Doctor of Philosophy
|Period||Coyntry, city||University||Additional info|
|2007–2010||Russia, Moscow||M.M. Shemyakin and Yu.A. Ovchinnikov Institute of bioorganic chemistry of the Russian Academy of Sciences|
|2002–2007||Russia, Moscow||Lomonosov Moscow State University||Graduated summa cum laude|
|Research fellow||Laboratory of human genes structure and functions|
|Doctor of Philosophy (Biological sciences)|
RNA types, structure and functions
I have a keen interest in RNA biology in general and non-coding RNAs in particular. The structural and functional diversity of RNAs is very high, and there still are many unanswered questions in the field of RNA biology. The most obvious function of RNA is to facilitate the conversion of DNA information into polypeptides; but RNA can also store genetic information as well as act as an enzyme. However, only a small percentage of the DNA in eukaryotic genomes encodes proteins, though the vast majority of eukaryotic genomes are transcribed into RNA. A significant fraction of non-coding RNAs have no identified function and thus these RNA molecules were dubbed “dark matter in the genome”. And while the question whether this “RNA dark matter” has functional relevance, or is a product of protein-coding genes transcription, or even just represents transcriptional noise is open, there is no doubt that a significant number of non-coding RNAs participate in regulatory processes.
Transcriptome analysis methods
All RNAs in a cell are involved in an ultra-complex network of interactions with each other, DNA, proteins, and other molecules, and constitute a transcriptome of a cell. The transcriptome is highly dynamic and is constantly changing in response to endogenous and exogenous stimuli. Thus, transcriptome studies are cornerstone of functional genomics and provide insight into which genes and non-coding RNAs are expressed in specific circumstances. Nevertheles, transcriptome studies were hampered until recently due to the lack of methods for quantitative and qualitative analysis of RNA pool. DNA microarrays and next-generation sequencing have made it possible to accurately measure compositions of transcriptomes of many eukaryotic and prokaryotic species. Next-generation sequencing produces huge amounts of sequencing data, but gives no information on secondary structures of sequences. Since most non-coding RNAs are characterized by a specific secondary and tertiary structure, the structure of RNA must be known before the relationship between its structure and function can be determined. Prediction of RNA structure as well as analysis of next-generation sequencing data requires solid knowledge of existing bioinformatics software tools and programming languages and good computational skills, so I started to study bioinformatics during my research on mycobacterial transcriptomes and eventually developed a strong interest in this area.
Bacterial transcriptomics is a very interesting field of research, and working with bacteria has its own advantages, disadvantages and peculiarities. Bacteria grow fast, large amounts of RNA can be easily isolated from their cultures, but bacterial RNAs are not polyadenylated, and thus cannot be amplified using oligo-dT primers. Rapid advancements in sequencing technology have shifted the paradigm that considers bacterial transcriptomes as rather simple in comparison with the far more complex eukaryotic ones. It was shown recently that bacterial cells utilize most, If not all, transcriptional regulatory mechanisms of their eukaryotic counterparts, i.e. alternative transcription, RNA processing and splicing, regulation of mRNA stability by polyadenylation and small RNAs, to name a few. This unraveling of a whole new level of bacterial complexity is of the highest importance, not only for fundamental science, but also for biotechnology and medicine, and thus my current research is focused on investigating the role of small RNAs in mycobacterial infections and their potential as therapeutic targets.
Pathogenomics and bacterial physiology
Genome-wide studies of gene expression provide a window into how bacterium’s genetic makeup enables it to function and adapt to its environment. In case of pathogenic bacteria such studies are invaluable for identifying the virulence factors that allow a pathogen to survive a host's defence mechanism. Nevertheless, host-pathogen interactions are too complex to be deciphered by transcriptome analyses alone. For example, it was shown that genomic differences between two strains of the same bacterial species may be very significant, leading to different transcriptional outcomes in the same environment. This influences the metabolic profiles of bacterial cells and might contribute to variations in pathogenicity. This is the reason of my vivid interest in various aspects of bacterial structural and comparative genomics as well as in physiology and biochemistry of bacteria.
Unusual nucleic acid structures
One of my oldest interests, dating back to high school, is in unusual DNA structures and conformations. I have never had a chance to participate in any research projects in this area, but since I find such noncanonical structures intriguing and firmly believe they must have interesting and important biological roles to play, I read a lot of literature on the subject.
Isolation and purification of DNA and RNA from bacterial and eukaryotic cells; PCR; qPCR; RT-PCR; RACE; qPCR; PCR site-directed mutagenesis; restriction and cloning; enzymatic modification of nucleic acids; working with bacterial cell cultures; preparation of DNA libraries (cDNA and genomic); agarose and polyacrylamide gel electrophoresis (both native and denaturating); Southern and Northern blotting methods; SAGE; nucleic acid sequence analysis; analysis of Illumina GA IIx and Roche GS FLX next-generation sequencing systems data; database mining.
GeneRunner, Vector NTI; Gel-Pro Analyzer; Chromas; GraphPad Prism; programming languages: Perl (beginner), R (beginner).
|I am currently working on the structural and functional characterization of several Mycobacterium tuberculosis proteins identified in earlier transcriptomic studies as potential therapeutical targets; I am also conducting a study of small RNAs in Mycobacterium avium|
|2007-2011||Junior Research Fellow|
|Developed a new method for transcriptional profiling of intracellular pathogens in vivo based on DNA-DNA hybridization and RNA-Seq; applied this method for mapping and quantifying transcriptomes of Mycobacterium tuberculosis and Mycobacterium avium during infection in a mouse model; identified a set of M. tuberculosis genes that invariably demonstrated increased expression in unfavorable for pathogen conditions.|
|Evaluated gene expression differences between two Mycobacterium tuberculosis strains; optimized real-time qPCR data analysis methods.|
|Conducted a study of Mycobacterium tuberculosis genomic DNA methylation using methyl-sensitive restriction analysis, affinity chromatography, bisulfite sequencing and methylated DNA immunoprecipitation (MeDIP).|