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CELL
CYCLE CONTROL
Gametogenesis
- Cyclin A's
We have identified a novel mammalian
A-type cyclin, cyclin Al, which we have shown to be expressed at highest
levels if not exclusively in the testis in mice and humans.
Targeted mutagenesis of Ccna1 results in viable progeny but
male sterility, while females are fully fertile. We
are currently extending our studies on the unique regulation and function
of cyclin Al in the male germ line and exploring the functional redundancy
of the two mammalian A-type cyclins.
Our current projects:
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identify
regulatory elements required for the proper in vivo expression
of Ccna1 |
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ask
if cyclin A2 can rescue the cyclin Al-deficient phenotype of meiotic
arrest in spermatogenesis, in transgenic mice in vivo |
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determine
the role of cyclin A1 in human infertility by sequencing CCNA1
from infertile men |
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identify interacting partners and substrates for the
A-type cyclins |
Cyclin
A1 and Oncogenesis Human
cyclin Al is highly expressed in leukemic cells from patients with acute
myeloid leukemia. To test the hypothesis that the aberrant high levels
of cyclin Al were causal in the leukemic phenotype, i.e., acting as
an oncogene, we generated transgenic mice in which cyclin Al was expressed
in myeloid precursor cells. The transgenic animals exhibited abnormal
myelopoiesis and developed acute myeloid leukemia. We have also recently
observed high levels of cyclin Al expression in testicular tumors of
the highly invasive embryonal carcinoma class but not in the more common
and less invasive seminoma. We propose that cyclin Al represents a novel
target for drug intervention in certain forms of cancer.
Our current projects:
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ask
if inhibiting the expression of cyclin Al in leukemic cells in
vitro and in vivo can revert the leukemic phenotype
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develop
efficient in vitro kinase assays for screening pharmacological
targets for inhibition specifically for cyclin Al/Cdk complexes |
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assess
the effects on normal hematopoiesis of the loss of cyclin A1 function
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examine
the importance of cytoplasmic versus nuclear localization of cyclin
A1 in leukemia cells |
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identify
interacting proteins and substrates of cyclin
Al/Cdk complexes in normal and leukemic cells |
Cell Cycle Arrest
and Apoptosis
Mice deficient for the cell cycle
regulating gene encoding Ccna1 are viable but male germ cells
arrest at the G2/M transition of meiotic prophase I. The cells
do not form haploid spermatids and rather, exhibit properties suggestive
of a rapid entry into an apoptotic cell death. Our hypothesis is
that there is a primary cell death response that is specifically activated
in response to cell cycle arrest. We also propose that disruption
of normal cell-cell associations, which are so critical for normal testicular
homeostasis, results in the subsequent loss of other cell types or ‘secondary
cell death.’ The mechanisms
by which these cells are lost remain to be determined but may involve
additional types of cell death and hence, different pathways.
Our current projects:
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examine
the role of p53 in apoptosis in Ccna -/-
spermatocytes
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morphological
analysis at the light end electron microscope level |
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accessment
of activity of various cell death pathways and activiation of markers
of cell death |
Cyclin
A2 and Heart Development
Irreversible exit from the cell
cycle during embryogenesis precludes the ability of cardiac muscle cells
to increase cell number after myocardial infarction. Cyclin A2 appears
to play a crucial role in cardiomyocyte development and proliferation,
and this role has not previously been studied adequately. We hypothesize
that cell cycle withdrawal during cardiogenesis is associated with a decrease
in cyclin A2 protein expression and that constitutive expression will maintain
proliferation through the later phases of development, the neonatal period,
and adulthood.
Our current projects:
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study
the role of cyclin A2 in cardiomyocyte development and proliferation
in transgenic mice that constitutively express cyclin A2 in the
cardiomyocyte lineage
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test
the hypothesis that the withdrawal of cardiomyocytes from the cell
cycle involves downregulation of cyclin A2 and ultimately a cascade
of inhibition that prevents phophorylation of an Rb-like protein |
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determine
how cyclin A2 is downregulated at the transcriptional level by investigating
promoter silencing through epigenetic modification such as methylation |
RETINOID
SIGNALING AND DEVELOPMENT OF THE REPRODUCTIVE SYSTEM
The
importance of dietary retinol (vitamin A) for the normal progression
of spermatogenesis and other aspects of male fertility has been recognized
for many years. Animals deprived of vitamin A in their diet exhibit
a variety of abnormalities, including male sterility. Most of
these effects can be corrected by supplementing the diet with retinoic
acid, with the exception of those involving vision and spermatogenesis.
Although retinoic acid is the active metabolite of vitamin A, retinol
is required for normal spermatogenesis to occur. This raises interesting
questions concerning the function of retinoids in the testis, with regard
both to their target genes which are required for spermatogenesis to
occur as well as to their metabolism within various testicular compartments.
Studies generating mutations in
specific receptors have clearly shown a role for the retinoid receptors
RAR-a and
RXR-ß in spermatogenesis.
Our current projects:
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characterize the phenotypic
abnormalities resulting in male sterility in the recently generated
strains of mice mutated in the RARa gene,
with particular emphasis on the developmental etiology of the abnormalities
in the testis
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test the hypothesis that the
mutations in the RARa
gene are essentially phenocopied by the effects of vitamin A deficiency
on spermatogenesis in mice by comparing the spermatogenic abnormalities
in the mutant mice with those in mice which have been vitamin A-deficient
from birth
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explore the role of the RARa
receptor in germ cells versus somatic cells by conditional mutogenesis
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identify target genes of the RARa
transcription factor |
THE
FSRG/BRD BROMODOMAIN-CONTAINING PROTEINS AND REPRODUCTIVE TISSUES
We
have cloned and characterized some properties of Fsrg1 (now
designated Brd2), a mouse gene that is homologous to the Drosophila
gene female sterile homeotic (fsh). We have also shown
that Brd2 is a member along with Brd3, Brd4,
and Brdt of a small sub-family of bromodomain-containing genes
that are differentially expressed during spermatogenesis. Given
the unique expression patterns of the Brd gene family, we are proposing
that they will have important functions in various aspects of the reproductive
system.
Our current projects:
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characterize
the biochemical properties of the Brd family of proteins by identifying
proteins with which they associate in vivo, using
a yeast 2-hybrid approach |
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determine
the effect of mutating Brd2
and Brdt in mice in vivo,
with particular focus on spermatogenesis and oogenesis
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