[Molecular mechanisms of regulaion of transcription by PARP1].

Poly-ADP-ribosylation is a covalent post-translational modification of nuclear proteins that plays a key role in the immediate response of cells to genotoxic stress. Poly(ADP-ribose) polymerase (PARP) synthesizes long and branched polymers of ADP-ribose onto acceptor regulator proteins, and thereby change their activity. Metabolism of poly-ADP regulates DNA repair, cell cycle, replication, aging and death of cells, as well as remodeling of chromatin structure and gene transcription. PARP1 is one of the most common nuclear proteins; it is responsible for production of -90% of the polymers of ADP-ribose in the cell. PARP1 inhibitors are promising antitumor agents. At the same time, the current inhibitors target the catalytic domain of PARP1 that leads to.a number of side effects. Therefore, considering the potential benefits of PARP1 inhibitors for the treatment of multiple diseases, it is necessary to develop new strategies of PARP1 inhibition. PARP1 has a modular structure and has catalytic, transcription and DNA-binding activities. The review focuses primarily on the role of PARP1 in transcriptional regulation; the structure and functional organization of PARP1, as well as multiple ways of regulation of chromatin remodeling, DNA methylation and transcription are covered in detail. Studies of the molecular mechanisms of regulation of transcription factor PARP1 can serve as a basis for search and design of new inhibitors.

[Molecular mechanisms of transcription through a nuclesome by RNA polymerase II].

The Pol II-type mechanism is conserved from yeast to human. After initiation of transcription, Pol II can be paused within the early transcribed region of a gene. Then Pol II overcomes the initial nucleosomal barrier, and efficiently proceeds through chromatin. At low- to moderate-level transcription progression of Pol II is characterized by displacement/exchange of only H2A/H2B dimer(s) and hexasome survival, likely mediated through formation of small intranucleosomal DNA loops. This mechanism helps to preserve the "histone" code during transcription. As the transcription rate is increased, the distance between transcribing Pol II complexes becomes shorter, and trailing Pol II complexes may encounter the hexasome formed after previous transcription round, before the H2A/H2B dimer re-binds to the hexasome. In this case an unstable intermediate with a smaller number of DNA-histone contacts is formed, resulting in eviction of the histone hexamer from DNA in vitro; therefore here all core histones are evicted/exchanged in vivo. Various protein factors and histone chaperones are involved in chromatin transcription by Pol II in vivo.

[Mechanisms of distant enhancer action on DNA and in chromatin].

Enhancers and insulators are regulatory DNA sequences that can work over a large distance. Efficient action over a distance clearly requires special mechanisms for facilitating communication between a regulatory region and its target. Studies from our laboratory identified DNA supercoiling as primary factor that mediates efficient enhancer-promoter communication over a distance in prokaryotes through a "DNA slithering" mechanism. These studies allowed rational design and construction of an insulator that can block enhancer action over a distance both in vitro and in vivo. Our most recent studies suggest that eukaryotic chromatin structure can support action over a distance using similar principles, but in a mechanistically distinct way.

[Chromatin remodeling by RNA polymerase II].

Tight compaction of eukaryotic DNA in chromatin having several levels of organization results in formation of a structure that is barely accessible to regulatory protein complexes and enzymes such as DNA-and RNA-polymerases. To deal with this intricate chromatin organization, numerous chromatin remodeling mechanisms operating at various levels of chromatin structure were developed. Recently it has become apparent that numerous intranuclear processes of DNA metabolism (such as transcription, repair etc.) are often regulated at the level of chromatin remodeling. The mechanisms of chromatin remodeling and its regulation are beginning to emerge. Thus the histone octamer has a remarkable ability to survive action of various processive enzymes, such as DNA- and RNA polymerases, as well as ATP-dependent remodelers. Here we discuss the mechanisms of chromatin remodeling during transcription with primary focus on chromatin remodeling by transcribing Pol II.

[Chromatin transcription]. / Transkriptsiia khromatina.

Eukaryotic genome is organized in DNA--histone complexes called chromatin. The nucleosome core (approximately 150 bp DNA tightly wrapped around the histone octamer in two superhelical coils) is a minimal unit of chromatin organization. Chromatin is designed to allow both tight DNA packaging and proper functioning and regulation of enzymes such as DNA- and RNA-polymerases. The mechanisms allowing DNA to be accessible to enzymes of DNA metabolism in regulated manner are beginning to emerge. Thus the histone octamer has a remarkable ability to survive transcription by stepping around the transcribing polymerase without ever going free in solution. Moreover, nucleosomes can present a high barrier for transcribing polymerases that dramatically decreases the rate of transcription and probably is regulated in vivo. The mechanism of transcription through nucleosomal barrier will be discussed in detail below.

[Study of the structure of Escherichia coli RNA polymerase and its complex with the lacUV5-promotor using protein-protein and DNA-protein crosslinks, formed by formaldehyde]. / Issledovanie struktury RNK-polimerazy Escherichia coli i ee komplekxa s lacUV5-promotorom s pomoshch'iu belok-belkovykh i DNK-belkovykh sshivok, obrazuemykh formal'degidom.

The protein-protein and DNA-protein crosslinks produced by formaldehyde were used to investigate the intersubunit and subunit-DNA interactions for free RNA polymerase and for an open complex of RNA polymerase with the lacUV5 promoter. In both cases the contacts between beta,beta' and beta', sigma subunits were observed, while there were no contacts between beta and sigma subunits. Only one of beta or beta' subunits and a sigma subunit crosslink to promoter DNA. We have chosen the conditions for fixing the RNA polymerase-DNA complexes on different stages of transcription initiation. The possibility to use limited fixation with low concentrations of formaldehyde to study specific DNA-protein interactions was shown.

[Enhancers, DNA loops and stable complexes: a mechanism of transcription activation]. / Enkhansery, petli DNK i stabil'nye kompleksy: mekhanizm aktivatsii transkriptsii.

A recurrent theme in molecular biology is that of "action at a distance" along DNA. We consider various mechanisms of action of transcriptional enhancers as a well-characterised example of interaction between widely separated DNA-bound proteins. The role of promoter- and enhancer-binding proteins in the formation of a stable promoter complex, and the mechanisms of interaction between multiple activatory DNA sequences is also discussed.

[The structure of nucleosomal core particles located on the transcribed genome regions]. / Struktura minimal'nykh nukleosom, raspolozhennykh na transkribiruemykh uchastkakh genoma.

The arrangement of histones along nucleosomal DNA within particular active and inactive genome regions was analysed by using protein-DNA crosslinking methods combined with hybridization tests. Two-dimensional gel electrophoresis was employed to compare the nucleosome conformation and nucleosomal DNA size. The arrangement of histones along DNA and general compactness of nucleosomes were shown to be very similar in transcriptionally active and inactive genomic regions. On the other hand, nucleosomes within transcriptionally active chromatin are characterized by a somewhat larger size of nucleosomal DNA produced by micrococcal nuclease digestion and some peculiarity in electrophoretic mobility. It appears that changes in transcribed chromatin structure (such as an enhanced nuclease sensitivity) occur on the supranucleosomal level rather than in the nucleosomal core structure.