Sequence-specific DNA binding and transcriptional regulation by the promyelocytic leukemia zinc finger protein.
Item
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Title
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Sequence-specific DNA binding and transcriptional regulation by the promyelocytic leukemia zinc finger protein.
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Identifier
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AAI9807958
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identifier
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9807958
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Creator
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Li, Jia-Yuan.
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Contributor
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Adviser: Jonathan D. Licht
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Date
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1997
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Language
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English
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Publisher
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City University of New York.
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Subject
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Biology, Molecular | Health Sciences, Oncology
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Abstract
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The Promyelocytic Leukemia Zinc Finger gene (PLZF) is rearranged in the chromosomal translocation t(11;17)(q23;21) associated with a retinoic acid resistant form of acute promyelocytic leukemia. The translocation fuses the retinoic acid receptor {dollar}\alpha{dollar} gene to PLZF producing reciprocal fusion proteins hypothesized to play a prominent role in leukemogenesis. PLZF encodes a 81 kD transcription factor with nine Kruppel like C2-H2 zinc fingers located in a single domain near the C-terminus of the protein. The last seven of PLZF's zinc fingers are retained in the t(11;17) fusion protein RAR{dollar}\alpha{dollar}-PLZF suggesting that RAR{dollar}\alpha{dollar}-PLZF plays a role in leukemogenesis by competing with wildtype PLZF for response elements. In this thesis, I identified several PLZF DNA binding sites which suggests that zinc finger proteins in general have the ability to recognize a spectrum of DNA sequences with different affinities. One binding site recognized specifically by the PLZF protein is the {dollar}\beta{dollar}-retinoic acid response element. The biological implications of such an interaction is that PLZF may regulate a subset of retinoic acid responsive genes and that restoration of the RAR{dollar}\alpha{dollar} pathway by all-trans retinoic acid may not compensate for transcriptional deregulation by RAR{dollar}\alpha{dollar}-PLZF, an aberrant transcription factor. Through PCR based binding site selection, I also identified a high affinity binding site for PLZF and showed that PLZF binds to this site through its most carboxyl seven zinc fingers. In cotransfection experiments, PLZF repressed transcription through this high affinity binding site. Transfected RAR{dollar}\alpha{dollar}-PLZF resulted in significant loss of repression (compared to that of wildtype PLZF). To identify important functional domains within its effector region, various fragments of the PLZF were fused to the Gal4p DNA binding domain. Constructs encoding these chimeric proteins were cotransfected with a reporter plasmid containing five copies of Gal4p DNA binding site. These experiments demonstrate that transcription repression by PLZF is mediated by two separate domains residing between amino acids 1-100 and amino acids 200-300. The presence of two non-contiguous repression domains implies that PLZF may repress transcription by effecting more than one molecular target. There is also a transcription activation domain within the PLZF effector domain which resides between amino acids 100-200. The presence of both activation and repression domains within one transcription factor implies that PLZF may be bi-functional depending on the presence of other factors, post-translation modification or the promoter architecture of target genes.
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Type
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dissertation
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Source
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PQT Legacy CUNY.xlsx
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degree
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Ph.D.
