Publications by authors named "Nikolay Zolotarev"

6 Publications

  • Page 1 of 1

A Prion-like Domain in Transcription Factor EBF1 Promotes Phase Separation and Enables B Cell Programming of Progenitor Chromatin.

Immunity 2020 Dec 6;53(6):1151-1167.e6. Epub 2020 Nov 6.

Max Planck Institute of Immunobiology and Epigenetics, 79108 Freiburg, Germany. Electronic address:

Establishment of B-lineage-specific gene expression requires the binding of transcription factors to inaccessible chromatin of progenitors. The transcription factor EBF1 can bind genomic regions prior to the detection of chromatin accessibility in a manner dependent on EBF1's C-terminal domain (CTD) and independent of cooperating transcription factors. Here, we studied the mechanism whereby the CTD enables this pioneering function. The CTD of EBF1 was dispensable for initial chromatin targeting but stabilized occupancy via recruitment of the chromatin remodeler Brg1. We found that the CTD harbors a prion-like domain (PLD) with an ability of liquid-liquid phase separation, which was enhanced by interaction of EBF1 with the RNA-binding protein FUS. Brg1 also partitioned into phase-separated FUS condensates and coincided with EBF1 and FUS foci in pro-B cells. Heterologous PLDs conferred pioneering function on EBF1ΔCTD. Thus, the phase separation ability of EBF1 facilitates Brg1-mediated chromatin opening and the transition of naive progenitor chromatin to B-lineage-committed chromatin.
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http://dx.doi.org/10.1016/j.immuni.2020.10.009DOI Listing
December 2020

Small Drosophila zinc finger C2H2 protein with an N-terminal zinc finger-associated domain demonstrates the architecture functions.

Biochim Biophys Acta Gene Regul Mech 2020 01 6;1863(1):194446. Epub 2019 Nov 6.

Department of the Control of Genetic Processes, Institute of Gene Biology, Russian Academy of Sciences, 34/5 Vavilov St., Moscow 119334, Russia. Electronic address:

Recently, the concept has arisen that a special class of architectural proteins exists, which are responsible not only for global chromosome architecture but also for the local regulation of enhancer-promoter interactions. Here, we describe a new architectural protein, with a total size of only 375 aa, which contains an N-terminal zinc finger-associated domain (ZAD) and a cluster of five zinc finger C2H2 domains at the C-terminus. This new protein, named ZAD and Architectural Function 1 protein (ZAF1 protein), is weakly and ubiquitously expressed, with the highest expression levels observed in oocytes and embryos. The cluster of C2H2 domains recognizes a specific 15-bp consensus site, located predominantly in promoters, near transcription start sites. The expression of ZAF1 by a tissue-specific promoter led to the complete blocking of the eye enhancer when clusters of ZAF1 binding sites flanked the eye enhancer in transgenic lines, suggesting that the loop formed by the ZAF1 protein leads to insulation. The ZAF1 protein also supported long-range interactions between the yeast GAL4 activator and the white promoter in transgenic Drosophila lines. A mutant protein lacking the ZAD failed to block the eye enhancer or to support distance interactions in transgenic lines. Taken together, these results suggest that ZAF1 is a minimal architectural protein that can be used to create a convenient model for studying the mechanisms of distance interactions.
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http://dx.doi.org/10.1016/j.bbagrm.2019.194446DOI Listing
January 2020

Removal of extra sequences with I-SceI in combination with CRISPR/Cas9 technique for precise gene editing in Drosophila.

Biotechniques 2019 04;66(4):198-201

Group of Molecular Organization of Genome, Russian Academy of Sciences, 34/5 Vavilov St, Moscow 119334, Russia.

The CRISPR/Cas9 system has recently emerged as a powerful tool for functional genomic studies and has been adopted for many organisms, including Drosophila. Previously, an efficient two-step strategy was developed to engineer the fly genome by combining CRISPR/Cas9 with recombinase-mediated cassette exchange (RMCE). This strategy allows the introduction of designed mutations into a gene of interest in vivo. However, the loxP or frt site remains in the edited locus. Here, we propose a modification of this approach for rapid and efficient seamless genome editing with CRISPR/Cas9 and site-specific recombinase-mediated integration (SSRMI) combined with recombination between homologous sequences induced by the rare-cutting endonuclease I-SceI. The induced homological recombination leads to the removal of the remaining extraneous sequences from the target locus.
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http://dx.doi.org/10.2144/btn-2018-0147DOI Listing
April 2019

Opbp is a new architectural/insulator protein required for ribosomal gene expression.

Nucleic Acids Res 2017 Dec;45(21):12285-12300

Institute of Gene Biology, Russian Academy of Sciences, 34/5 Vavilova St., Moscow 119334, Russia.

A special class of poorly characterized architectural proteins is required for chromatin topology and enhancer-promoter interactions. Here, we identify Opbp as a new Drosophila architectural protein, interacting with CP190 both in vivo and in vitro. Opbp binds to a very restrictive set of genomic regions, through a rare sequence specific motif. These sites are co-bound by CP190 in vivo, and generally located at bidirectional promoters of ribosomal protein genes. We show that Opbp is essential for viability, and loss of opbp function, or destruction of its motif, leads to reduced ribosomal protein gene expression, indicating a functional role in promoter activation. As characteristic of architectural/insulator proteins, the Opbp motif is sufficient for distance-dependent reporter gene activation and enhancer-blocking activity, suggesting an Opbp-mediated enhancer-promoter interaction. Rather than having a constitutive role, Opbp represents a new type of architectural protein with a very restricted, yet essential, function in regulation of housekeeping gene expression.
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http://dx.doi.org/10.1093/nar/gkx840DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5716193PMC
December 2017

Architectural protein Pita cooperates with dCTCF in organization of functional boundaries in Bithorax complex.

Development 2017 07 15;144(14):2663-2672. Epub 2017 Jun 15.

Department of the Control of Genetic Processes, Institute of Gene Biology, Russian Academy of Sciences,  Moscow 119334, Russia

Boundaries in the Bithorax complex (BX-C) of delimit autonomous regulatory domains that drive parasegment-specific expression of homeotic genes. BX-C boundaries have two crucial functions: they must block crosstalk between adjacent regulatory domains and at the same time facilitate boundary bypass. The C2H2 zinc-finger protein Pita binds to several BX-C boundaries, including and To study Pita functions, we have used a boundary replacement strategy by substituting modified DNAs for the boundary, which is located between the and regulatory domains. Multimerized Pita sites block crosstalk but fail to support regulation of (bypass). In the case of , we used a novel sensitized background to show that the two Pita-binding sites contribute to its boundary function. Although is from BX-C, it does not function appropriately when substituted for : it blocks crosstalk but does not support bypass. Mutation of the Pita site disrupts blocking activity and also eliminates dCTCF binding. In contrast, mutation of the dCTCF site does not affect Pita binding, and this mutant boundary retains partial function.
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http://dx.doi.org/10.1242/dev.149815DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5536930PMC
July 2017

Architectural proteins Pita, Zw5,and ZIPIC contain homodimerization domain and support specific long-range interactions in Drosophila.

Nucleic Acids Res 2016 09 2;44(15):7228-41. Epub 2016 May 2.

Institute of Gene Biology, Russian Academy of Sciences, Vavilova str. 34/5, Moscow 119334, Russia

According to recent models, as yet poorly studied architectural proteins appear to be required for local regulation of enhancer-promoter interactions, as well as for global chromosome organization. Transcription factors ZIPIC, Pita and Zw5 belong to the class of chromatin insulator proteins and preferentially bind to promoters near the TSS and extensively colocalize with cohesin and condensin complexes. ZIPIC, Pita and Zw5 are structurally similar in containing the N-terminal zinc finger-associated domain (ZAD) and different numbers of C2H2-type zinc fingers at the C-terminus. Here we have shown that the ZAD domains of ZIPIC, Pita and Zw5 form homodimers. In Drosophila transgenic lines, these proteins are able to support long-distance interaction between GAL4 activator and the reporter gene promoter. However, no functional interaction between binding sites for different proteins has been revealed, suggesting that such interactions are highly specific. ZIPIC facilitates long-distance stimulation of the reporter gene by GAL4 activator in yeast model system. Many of the genomic binding sites of ZIPIC, Pita and Zw5 are located at the boundaries of topologically associated domains (TADs). Thus, ZAD-containing zinc-finger proteins can be attributed to the class of architectural proteins.
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http://dx.doi.org/10.1093/nar/gkw371DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5009728PMC
September 2016