Download Department of Cellular and Molecular Biology (1

Laboratory of the Molecular Biology of Stem Cells
Alexey TOMILIN
Ph.D., D.Sci.,
Corresponding Member of the Russian Academy of Sciences,
Head of the Laboratory
Institute of Cytology
Russian Academy of Science (INC RAS)
Tikhoretsky avenue 4
194064 St-Petersburg
Russian Federation
Skoltech Center for Stem Cell Research,
Skolkovo Institute of Science and Technology,
Novaya Street 100,
143025 Skolkovo, Moscow Region
Russian Federation
E-mail: [email protected]
Тel: +7-(812)-297-4519
Fax: +7-(812)-297-3541
BRIEF CURRICULUM VITAE
1987-1992
Undergraduate Studies, School of Physics, Dept. Biophysics, Polytechnical State
Institute, Leningrad, USSR/Russia.
1993-1997
Ph.D. studies in the Institute of Cytology, Academy of Sciences, St-Petersburg,
Russia and University of Caen, France.
1998-2002
Postdoctoral studies at the EMBL, Heidelberg, Germany, and University of
Pennsylvania, USA (Prof. Hans R. Schöler).
2002-2006
Group Leader, Department Developmental Biology (head Prof. Davor Solter), MaxPlanck-Institute for Immunobiology, Freiburg, Germany.
2007-now
Head of the Lab, Institute of Cytology, Russian Academy of Science, St-Petersburg,
Russia
CURRENT RESEARCH TOPICS
The laboratory, established in the Institute of Cytology in 2006, has broad interests in
cell and molecular biology with a primary focus on pluripotent stem cells, such as
embryonic stem (ES) and induced pluripotent stem (iPS) cells. Study of the molecular
mechanisms of stem cell maintenance and development is our primary emphasis,
however, we are increasingly working on developing applications of pluripotent stem
cells in clinics. The most recent and exciting current topics are as follows.
Mechanisms of transcriptional regulation of Oct4 gene
Oct4 gene is a central cellular player involved in the maintenance of cellular
pluripotency, as well as the establishment of pluripotent
cell state during in vitro reprogramming. Considering
the importance of this function, a highly relevant pursuit
is to dissect molecular mechanisms controlling the
expression of Oct4 gene itself. We have recently
identified novel factors binding and controlling Oct4
expression via the well-known distal enhancer
previously shown to be essential and sufficient for Oct4
expression in pluripotent and germ cells. Studies
unraveling biological role molecular details and of this
regulation are currently under way. Besides
fundamental aspects, the obtained knowledge may provide an important tool, for
example, to improve the efficiency of iPS cell generation.
Oct4 functions in stem cells
It is becoming increasingly clear that Oct4 functions go beyond the control of identity
of pluripotent and germ cells. A recent collaborative study has provided the first
genetic evidence that Oct4 plays a role in some somatic cells and tissues that
undergo stress during pathogenesis and tissue responses to injury (manuscript in
preparation). Molecular mechanism of Oct4 induction and target genes under these
stress conditions are in focus of our research.
Structure of eukaryotic chromatin
The nucleosome as structural unit of chromatin is sufficiently stable to maintain
structures of higher order. At the same time, it must
ensure the availability the information encoded in the
DNA. Much data allows us to consider the nucleosome
as a dynamic structure that can reversibly change its
conformation and composition. We have studied
mononucleosomes reconstituted from a DNA core
histone octamer by atomic force microscopy (AFM) and
found their remarkable structural diversity caused by
disassembly
of
octasome
and
formation
of
subnucleosomal structures. These structures were due to
the quantized DNA unwrapping and partial histone
octamer dissociation. The equilibrium of these
subpopulations was significantly affected by ionic
environment (manuscript in preparation). The observed nucleosome structural
diversity serves as a paradigm of their functional plasticity dictated by physiological
environment.
Human artificial chromosomes (HACs) for ES/iPS cell-based regenerative
medicine and gene therapies
Human artificial chromosomes (HACs) are a powerful DNA vector system developed
recently to introduce large chromosomal fragments, genes and regulatory elements
into cultured mammalian cells without affecting the host genome. This approach is
devoid of known problems of viral or other
vector tools such as insertional mutagenesis
and unstable expression. However the delivery
of HACs directly into cells of living organism is
feasible so far only via cultured cells that are
first to be targeted with HACs, and then
incorporated into desired tissues and organs.
Pluripotent stem cells such as embryo-derived
embryonic stem (ES) cells and autologous
induced pluripotent stem (iPS) cells seem to be
an ideal choice for HAC delivery via tissuereplacement because they possess the
capacity for unlimited self-renewal ex vivo and can differentiate into virtually any cell
type of the organism both in vivo and in vitro (reviewed in Kouprina et al., 2014,
Expert Opin. Drug Del. 11(4): 1-19). As initial step of the approach, we have recently
generated and functionally evaluated mouse ES cells carrying alphoid tetO-HAC, the
newest generation of in vitro assembled HACs, was transferred into mouse ES cells.
Autonomous maintenance of this HAC was checked via FISH analysis of
chromosome spreads. Our data suggest that alphoidtetO-HACs can be stably
maintained and expressed in pluripotent and derived thereof differentiated cells
without any detrimental effects on developmental processes (Liskovykh et al., 2015,
Cell Cycle, in press). The result thus serves as a paradigm for the development of
further HAC-based approaches to tackling a broad range of hereditary recessive
diseases in human, which is currently being actively pursued in our lab using different
mouse models.
LAB MEMBERS
Main Group
Dr. Sergey SINENKO
Staff Scientist
[email protected]
Elena SKVORTSOVA
Research Assistant
[email protected]
Alexander KHUDYAKOV
PhD student
[email protected]
Dr. Igor NAZAROV
Staff Scientist
[email protected]
Evgeny BAKHMET
PhD student
[email protected]
Natalia KHRAMOVA
Technitian
[email protected]
Sergey PONOMARTSEV
PhD student
[email protected]
Natalia KATOLIKOVA
Skoltech PhD student
[email protected]
Elena KUZMINA
Student
[email protected]
Alexandra KONDRASHKINA
Student
[email protected]
Andrey KUZMIN
Student
[email protected]
Cancer Stem Cell Group
Dr. Elena TOLKUNOVA
Staff Scientist
[email protected]
Grigory TIMIN
Student
[email protected]
Since 2009 the lab includes the Biophysics Group
Former lab members:
Dr. Mikhail Liskovykh, currently post-doc, National Cancer Institute, NIH, Bethesda,
USA;
Dr. Alexander Davydov-Sinitsyn, currently working for a company;
Dina Safina, currently PhD student, Department for Cell Morphology and Molecular
Neurobiology, Ruhr-University Bochum, Bochum, Germany;
Lyubov Chechik, currently PhD student Max-Planck Institute of Immunobiology and
Epigenetics, Freiburg (Germany);
Dr. Anna MALASHICHEVA, currently Head of Laboratory, Almazov Federal Heart,
Blood and Endocrinology Centre, St-Petersburg (Russia);
Birgit KOSCHORTZ, MPI-Freiburg;
Robert TUREK, MPI-Freiburg;
Volker KOELIN, MPI-Freiburg;
COLLABORATORS
Dr. Vladimir Larionov and Dr. Natalia Kouprina, National Cancer Institute, NIH,
Bethesda, USA;
Prof. Gary K. Owens and Dr. Olga Cherepanova, Virginia University, USA;
Prof. Michael Bader and Dr. Natalia Alenina, Max-Delbruck Center for Biomedicine,
Berlin, Germany;
PUBLICATIONS BY LAB MEMBERS
(starting from the most recent ones; lab members are highlighted in bold)
2010-present
Liskovykh M, Ponomartsev S, Popova E, Bader M, Kouprina N, Larionov V,
Alenina N, Tomilin A. (2015) Stable maintenance of de novo assembled human
artificial chromosomes in embryonic stem cells and their differentiated progeny in
mice. Cell Cycle, Feb 19: 0. [Epub ahead of print].
Kulichkova VA, Artamonova TO, Zaykova JJ, Ermolaeva JB, Khodorkovskii MA,
Barlev NA, Tomilin A, Tsimokha AS (2014) Simultaneous EGFP and Tag Labeling of
the b7 Subunit for Live Imaging and Affinity Purification of Functional Human
Proteasomes. Mol Biotechnol. Aug 28. [Epub ahead of print]
Bajenova O, Chaika N, Tolkunova E, Davydov-Sinitsyn A, Gapon S, Thomas P,
O'Brien S. (2014) Carcinoembryonic antigen promotes colorectal cancer progression
by targeting adherens junction complexes. Exp Cell Res. Epub 2014 Apr 12.
Kouprina N, Tomilin A, Masumoto H, Earnshaw WC, Larionov V. (2014) HAC-based
gene delivery vectors for gene function studies, gene therapy and pharmacology.
Expert Opin. Drug Del. 11(4): 1-19.
Yang CS, Sinenko SA, Thomenius MJ, Robeson AC, Freel CD, Horn SR, Kornbluth
S. The deubiquitinating enzyme DUBAI stabilizes DIAP1 to suppress Drosophila
apoptosis. Cell Death Differ. 2014 Apr;21(4):604-11. doi: 10.1038/cdd.2013.184.
Epub 2013 Dec 20.
DeVeale B, Brokhman I, Mohseni P, Babak T, Yoon C, Lin A, Onishi K, Tomilin A,
Pevny L, Zandstra PW, Nagy A, van der Kooy D (2013) Oct4 is required for e7.5 for
proliferation in the primitive streak. PLoS Genet. 9(11): e1003957.
Wu G, Han D, Gong Y, Sebastiano V, Gentile L, Singhal N, Adachi K, Fischedick G,
Ortmeier C, Sinn M, Radstaak M, Tomilin A, Schöler HR. (2013) Establishment of
totipotency does not depend on Oct4. Nat Cell Biol. 15(9): 1089-97.
Lee N.C.O, K. A., Lee H.S, Tolkunova E.N., Liskovykh М.A., Masumoto H.,
Earnshaw W.C., Tomilin A., Larionov V., Kouprina N. (2013) Protecting a transgene
expression from the HAC-based vector by different chromatin insulators. Cell. Mol.
Life Sci. 70(19): 3723-37.
Nazarov I, Krasnoborova V, Mitenberg A, Chikhirzhina E, Davidov-Sinitsin A,
Liskovykh M, and Tomilin A (2014) Transcription Regulation of Oct4 (Pou5F1)
Gene by Its Distal Enhancer. Cell and Tissue Biol 8(1): 27-32.
Davydov-Sinitsyn A, O. V. Bazhenova, M. A. Liskovykh, L. L. Chechik, S. V.
Ponomartsev S, Tomilin A, Tolkunova E. (2013) In vitro Derivation and
Characterization of a Colorectal Cancer Stem Cell Subpopulation. Cell and Tissue
Biol 7(4): 320-325.
Davydov-Sinitsyn A, Bajenova O, Liskovykh M, Ponomartsev S, Chechik L,
Tomilin A, Tolkunova E. (2012) Comparative Analysis of Colorectal Carcinoma Cell
Lines That Differ in Metastatic Potential. Cell and Tissue Biol 7(5): 407-412.
***Liskovykh M., Chuykin I., Ranjan A., Safina D., Popova E., Tolkunova E.,
Mosienko V., Minina Iu., Zhdanova N., Mullins J.J., Michael Bader M., Alenina N.,
Tomilin A. (2011) Derivation, Characterization, and Stable Transfection of Induced
Pluripotent Stem Cells from Fischer344 Rats // PLoS ONE 11(6): e27345.
Liskovykh M, Davydov-Sinitcyn A, Marilovtceva E, Tomilin A, Tolkunova E.
(2011) Interaction between the CDX2 Transcription Factor and DDX5 Protein. Cell
and Tissue Biol. 6(1): 20-25.
Mondal BC, Mukherjee T, Mandal L, Evans CJ, Sinenko SA, Martinez-Agosto JA,
Banerjee U. (2011) Interaction between differentiating cell- and niche-derived signals
in hematopoietic progenitor maintenance. Cell 147(7): 1589-600.
Sinenko SA, Shim J, Banerjee U. (2011) Oxidative stress in the haematopoietic
niche regulates the cellular immune response in Drosophila. EMBO Rep. 13(1):83-9
Sinenko SA, Hung T, Moroz T, Tran QM, Sidhu S, Cheney MD, Speck NA, Banerjee
U (2010) Genetic manipulation of AML1-ETO-induced expansion of hematopoietic
precursors in a Drosophila model. Blood 116(22): 4612-20.U.
Before 2010
Kuckenberg P., Buhl S., Woynecki T., van Fürden B., Tolkunova E., Seiffe F., Moser
M., Tomilin A., Winterhager E., Schorle H. (2010) The transcription factor
TCFAP2C/AP-2gamma cooperates with CDX2 to maintain trophectoderm formation.
Mol. Cell. Biol. 30: 3310-3320. download
Tomilin A., Tolkunova E., Liskovykh M. (2009) A method of stem cell application
for tissue-replacement therapies. Russian Federation patent No. 2009143025.
Saxe J., Tomilin А., Schöler H.R., Plath K., Huang J. (2009) Post-Translational
Regulation of Oct4 Transcriptional Activity. PLOS One 4: 1-9. download +
supplement
Tolkunova E., Malashicheva A., Chikhirzhina E., Kostyleva E., Zeng W., Luo J.,
Dobrinski I., Hierholzer A., Kemler R., and Tomilin A. (2009) E-Cadherin as a Novel
Surface Marker of Spermatogonial Stem Cells. Cell and Tissue Biol. 3: 103.
download
Popov B., Petrov N., Mikhailov V., Tomilin A., Alekseenko L., Grinchuk T., and
Zaichik A. (2009) Spontaneous Transformation and Immortalization of Mesenchymal
Stem Cells in vitro. Cell and Tissue Biol. 3: 110-115.
Sinenko SA, Mandal L, Martinez-Agosto JA, Banerjee U. (2009) Dual role of
wingless signaling in stem-like hematopoietic precursor maintenance in Drosophila.
Dev Cell 16(5): 756-63.
Tolkunova A., Malashicheva A., Parfenov V.N., Sustmann C., Grosschedl R.,
Tomilin A. (2007) PIAS proteins as repressors of Oct4 function. J. Mol. Biol. 374:
1200-1212. download + supplement
Lengner C.J., Camargo F.D., Hochedlinger K., Welstead G.G., Zaidi S., Gokhale S.,
Schöler H.R., Tomilin A., Jaenisch R. (2007) Oct4 expression is not required for
mouse somatic stem cell self-renewal. Cell Stem Cell 1: 403–415. download +
supplement
Malashicheva A., Kanzler B., Tolkunova E., Trono D., Tomilin A. (2007) Lentivirus
as a tool for lineage-specific gene manipulations. Genesis 45, 456-459. download +
supplement
Tolkunova E., Cavaleri F., Eckardt S., Reinbold R., Christenson L.K., Schöler H.R. ,
Tomilin A. (2006) The Caudal-Related Protein Cdx2 Promotes Trophoblast
Differentiation of Mouse ES Cells. Stem Cells 24: 139-144. download + supplement
Kehler J, Tolkunova E., Koschorz B., Pesce M., Gentile L., Boiani M, Lomeli H,
Nagy A, McLaughlin K.J., Scholer H.R., Tomilin A. (2004) Oct4 is required for
primordial germ cell survival. EMBO Rep. 5: 1078-83. download + supplement
Lins K., Reményi A., Tomilin A., Massa S., Wilmanns M., Matthias P. and Schöler
H.R. (2003) OBF1 enhances transcriptional potential of Oct1. EMBO J. 22: 21882198. download
Seralini G.E., Tomilin A., Auvray P., Nativelle-Serpentini C., Sourdaine P., Moslemi
S. (2003) Molecular characterization and expression of equine testicular cytochrome
P450 aromatase. Biochim Biophys Acta. 1625: 229-238. download
Kobayashi M., Fujioka M., Tolkunova E., D. Deka, M. Abu-Shaar, R. S. Mann, and
J.B. Jaynes. (2003) Engrailed cooperates with extradenticle and homothorax to
repress target genes in Drosophila. Development 130: 741-51. download
Remenyi A, Tomilin A, Scholer HR, Wilmanns M. (2002) Differential activity by DNAinduced quarternary structures of POU transcription factors. Biochem Pharmacol.
64:979-84. download
Perez-Martinez X, Funes S, Tolkunova E, Davidson E, King MP, Gonzalez-Halphen
D. (2002) Structure of nuclear-localized cox3 genes in Chlamydomonas reinhardtii
and in its colorless close relative Polytomella sp. Curr Genet. 40: 399-404. download
*Remenyi A., *Tomilin A., Pohl E., Lins K., Philippsen A., Reinbold R., Schöler H.R.,
and Wilmanns M. (2001) Differential transcriptional activity of dimeric Oct-1 by DNAmotif induced domain swapping. Mol. Cell 8: 569-580. download
*equal contributors
Perez-Martinez X., Antaramian A., Vazquez-Acevedo M, Funes S, Tolkunova E,
d'Alayer J, Claros MG, Davidson E, King MP, Gonzalez-Halphen D. (2001) Sububunit
II of cytochrome c oxidase in Chlamydomonad algae is a heterodimer encoded by
two independent nuclear genes. J Biol Chem 276: 11302-9. download
#Tomilin
A., #Remenyi A., Lins K., Bak H., Leidel S., Vriend G., Wilmanns M.,
Scholer H.R. (2000) Synergism with the coactivator OBF-1 (OCA-B, BOB-1) is
mediated by a specific POU dimer configuration. Cell 103: 853-864. download
#equal contributors
Perez-Martinez X., Vazquez-Acevedo M., Tolkunova E., Funes S., Claros MG,
Davidson E., King D., Gonzales-Halphen D. (2000) Unusual location of mitochondrial
gene. Subunit III of cytochtome C oxidase is encoded in the nucleus of C. algae. J
Biol Chem. 275: 30144-30152. download
Tolkunova E., Park H., Xia J., King MP., Davidson E. (2000) The human lysyl-tRNA
synthetase gene encodes both the cytoplasmic and mitochondrial enzymes by
means of an unusual alternative splicing of the primary transcript. J Biol Chem 275:
35063-35069. download
Kropotov A., Sedova V., Ivanov V., Sazeeva N., Tomilin A., Krutilina R., Oei S.L.,
Griesenbeck J., Buchlow G., Tomilin N. (1999) A novel human DNA-binding protein
with sequence similarity to a subfamily of redox proteins which is able to repress
RNA-polymerase-III-driven transcription of the Alu-family retroposons in vitro. Eur. J.
Biochem. 260: 336-346.
Tomilin A., Vorob'ev V., Drosdowsky M., Seralini G.-E. (1998) Oct3/4-associating
proteins from embryonal carcinoma and spermatogenic cells of mouse. Mol. Biol.
Rep. 25: 103-109.
Tolkunova E., Fujioka M., Kobayashi M., Deka D., Jaynes J. (1998) Two distinct
types of repression domain in engrailed: one interacts with groucho co-repressor and
is preferentially active in integrated target genes. Mol. Cell. Biol. 18(5): 2804-2814.
download
Belokopytova I.A., Kostyleva E.I., Tomilin A., Vorob'ev V.I. (1993) Human male
infertility may be due to a decrease of the protamine P2 content in sperm chromatin.
Mol. Reprod. Dev. 34: 53-57.
FINANCIAL SUPPORT
Current financial support:

Skolkovo Foundation endowment to the Center of Research and
Enterprenership (CREI) for Stem Cells in the Skolkovo Institute of Science and
Technology (SkolTech);

Russian Science Foundation grant 14-15-00718;

Russian Science Foundation grant 14-50-00068;

Russian Foundation for Basic Research (13-02-00923a)

Russian Foundation for Basic Research (14-04-31556_mol_a) to Liskovykh
M./Ponomartsev S.
Past financial support:

Grant program „Molecular and Cellular Biology“ by the Presidium of the
Russian Academy of Science (2006-2012);

State contracts with the Ministry of Science and Education (02.512.11.2253,
16.512.11.2085, 16.512.11.2242, 2008-2012);

Russian Foundation for Basic Research (RFBR) – Helmholtz Society
(Germany) joint grant (07-04-92281/HRJRG-024, 2009-2011);

Russian Foundation for Basic Research (RFBR, 07-04-01154a, 2007-2008);