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DAVID REISMAN, Ph.D.
Boggs,
K. and Reisman, D. The induction of p53 transcription prior
to DNA synthesis is regulated through a novel regulatory
element within the p53 promoter. Oncogene 2005 in press.
p53 mRNA levels are tightly regulated
during the cell cycle with its transcription being induced
prior to DNA synthesis. However, the mechanism controlling
this regulation is not well defined. Through characterizing
an additional 1000bp of upstream DNA sequences of the murine
p53 gene, we identified new positive and negative regulatory
elements. Furthermore, we found a trans-acting factor(s)
that binds within a positive cis-acting element (-972/-953)
in a manner indicative of cell cycle regulation. When
Swiss3T3 cells are arrested by serum depletion p53 mRNA
levels decrease and binding of this regulatory factor(s) to
the promoter is reduced. Upon serum stimulation the
regulatory factor(s) binds the promoter and p53 mRNA levels
increase prior to the cells entering S phase. When the
factors are experimentally sequestered from the promoter or
when the regulatory element is deleted from the promoter
overall p53 promoter activity is reduced. There is no
further reduction in p53 promoter activity upon serum
depletion and the kinetics of induction is delayed by
approximately 5 hours. These findings indicate that a
factor(s) binding within the -972/-953 regulatory element on
the p53 promoter is important for the proper regulation of
p53 mRNA expression in response to mitogen stimulation.
Initial analysis of proteins that bind to this element
indicate that a member of the C/EBP family of transcription
factors may play a role in this regulation.
Edwards, S. and Reisman, D.
Localization of a mutant p53 response element on the Tissue
Inhibitor of Metalloproteinase-3 promoter: mutant p53
activities are distinct from wild-type Cancer Lett. 2005 in
press
Missense mutations in the p53 gene have
been observed in greater than 60% of all human tumors.
Recent evidence indicates that some mutations in p53 arise
as the cancer progresses from a benign tumor to a metastatic
tumor and that these mutations in p53 actively contribute to
the process of cancer progression. Previously, we reported
that the expression of the gene encoding the tissue
inhibitor of metalloproteinase-3 (TIMP-3) is repressed in
cells expressing codons 248 and 281 mutant p53 alleles. The
ability of tumor-derived p53 mutants to inhibit TIMP-3
expression provides a novel mechanism for understanding how
p53 mutations might contribute to tumorigenesis. Since
mutant p53 is often expressed at elevated levels in a
variety of cancers, the generation of cells in a tumor
carrying certain mutations in p53 would cause
inappropriately reduced expression of TIMP-3 and lead to
elevated matrix metalloproteinase activity. We present the
results of experiments that begin to determine the mechanism
by which mutant p53 represses TIMP-3 gene expression. By
generating deletion derivatives of the TIMP-3 promoter and
testing them for expression and by performing DNA protein
binding assays on the regions determined to be required for
repression, we have identified elements that are essential
for mutant p53-mediated transcriptional repression. These
elements respond specifically to mutant but not wild type
p53. While mutant p53 itself does not bind to the TIMP-3
promoter, we provide evidence for the presence of DNA
binding proteins whose activity is enhanced in the presence
of mutant p53.
Reisman, D., Wallace, J., and Lu, G.
2004. Loss of heterozygosity and p53 expression in Pterygium.
Cancer Letters, 2004 206, 77-83.
While the pathogenesis of pterygium is
still not well understood, environmental factors such at UV
light, appear to play an important role in its development.
UV radiation can cause mutations in genes such as the p53
tumor suppressor gene, that when inactivated through
mutation and LOH can lead to cell proliferation and genomic
instability. However, aside from mutations in the gene,
other mechanisms have been identified that can lead to loss
of p53 function. These include the interaction of the p53
protein with cellular or viral gene products that lead to
the inactivation of p53 or to its rapid degradation as well
as the silencing of transcription of the p53 gene through
the aberrant expression of factors that control p53
expression. We have analyzed the status and expression of
the p53 gene in epithelial cells derived from pterygium and
have demonstrated that the p53 gene has undergone a
monoallelic deletion. Assays for both p53 protein and mRNA
revealed that the remaining allele in these cells is not
expressed at detectable levels. Furthermore, the remaining
allele, by DNA sequence analysis appears to remain wild
type. The mechanism of silencing the p53 gene and the loss
of p53 expression in these cells is currently under
investigation.
Durland-Busbice, S. and Reisman, D.
2002. The lack of p53 expression in human myeloid leukemias
is not due to mutations in transcriptional regulatory
regions of the gene. Leukemia, 2002 16, 2165-2167.
The normal or wild type p53 gene encodes
a DNA-binding transcription factor that is responsible for
cell cycle checkpoints that are activated after exposure to
DNA damaging agents (1). A hallmark of many tumor cells is
that the p53 gene is mutated and expressed at greatly
elevated levels. We have been interested in determining the
cause of lack of expression of the p53 gene in cell lines
derived from myeloid leukemias. Although numerous tumor
types express elevated levels of mutant p53, it has been
demonstrated that some astrocytomas (2), a high proportion
of breast cancers (3) and many human myeloid leukemias do
not express detectable levels of p53 mRNA (4). The mechanism
responsible for the silencing of p53 expression in myeloid
tumors remains unknown. The loss of p53 gene expression in
these cells is likely to be an important step in cancer
progression and could result from cis-acting mutations
within the regulatory regions of the gene or from the loss
of required trans-acting factors. Our goal has been to
determine whether mutational events in the p53
transcriptional regulatory regions of the gene represent a
mechanism by which p53 expression is silenced.
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