Molecular basis of gene expression
Transcription, copying genetic information into RNA, is the first step of gene expression. In all living organisms transcription is accomplished by conserved multi-subunit RNA polymerases, one of the most ancient enzymes on the planet. Understanding functions of RNA polymerase are essential for understanding evolution of Life of the planet. Importantly, malfunctions of RNA polymerases are linked to various human diseases including cancer and Alzheimer. Also transcription is the potent target for antimicrobials. Many mechanistic details of the functioning of RNA polymerases are not clear. Furthermore RNA polymerases are imbedded in intricate relations with other cellular machineries, such as translation in prokaryotes or replication and repair in all organisms. Regulation of these interactions is pivotal for coordination of cellular processes, correct gene expression and ultimately for survival of organisms. The mechanisms underlining these interactions and their regulation are also poorly understood.
We are investigating transcription. All possible aspects: from mechanisms of reactions and inhibition by antibiotics to regulation and interactions with other cellular machineries, such as translation and replication. We use classical biochemistry and molecular biology, some unique experimental systems as well as novel techniques. In vitro we investigate bacterial and archaeal RNA polymerases as well as eukaryotic RNA polymerases I, II and III. In vivo we are focusing on bacterial transcription.
The main goal of our study is to understand how RNA polymerase evolved to the enzyme we know today, and how it functions and is regulated in the modern cells. Another important aspect of our work is to look for the ways of manipulation of RNA polymerases (such as specific inhibition) in order to control pathogenicity.
Bacterial defence systems
Bacteria invented several mechanisms to fight against their enemies –bacteriophages and antibiotics. One of such systems, toxin-antitoxin (TA) systems, involves a large number of small proteins, called toxins, which reversibly target essential processes in the bacterial cell to stop these processes and bring the cell to dormancy. Such dormant cells become temporarily resistant to antibiotics (phenomenon called persistence) because antibiotic targets are temporarily non-functional in them, as well as become “not interesting” to bacteriophages, because they cannot propagate in dormant cells. There are also some other biological important phenomena, such as programmed cell death and addiction to plasmids, linked to TA systems. TA systems involve large number of activities and targets, many of which are not yet known or characterised.
We are interested in mechanistic and catalytic details of functioning of TAs and the outcomes of their action for their targets.
Toxin-antitoxin system described above is a “passive” one, i.e. the cell uses it to wait through the harsh conditions. Recently an “active” defensive immune system of bacteria (CAS/CRISPR system) was discovered. Bacteria store information about bacteriophages that have attacked them in the past, and use this information to kill bacteriophages if they enter the cell again. Bacterial immune system, however, is not related to the known immune systems. Information about the phage is stored in bacterial genome as short sequences (called CRISPRs) complementary to the phage genome. Upon phage invasion, the sequences are transcribed and the processed RNA along with large protein complex (CAS proteins) recognises phage genome and/or phage mRNA and destroys it.
The process of phage recognition and destruction is well studied. We are interested in the mechanism of acquisition of the immunity memory by CAS/CRISPR system, which remain a mystery.
Specific: cellular machineries working with nucleic acids.
General: molecular evolution, physics of high gravities and velocities.
1. Castro-Roa, D., Garcia-Pino, A, van Nuland, N. A. J., Loris, R., and Zenkin, N*. (2013) The Fic protein Doc uses an inverted substrate to phosphorylate and inactivate EF-Tu. Nat Chem Biol 9:811-7
2. Germain, E., Daniel Castro-Roa, D., Zenkin, N*., and Gerdes, K. (2013) Molecular Mechanism of Bacterial Persistence by HipA. Mol Cell 52:248-54
3. Nielsen, S.U., Yuzenkova, Y., and Zenkin, N*. (2013). Mechanism of RNA polymerase III transcription termination Science, 340: 1577-1580.
4. Bochkareva, A., Yuzenkova, Y., Tadigotla, V. and Zenkin, N*. (2012). Factor-independent transcription pausing caused by recognition of the RNA-DNA hybrid sequence. EMBO J 31, 630-639
5. Yuzenkova, Y., Tadigotla, V.R., Severinov, K., and Zenkin, N*. (2011). A new basal promoter element recognized by RNA polymerase core enzyme. EMBO J 30, 3766-3775.
6. Yuzenkova, Y., Zenkin, N*. (2010) Central role of the RNA polymerase trigger loop in intrinsic RNA hydrolysis Proc Natl Acad Sci U S A 107(24):10878-83
7. Zenkin, N., Kulbachinskiy, A., Yuzenkova, Y., Mustaev, A., Bass, I., Severinov, K. and Brodolin, K. (2007). Region 1.2 of the RNA polymerase sigma subunit controls recognition of the -10 promoter element. EMBO J 26, 955-64.
8. Zenkin, N*., Yuzenkova, Y. and Severinov, K. (2006) Transcript-assisted transcriptional proofreading. Science, 313, 518-20.
9. Zenkin, N*., Naryshkina, T., Kuznedelov, K. and Severinov, K. (2006) The mechanism of DNA primer synthesis by RNA polymerase. Nature, 439, 617