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To approach this problem we are using Drosophila transcriptional
factor E2F as a model system. E2F is a critical regulator
of cell cycle progression and plays a key role in orchestration
of the cell cycle and cell growth. E2F family members
are one of the most important downstream targets of
retinoblastoma tumor suppressor protein (pRB) and deregulation
of E2F is thought to drive proliferation of most tumor
cells. Studies of E2F in mammalian cells are hampered
by the large numbers of E2F and pRB family members.
In flies, there are only two E2Fs and unlike mammalian
cells they have clearly distinct biochemical properties.
Another advantage of Drosophila is the ability to utilize
a variety of genetic tools in order to identify important
functional interactions in signaling pathways.In particular,
the most attractive potential of this approach is the
ability to screen through large numbers of randomly
generated mutations to isolate important interacting
modifiers in an unbiased fashion. By applying methods
of molecular genetics, biochemistry and cell biology
we are dissecting which repressor activities are the
most important for E2F function in vivo. In the reverse
approach, we are studying the effects on E2F dependent
repression resulting from systematical removal of members
of the histone lysine methyltransferase family.
The basic unit of eukaryotic chromatin is a nucleosome
comprising of DNA wrapped around a histone octamer.
Each histone contains a globular region and a less-structured
N-terminal tail that protrudes outwards. These tails
are targeted by a variety of post-translational modifications
which are thought to affect the properties of chromatin
and regulate gene expression. At present, one of the
most puzzling issues in the chromatin field is whether
histone methylation is reversible. Can methyl groups
be removed from histones enzymatically? Since de-methylase
activity has yet to be discovered, a prevailing view
is that methylation marks are more permanent and provide
epigenetic cue for a heterochromatic state. However,
there are some documented examples of the loss of methylation
following gene activation. We started to tease this
problem by performing a large scale genetic screen to
look for the mutations which elevate the level of histone
methylation and therefore could potentially uncover
pathways affecting putative de-methylation enzymes.
Several mutations were isolated from the screen and
their phenotypes are being analyzed. Current aim is
to map and clone the genes disrupted by the mutations
and evaluate the role of mammalian orthologs.
Selected Publications:
Frolov, M.V., E.V. Benevolenskaya and J.A. Birchler.
1998. Regena (Rga), a Drosophila homolog of the global
negative transcriptional regulator CDC36 (NOT2) from
yeast, modifies gene expression and suppresses position
effect variegation. Genetics 148: 317-329.
Frolov, M.V. and J.A. Birchler. 1998. Mutation in P0,
a dual function ribosomal protein/apurinic/apyrimidinic
endonuclease, modifies gene expression and position
effect variegation in Drosophila. Genetics 150: 1487-1495.
Alatortsev, V.E., A. Fadeeva, and M.V. Frolov. 2000.
P[lacW] transposon insertions with gradual expression
of reporter genes. Genetika 36: 630-635.
Benevolenskaya, E.V., M.V. Frolov and J.A. Birchler.
2000. Krüppel homolog (Kr h) is a dosage-dependent
modifier of gene expression in Drosophila. Genetical
Research 75: 137-142.
Frolov, M.V., E.V. Benevolenskaya, and J.A. Birchler.
2000. The oxen gene of Drosophila encodes a homolog
of subunit 9 of yeast ubiquinol-cytochrome c oxidoreductase
complex: evidence for modulation of gene expression
in response to mitochondrial activity. Genetics 156:
1727-1736.
Frolov, M.V., and V.E. Alatortsev. 2001. Molecular
analysis of novel Drosophila gene, Gap69C, encoding
a homolog of ADP-ribosylation factor GTPase-activating
protein. DNA & Cell Biology 20: 107-113.
Frolov, M.V., E.V. Benevolenskaya, and J.A. Birchler.
2001. Molecular analysis of a novel Drosophila diacylglycerol
kinase, DGK?. Biochimica & Biophysica Acta 1538:
339-352.
Frolov, M.V., D.S. Huen, O. Stevaux, D. Dimova, K.
Balczarek-Strang, and N.J. Dyson. 2001. Functional antagonism
between E2F family members. Genes & Development
15: 2146-2160.
Stevaux O, D. Dimova, M.V. Frolov, B. Taylor-Harding,
E.J. Morris, and N.J. Dyson. 2002. Distinct mechanisms
of E2F regulation by Drosophila RBF1 and RBF2. EMBO
J. 21: 4927-4937.
Frolov M.V., O. Stevaux, N.S. Moon, D. Dimova, E.J.
Kwon, E.J. Morris, and N.J. Dyson. 2003. G1 cyclin-dependent
kinases are insufficient to reverse dE2F2-mediated repression.
Genes & Development 17: 723-728.
Dimova, D., O. Stevaux, M.V. Frolov, and N.J. Dyson.
2003. Cell cycle-dependent and cell cycle-independent
control of transcription by the Drosophila E2F/RB pathway.
Genes & Development 17: 2308-2320.
Frolov, M.V. and N.J. Dyson. 2004. Molecular mechanisms
of E2F-dependent activation and pRB-mediated repression.
Journal of Cell Science 117: 2173-2181.
Dyson, N.J. and Frolov, M.V. The Retinoblastoma Protein.
2004. In Encyclopedia of Biological Chemistry. Elsevier/Academic
Press. San Diego.
Frolov M.V., N.S. Moon, and N.J. Dyson. 2005. DP is
needed for normal cell proliferation. Molecular and
Cellular Biology. 25: 3027-3039.
Stevaux O., D.K. Dimova, J.Y. Ji, N.S. Moon, M.V. Frolov,
N.J. Dyson. 2005. Retinoblastoma Family 2 is Required
In Vivo for the Tissue-Specific Repression of dE2F2
Target Genes. Cell Cycle 4: 1272-1280.
Moon N.S., M.V. Frolov, E.-J. Kwon, L. Di Stefano,
D.K. Dimova, E. Morris, B. Taylor-Harding, K. White
and N.J. Dyson. 2005. Drosophila E2F1 has context specific
pro- and anti- apoptotic properties during development.
Developmental Cell. In Press.
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