[Restraint Stress-Induced Expression of Fos and Several Related Genes in the Hypothalamus of Hypertensive ISIAH Rats].

Stress can play a significant role in arterial hypertension and many other complications of cardiovascular diseases. Considerable attention is paid to the study of the molecular mechanisms involved in the body response to stressful influences, but there are still many blank spots in understanding the details. ISIAH rats model the stress-sensitive form of arterial hypertension. ISIAH rats are characterized by genetically determined enhanced activities of the hypothalamic-pituitary-adrenocortical and sympathetic-adrenomedullary systems, suggesting a functional state of increased stress reactivity. For the first time, the temporal expression patterns of Fos and several related genes were studied in the hypothalamus of adult male hypertensive ISIAH rats after a single exposure to restraint stress for 30, 60, or 120 min. Fos transcription was activated and peaked 1 h after the start of restraint stress. The time course of Fos activation coincided with that of blood pressure increase after stress. Activation of hypothalamic neurons also alters the transcription levels of several transcription factor genes (Jun, Nr4a3, Jdp2, and Ppargc1a), which are associated with the development of cardiovascular diseases. Because Fos induction is a marker of brain neuron activation, activation of hypothalamic neurons and an increase in blood pressure were concluded to accompany increased stress reactivity of the hypothalamic-pituitary-adrenocortical and sympathoadrenal systems in hypertensive ISIAH rats during short-term restraint.

RatDEGdb: a knowledge base of differentially expressed genes in the rat as a model object in biomedical research.

The animal models used in biomedical research cover virtually every human disease. RatDEGdb, a knowledge base of the differentially expressed genes (DEGs) of the rat as a model object in biomedical research is a collection of published data on gene expression in rat strains simulating arterial hypertension, age-related diseases, psychopathological conditions and other human afflictions. The current release contains information on 25,101 DEGs representing 14,320 unique rat genes that change transcription levels in 21 tissues of 10 genetic rat strains used as models of 11 human diseases based on 45 original scientific papers. RatDEGdb is novel in that, unlike any other biomedical database, it offers the manually curated annotations of DEGs in model rats with the use of independent clinical data on equal changes in the expression of homologous genes revealed in people with pathologies. The rat DEGs put in RatDEGdb were annotated with equal changes in the expression of their human homologs in affected people. In its current release, RatDEGdb contains 94,873 such annotations for 321 human genes in 836 diseases based on 959 original scientific papers found in the current PubMed. RatDEGdb may be interesting first of all to human geneticists, molecular biologists, clinical physicians, genetic advisors as well as experts in biopharmaceutics, bioinformatics and personalized genomics. RatDEGdb is publicly available at https://www.sysbio.ru/RatDEGdb.

InterTransViewer: a comparative description of differential gene expression profiles from different experiments.

Meta-analysis of transcriptomic data from different experiments has become increasingly prevalent due to a significantly increasing number of genome-wide experiments investigating gene expression changes under various conditions. Such data integration provides greater accuracy in identifying candidate genes and allows testing new hypotheses, which could not be validated in individual studies. To increase the relevance of experiment integration, it is necessary to optimize the selection of experiments. In this paper, we propose a set of quantitative indicators for a comprehensive comparative description of transcriptomic data. These indicators can be easily visualized and interpreted. They include the number of differentially expressed genes (DEGs), the proportion of experiment-specific (unique) DEGs in each data set, the pairwise similarity of experiments in DEG composition and the homogeneity of DEG profiles. For automatic calculation and visualization of these indicators, we have developed the program InterTransViewer. We have used InterTransViewer to comparatively describe 23 auxin- and 16 ethylene- or 1-aminocyclopropane-1-carboxylic acid (ACC)-induced transcriptomes in Arabidopsis thaliana L. We have demonstrated that analysis of the characteristics of individual DEG profiles and their pairwise comparisons based on DEG composition allow the user to rank experiments in the context of each other, assess the tendency towards their integration or segregation, and generate hypotheses about the influence of non-target factors on the transcriptional response. As a result, InterTransViewer identifies potentially homogeneous groups of experiments. Subsequent estimation of the profile homogeneity within these groups using resampling and setting a significance threshold helps to decide whether these data are appropriate for meta-analysis. Overall, InterTransViewer makes it possible to efficiently select experiments for meta-analysis depending on its task and methods.

A bioinformatic search for correspondence between differentially expressed genes of domestic versus wild animals and orthologous human genes altering reproductive potential.

One of the greatest achievements of genetics in the 20th century is D.K. Belyaev's discovery of destabilizing selection during the domestication of animals and that this selection affects only gene expression regulation (not gene structure) and inf luences systems of neuroendocrine control of ontogenesis in a stressful environment. Among the experimental data generalized by Belyaev's discovery, there are also f indings about accelerated extinc tion of testes' hormonal function and disrupted seasonality of reproduction of domesticated foxes in comparison with their wild congeners. To date, Belyaev's discovery has already been repeatedly conf irmed, for example, by independent observations during deer domestication, during the use of rats as laboratory animals, after the reintroduction of endangered species such as Przewalski's horse, and during the creation of a Siberian reserve population of the Siberian grouse when it had reached an endangered status in natural habitats. A genome-wide comparison among humans, several domestic animals, and some of their wild congeners has given rise to the concept of self-domestication syndrome, which includes autism spectrum disorders. In our previous study, we created a bioinformatic model of human self-domestication syndrome using differentially expressed genes (DEGs; of domestic animals versus their wild congeners) orthologous to the human genes (mainly, nervous-system genes) whose changes in expression affect reproductive potential, i. e., growth of the number of humans in the absence of restrictions caused by limiting factors. Here, we applied this model to 68 human genes whose changes in expression alter the reproductive health of women and men and to 3080 DEGs of domestic versus wild animals. As a result, in domestic animals, we identif ied 16 and 4 DEGs, the expression changes of which are codirected with changes in the expression of the human orthologous genes decreasing and increasing human reproductive potential, respectively. The wild animals had 9 and 11 such DEGs, respectively. This difference between domestic and wild animals was signif icant according to Pearson's χ2 test (p < 0.05) and Fisher's exact test (p < 0.05). We discuss the results from the standpoint of restoration of endangered animal species whose natural habitats are subject to an anthropogenic impact.

Migration of primordial germline cells is negatively regulated by surrounding somatic cells during early embryogenesis in Drosophila melanogaster.

Cell migration is an important morphogenetic process necessary at different stages of individual development and body functioning. The initiation and maintenance of the cell movement state requires the activation of many factors involved in the regulation of transcription, signal transduction, adhesive interactions, modulation of membranes and the cytoskeleton. However, cell movement depends on the status of both migrating and surrounding cells, interacting with each other during movement. The surrounding cells or cell matrix not only form a substrate for movement, but can also participate in the spatio-temporal regulation of the migration. At present, there is no exact understanding of the genetic mechanisms of this regulation. To determine the role of the cell environment in the regulation of individual cell migration, we studied the migration of primordial germline cells (PGC) during early embryogenesis in Drosophila melanogaster. Normally, PGC are formed at the 3rd stage of embryogenesis at the posterior pole of the embryo. During gastrulation (stages 6-7), PGC as a consolidated cell group passively transfers into the midgut primordium. Further, PGC are individualized, acquire an amoeboid form, and actively move through the midgut epithelium and migrate to the 5-6 abdominal segment of the embryo, where they form paired embryonic gonads. We screened for genes expressed in the epithelium surrounding PGC during early embryogenesis and affecting their migration. We identified the myc, Hph, stat92E, Tre-1, and hop genes, whose RNA interference leads to premature active PGC migration at stages 4-7 of embryogenesis. These genes can be divided into two groups: 1) modulators of JAK/STAT pathway activity inducing PGC migration (stat92E, Tre-1, hop), and 2) myc and Hph involved in epithelial morphogenesis and polarization, i. e. modifying the permeability of the epithelial barrier. Since a depletion of each of these gene products resulted in premature PGC migration, we can conclude that, normally, the somatic environment negatively regulates PGC migration during early Drosophila embryogenesis.

Hidden heterogeneity of transcription factor binding sites: A case study of SF-1.

Steroidogenic factor 1 (SF-1) belongs to a small group of the transcription factors that bind DNA only as a monomer. Three different approaches-Sitecon, SiteGA, and oPWM-constructed using the same training sample of experimentally confirmed SF-1 binding sites have been used to recognize these sites. The appropriate prediction thresholds for recognition models have been selected. Namely, the thresholds concordant by false positive or negative rates for various methods were used to optimize the discrimination of steroidogenic gene promoters from the datasets of non-specific promoters. After experimental verification, the models were used to analyze the ChIP-seq data for SF-1. It has been shown that the sets of sites recognized by different models overlap only partially and that an integration of these models allows for identification of SF-1 sites in up to 80% of the ChIP-seq loci. The structures of the sites detected using the three recognition models in the ChIP-seq peaks falling within the [-5000, +5000] region relative to the transcription start sites (TSS) extracted from the FANTOM5 project have been analyzed. The MATLIGN classified the frequency matrices for the sites predicted by oPWM, Sitecon, and SiteGA into two groups. The first group is described by oPWM/Sitecon and the second, by SiteGA. Gene ontology (GO) analysis has been used to clarify the differences between the sets of genes carrying different variants of SF-1 binding sites. Although this analysis in general revealed a considerable overlap in GO terms for the genes carrying the binding sites predicted by oPWM, Sitecon, or SiteGA, only the last method elicited notable trend to terms related to negative regulation and apoptosis. The results suggest that the SF-1 binding sites are different in both their structure and the functional annotation of the set of target genes correspond to the predictions by oPWM+Sitecon and SiteGA. Further application of Homer software for de novo identification of enriched motifs in ChIP-Seq data for SF-1ChIP-seq dataset gave the data similar to oPWM+Sitecon.

Analysis and recognition of the GAGA transcription factor binding sites in Drosophila genes.

The transcription factor GAGA, encoded by the gene Trl, controls expression of many Drosophila melanogaster genes. We have compiled the presently largest sample (120 sites) of published nucleotide sequences with experimentally confirmed binding to GAGA protein. Analysis of the sample has demonstrated that despite an apparent structural diversity of the GAGA sites, they fall into four distinct groups, namely, (1) the sites containing two GAG trinucleotides with no more than one nucleotide substitution in each and separated by spacers with a length of 1 or 3 nucleotides (GAGnGAG and GAGnnnGAG); (2) the sites containing a single GAGAG motif; (3) (GA)(3-9) microsatellite repeats; and (4) the sites corresponding to three and more direct repeats of GAG trinucleotide homolog and its inverted repeats separated by spacers of various lengths. Using the software package SITECON, the methods were elaborated for recognizing the sites of GAGnGAG (method 1) and GAGnnnGAG (method 2) types in DNA sequences. Experimental verification confirmed the ability to interact with the GAGA factor for 72% of the sites predicted using method 1 and 94.5% of the sites predicted by method 2. Application of the experimentally verified methods to analyzing the localization of potential GAGA binding sites in the target genes of this transcription factor has demonstrated that the 5'-untranslated regions (5'UTRs) and first introns are enriched for these sites (two-threefold relative to the average occurrence frequency in the D. melanogaster genome) as compared with a moderate enrichment (not exceeding 1.5-fold) of promoter regions (-4000/+200 bp or -1000/+100 bp).

Detection of target genes of FOXA transcription factors involved in proliferation control.

To reveal the mechanism of tumor-suppressing activity of FOXA proteins in liver, a search for potential target genes of these transcription factors involved in proliferation control was carried out. In the first step, we have used data from the literature concerning gene expression in mouse liver (high content of FOXA proteins) and kidney (FOXA expression is absent) obtained by hybridization on microchips. A search for FOXA binding sites in regulatory regions of forty differentially expressing genes involved in proliferation control was carried out using the computer method SITECON. Eleven genes containing clusters of potential FOXA sites incorporating 3-6-fold repeats of TTTG were revealed. The FOXA-specific interaction with such microsatellite sites was confirmed by gel-retardation technique using the GST-fused protein containing the DNA-binding domain of FOXA2. Six genes containing clusters of confirmed binding sites--Cul2, Cdc73, Ptk, Pdcd, Creb, and Ppp2r5d--were selected. The effect of hepatocarcinogen orthoaminoazotoluene (OAT), which lowers the FOXA activity, on expression of these genes was studied by the real-time PCR. OAT was shown to increase sharply the level of mRNA of the Cul2 and Cdc73 genes.

Bioinformatical and experimental approaches to investigation of transcription factor binding sites in vertebrate genes.

The development of computer-assisted methods for transcription factor binding sites (TFBS) recognition is necessary for study the DNA regulatory transcription code. There are a great number of experimental methods that enable TFBS identification in genome sequences. The experimental data can be used to elaborate multiple computer approaches to recognition of TFBS, each of which has its own advantages and limitations. A short review of the characteristics of computer methods of TFBS prediction based on various principles is presented. Methods used for experimental monitoring of predicted sites are analyzed. Data concerning DNA regulatory potential and its realization at the chromatin level, obtained using these methods, are discussed along with approaches to recognition of target genes of certain transcription factors in the genome sequences.