Effects of Chromatin Structure Modifiers on the trans-Acting Heterochromatin Position Effect in Drosophila melanogaster.

The heterochromatin position effect is manifested in the inactivation of euchromatin genes transferred to heterochromatin. In chromosomal rearrangements, genes located near the new eu-heterochromatin boundary in the rearrangement (cis-inactivation) and, in rare cases, genes of a region of the normal chromosome homologous to the region of the eu-heterochromatin boundary of the chromosome with the rearrangement (trans-inactivation) are subject to inactivation. The In(2)A4 inversion is able to trans-inactivate the UAS-eGFP reporter gene located on the normal chromosome. We knockdown a number of chromatin proteins using temperature-controlled RNA interference and investigated the effect of knockdown on trans-inactivation of the reporter. We found suppression of trans-inactivation by knockdowns of Su(var)2-HP2, a protein that binds to the key heterochromatin protein HP1a, SAYP, a subunit of the chromatin remodelling complex, and Eggless histone methyltransferase (SETDB1), which introduces a H3K9me3 histone mark, recognized by the HP1a protein. The method of studying the effects of gene knockdown on heterochromatin position effects presented in this work is of independent methodological interest.

High Level of Gene Transcription at the Embryonic Stage Leads to the Suppression of Heterochromatic Trans-Inactivation in Drosophila melanogaster Adults.

In some cases, gene transfer from euchromatin to constitutive heterochromatin as a result of chromosomal rearrangement is accompanied by epigenetic inactivation of this gene (cis-inactivation). In the case of trans-inactivation, transgenes in the normal chromosome are repressed by the cis-inactivation-causing rearranged homologous chromosome. Trans-inactivation is a result of the somatic pairing of homologs and the transfer of the normal chromosomal segment to the heterochromatic compartment of the nucleus. Previously, we have shown that the degree of trans-inactivation of the UAS-eGFP reporter gene in adult flies depends on its transcription level that can be regulated by temperature using the GAL4 transcription activator and its temperature-sensitive inhibitor GAL80ts. In this paper, we investigated the epigenetic inheritance of the active/repressed state of the trans-inactivated reporter gene at different expression levels by measuring eGFP fluorescence in the individual cells of Malpighian tubules in adult flies. High expression levels at the embryonic stage protected the eGFP gene from trans-inactivation in adult flies. The activated state was inherited over the entire period of development and differentiation, while the activating effect of GAL4 was turned off.

Characteristics of miniature pulsed penning ion source: Experiment and PIC simulation.

In the present work, the results of the experimental and particle in cell (PIC) simulation studies of the discharge combustion modes in a miniature Penning ion source (PIS) under the pulse-periodic power supply conditions are presented. Dynamics of discharge ignition and discharge operation mode at a pulsed anode voltage supply are investigated for different values of anode voltage and gas pressure in various magnetic field configurations. Typical examples of current pulse waveforms are shown. Also, numerical simulations of the PIS were performed using 3D PIC combined with Monte Carlo collisions in the code VSim. Temporal dependencies of electron, ion, and potential distributions in the Penning cell are simulated. Differences between the numerical and experimental results are discussed.

Regulated Gene Expression as a Tool for Analysis of Heterochromatin Position Effect in Drosophila.

Position effect variegation (PEV) is a perturbation of genes expression resulting from the changes in their chromatin organization due to the abnormal juxtaposition with heterochromatin. The exact molecular mechanisms of PEV remain enigmatic in spite of the long history of PEV studies. Here, we developed a genetic model consisting of PEV-inducing chromosome rearrangement and a reporter gene under control of the UAS regulatory element. Expression of the reporter gene could be regulated by adjustment of the GAL4 transactivator activity. Two UAS-based systems of expression control were tested - with thermosensitive GAL4 repressor GAL80ts and GAL4-based artificial transactivator GeneSwitch. Both systems were able to regulate the expression of the UAS-controlled reporter gene over a wide range, but GAL80ts repressed the reporter gene more efficiently. Measurements of the heterochromatin-mediated repression of the reporter gene in the GAL4+GAL80ts system point to the existence of a threshold level of expression, above which no PEV is observed.