results of TUNEL demonstrated that retinoids and TTNPB, but not Ma, could induce apoptosis of Tca8113 cells as compared with the control group (P<0.05). Except for Ma, all the agents up-regulated caspase-3 expression, and the effect of TTNPB was the strongest (P<0.05). CONCLUSIONS: Retinoids can suppress the proliferation of and induce apoptosis of Tca8113 cells, the effect of which involves activation of RAR but not RXR. caspase-3 pathway is involved in apoptosis-inducing effects of retinoids.

[Apoptosis induced by curcumin and its effect on c-myc and caspase-3 expressions in human melanoma A375 cell line].

OBJECTIVE: To investigate the effect of curcumin on cell apoptosis and c-myc and caspase-3 expressions in human melanoma A375 cell line. METHODS: A375 cells were exposed to curcumin treatment and growth inhibition of the cells was examined by MTT assay. Annexin V/propidium iodide double staining and DNA fragmentation analysis were employed for assay of the cell apoptosis and morphological changes of the cells were observed with inverted microscopy and transmission electron microscopy, respectively. In situ hybridization and SABC immunohistochemistry were performed for detection of the expressions of c-Myc and caspase-3 in the A375 cells. RESULTS: Curcumin inhibited the growth of A375 cells in both time- and concentration-dependent manners. After treatment with 30 micromol/L curcumin for 48 h, apoptotic morphological changes were observed in the cells and an oligonucleosomal DNA ladder was clearly visualized in DNA fragmentation analysis. The apoptotic rates of the cells treated with curcumin at the concentration above 20 micromol/L were significantly higher than that of the control cells. c-myc expression level was decreased whereas caspase-3 expression increased with the increase in curcumin concentrations. CONCLUSION: Curcumin can inhibit the proliferation and induce apoptosis of A375 cells in vitro, and the genes encoding c-myc and caspase-3 may play a role in the process.

Small-interfering RNA-mediated silencing of the MAPK p42 gene induces dual effects in HeLa cells.

The genesis and progression of cervical cancer involve the mutation or deviant expression of numerous genes, including the activation of oncogenes (Ha-ras, C-myc, C-erbB2 and Bcl-2) and inactivation of tumor-suppressor genes (p53 and Rb). Previous studies showed that small-interfering RNAs (siRNAs) targeting the MAPK p42 gene partly inhibit proliferation and increase apoptosis in human cervical carcinoma HeLa cells. Results of a microarray analysis showed that MAPK p42 siRNA inhibited cell growth through the regulation of cell cycle control and apoptosis and induced interferon-like response in HeLa cells. In order to confirm the dual effects of MAPK p42 siRNA, we compared the roles of siRNA and U0126, an inhibitor of MAPK p42, in HeLa cells. Short 21-mer double-stranded/siRNAs were synthesized to target MAPK p42 mRNA in HeLa cells. The siRNAs were transfected into HeLa cells using Lipofectamine. The cells were treated with siRNA or U0126 at different concentrations for a period of 48 h. The biological effect of siRNA and U0126 on HeLa cells was measured by MTT and flow cytometry. MAPK1, NUP188, P38, STAT1, STAT2, PML and OAS1 were analyzed by real-time quantitative PCR. HeLa cell growth was inhibited by siRNA or U0126, and the effect of siRNA inhibition was greater than that of U0126. Cell cycle phases were different for siRNA or U0126, but HeLa cell growth was arrested at the S phase by siRNA and at G1 phase by U0126. A down-regulation in MAPK p42 expression by siRNA and up-regulation by U0126 were noted. The results of real-time quantitative PCR showed that P38 was up-regulated and NUP188 was down-regulated by siRNA in comparison with the control groups, and the results were consistent with those of U0126. Expression levels of STAT1, STAT2, PML and OAS1 induced by siRNA differed from those induced by U0126. siRNA-mediated silencing and deactivation induced by U0126 in MAPK p42 led to growth inhibition in the HeLa cells. The effects of siRNA on HeLa cell growth were different from those of U0126. Dual effects of MAPK p42 siRNA-2 on HeLa cell growth were noted: one consisted of a specific effect induced by siRNA-mediated p42 MAPK silencing and the other exhibited a non-specific interferon-like response.

[Expression of E-cadherin and beta-catenin in Bowen's disease and squamous cell carcinoma].

OBJECTIVE: To investigate the involvement of E-cadherin-catenin adhesion system in Bowen's disease (BD) and cutaneous squamous cell carcinoma (SCC). METHODS: Fifteen normal skin, 28 BD and 18 SCC specimens were stained with monoclonal antibodies against E-cadherin and beta-catenin. Evaluation of the staining results was performed with semi-quantification of the pattern and intensity of staining, percentage of positive cells, and cytoplasmic staining. RESULT: Normal skins strongly expressed membranous E-cadherin and beta-catenin, but their expression was remarkably reduced in BD and SCC. Abnormal staining of beta-catenin was observed in the cytoplasm or cell nuclei of BD and SCC. CONCLUSION: Abnormal expression of the E-cadherin/catenin complex is common in SCC and BD.