Effects of Dicer and Argonaute down-regulation on mRNA levels in human HEK293 cells

RNA interference and the microRNA (miRNA) pathway can induce sequence-specific mRNA degradation and/or translational repression. The human genome encodes hundreds of miRNAs that can post-transcriptionally repress thousands of genes. Using reporter constructs, we observed that degradation of mRNAs bearing sites imperfectly complementary to the endogenous let-7 miRNA is considerably stronger in human HEK293 than HeLa cells. The degradation did not result from the Ago2-mediated endonucleolytic cleavage but it was Dicer- and Ago2-dependent. We used this feature of HEK293 to address the size of a pool of transcripts regulated by RNA silencing in a single cell type. We generated HEK293 cell lines depleted of Dicer or individual Ago proteins. The cell lines were used for microarray analyses to obtain a comprehensive picture of RNA silencing. The 3'-untranslated region sequences of a few hundred transcripts that were commonly up-regulated upon Ago2 and Dicer knock-downs showed a significant enrichment of putative miRNA-binding sites. The up-regulation upon Ago2 and Dicer knock-downs was moderate and we found no evidence, at the mRNA level, for activation of silenced genes. Taken together, our data suggest that, independent of the effect on translation, miRNAs affect levels of a few hundred mRNAs in HEK293 cells

Law of Genome Evolution Direction : Coding Information Quantity Grows

The problem of the directionality of genome evolution is studied. Based on the analysis of C-value paradox and the evolution of genome size we propose that the function-coding information quantity of a genome always grows in the course of evolution through sequence duplication, expansion of code, and gene transfer from outside. The function-coding information quantity of a genome consists of two parts, p-coding information quantity which encodes functional protein and n-coding information quantity which encodes other functional elements except amino acid sequence. The evidences on the evolutionary law about the function-coding information quantity are listed. The needs of function is the motive force for the expansion of coding information quantity and the information quantity expansion is the way to make functional innovation and extension for a species. So, the increase of coding information quantity of a genome is a measure of the acquired new function and it determines the directionality of genome evolution.Comment: 16 page

Transgenerational Stress Memory Is Not a General Response in Arabidopsis

Adverse conditions can trigger DNA damage as well as DNA repair responses in plants. A variety of stress factors are known to stimulate homologous recombination, the most accurate repair pathway, by increasing the concentration of necessary enzymatic components and the frequency of events. This effect has been reported to last into subsequent generations not exposed to the stress. To establish a basis for a genetic analysis of this transgenerational stress memory, a broad range of treatments was tested for quantitative effects on homologous recombination in the progeny. Several Arabidopsis lines, transgenic for well-established recombination traps, were exposed to 10 different physical and chemical stress treatments, and scored for the number of somatic homologous recombination (SHR) events in the treated generation as well as in the two subsequent generations that were not treated. These numbers were related to the expression level of genes involved in homologous recombination and repair. SHR was enhanced after the majority of treatments, confirming previous data and adding new effective stress types, especially interference with chromatin. Compounds that directly modify DNA stimulated SHR to values exceeding previously described induction rates, concomitant with an induction of genes involved in SHR. In spite of the significant stimulation in the stressed generations, the two subsequent non-treated generations only showed a low and stochastic increase in SHR that did not correlate with the degree of stimulation in the parental plants. Transcripts coding for SHR enzymes generally returned to pre-treatment levels in the progeny. Thus, transgenerational effects on SHR frequency are not a general response to abiotic stress in Arabidopsis and may require special conditions

Maternal Genome-Wide DNA Methylation Patterns and Congenital Heart Defects

The majority of congenital heart defects (CHDs) are thought to result from the interaction between multiple genetic, epigenetic, environmental, and lifestyle factors. Epigenetic mechanisms are attractive targets in the study of complex diseases because they may be altered by environmental factors and dietary interventions. We conducted a population based, case-control study of genome-wide maternal DNA methylation to determine if alterations in gene-specific methylation were associated with CHDs. Using the Illumina Infinium Human Methylation27 BeadChip, we assessed maternal gene-specific methylation in over 27,000 CpG sites from DNA isolated from peripheral blood lymphocytes. Our study sample included 180 mothers with non-syndromic CHD-affected pregnancies (cases) and 187 mothers with unaffected pregnancies (controls). Using a multi-factorial statistical model, we observed differential methylation between cases and controls at multiple CpG sites, although no CpG site reached the most stringent level of genome-wide statistical significance. The majority of differentially methylated CpG sites were hypermethylated in cases and located within CpG islands. Gene Set Enrichment Analysis (GSEA) revealed that the genes of interest were enriched in multiple biological processes involved in fetal development. Associations with canonical pathways previously shown to be involved in fetal organogenesis were also observed. We present preliminary evidence that alterations in maternal DNA methylation may be associated with CHDs. Our results suggest that further studies involving maternal epigenetic patterns and CHDs are warranted. Multiple candidate processes and pathways for future study have been identified

At-risk individuals display altered brain activity following stress

Stress is a major risk factor for almost all psychiatric disorders, however, the underlying neurobiological mechanisms remain largely elusive. In healthy individuals, a successful stress response involves an adequate neuronal adaptation to a changing environment. This adaptive response may be dysfunctional in vulnerable individuals, potentially contributing to the development of psychopathology. In the current study, we investigated brain responses to emotional stimuli following stress in healthy controls and at-risk individuals. An fMRI study was conducted in healthy male controls (N = 39) and unaffected healthy male siblings of schizophrenia patients (N = 39) who are at increased risk for the development of a broad range of psychiatric disorders. Brain responses to pictures from the International Affective Picture System (IAPS) were measured 33 min after exposure to stress induced by the validated trier social stress test (TSST) or a control condition. Stress-induced levels of cortisol, alpha-amylase, and subjective stress were comparable in both groups. Yet, stress differentially affected brain responses of schizophrenia siblings versus controls. Specifically, control subjects, but not schizophrenia siblings, showed reduced brain activity in key nodes of the default mode network (PCC/precuneus and mPFC) and salience network (anterior insula) as well as the STG, MTG, MCC, vlPFC, precentral gyrus, and cerebellar vermis in response to all pictures following stress. These results indicate that even in the absence of a psychiatric disorder, at-risk individuals display abnormal functional activation following stress, which in turn may increase their vulnerability and risk for adverse outcomes

Improving metabolic health in obese male mice via diet and exercise restores embryo development and fetal growth

Paternal obesity is now clearly associated with or causal of impaired embryo and fetal development and reduced pregnancy rates in humans and rodents. This appears to be a result of reduced blastocyst potential. Whether these adverse embryo and fetal outcomes can be ameliorated by interventions to reduce paternal obesity has not been established. Here, male mice fed a high fat diet (HFD) to induce obesity were used, to determine if early embryo and fetal development is improved by interventions of diet (CD) and/or exercise to reduce adiposity and improve metabolism. Exercise and to a lesser extent CD in obese males improved embryo development rates, with increased cell to cell contacts in the compacting embryo measured by E-cadherin in exercise interventions and subsequently, increased blastocyst trophectoderm (TE), inner cell mass (ICM) and epiblast cell numbers. Implantation rates and fetal development from resulting blastocysts were also improved by exercise in obese males. Additionally, all interventions to obese males increased fetal weight, with CD alone and exercise alone, also increasing fetal crown-rump length. Measures of embryo and fetal development correlated with paternal measures of glycaemia, insulin action and serum lipids regardless of paternal adiposity or intervention, suggesting a link between paternal metabolic health and subsequent embryo and fetal development. This is the first study to show that improvements to metabolic health of obese males through diet and exercise can improve embryo and fetal development, suggesting such interventions are likely to improve offspring health.Nicole O. McPherson, Hassan W. Bakos, Julie A. Owens, Brian P. Setchell, Michelle Lan

microRNAome changes in bystander three-dimensional human tissue models suggest priming of apoptotic pathways Carcinogenesis Advance

ABSTRACT The radiation-induced bystander effect (RIBE) is a phenomenon whereby unexposed cells exhibit molecular symptoms of stress exposure when adjacent or nearby cells are traversed by ionizing radiation (IR). Recent data suggest that RIBE may be epigenetically mediated by microRNAs (miRNAs), which are small regulatory molecules that target mRNA transcripts for translational inhibition. Here we analyzed microRNAome changes in bystander tissues after α-particle microbeam irradiation of three-dimensional artificial human tissues using microRNA microarrays. Our results indicate that IR leads to a deregulation of miRNA expression in bystander tissues. We report that major bystander endpoints, including apoptosis, cell cycle deregulation, and DNA hypomethylation, may be mediated by altered expression of miRNAs. Specifically, c-MYC-mediated upregulation of the miR-17 family was associated with decreased levels of E2F1 and RB1, suggesting a switch to a proliferative state in bystander tissues, while priming these cells for impending death signals. Upregulation of the miR-29 family resulted in decreased levels of its targets DNMT3a and MCL1, consequently affecting DNA methylation and apoptosis. Altered expression of miR-16 led to changes in expression of BCL2, suggesting modulation of apoptosis. Thus, our data clearly show that miRNAs play a profound role in the manifestation of late RIBE endpoints. In summary, this study creates a roadmap for understanding the role of microRNAome in RIBE and for developing of novel RIBE biomarkers.