Sp2 DNA Binding Activity and trans-Activation Are Negatively Regulated in Mammalian Cells
Previous studies have indicated that Sp2 binds poorly to GC-rich sequences bound by Sp1 and Sp3, and further functional analyses of Sp2 have been limited. To study Sp2-mediated transcription, we employed a PCR-based protocol to determine the Sp2 consensus DNA-binding sequence (5â²-GGGCGGGAC-3â²) a...
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Published in | The Journal of biological chemistry Vol. 279; no. 14; pp. 13911 - 13924 |
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Main Authors | , , |
Format | Journal Article |
Language | English |
Published |
United States
American Society for Biochemistry and Molecular Biology
02.04.2004
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Subjects | |
Online Access | Get full text |
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Summary: | Previous studies have indicated that Sp2 binds poorly to GC-rich sequences bound by Sp1 and Sp3, and further functional analyses
of Sp2 have been limited. To study Sp2-mediated transcription, we employed a PCR-based protocol to determine the Sp2 consensus
DNA-binding sequence (5â²-GGGCGGGAC-3â²) and performed kinetic experiments to show that Sp2 binds this consensus sequence with
high affinity (225 p m ) in vitro . To determine the functional consequence of Sp2 interaction with this sequence in vivo , we transformed well characterized Sp-binding sites within the dihydrofolate reductase (DHFR) promoter to consensus Sp2-binding
sites. Incorporation of Sp2-binding sites within the DHFR promoter increased Sp2-mediated trans -activation in transient co-transfection experiments but also revealed Sp2 to be a relatively weak trans -activator with little or no capacity for additive or synergistic trans -activation. Using chimeric molecules prepared with portions of Sp1 and Sp2 and the human prostate-specific antigen promoter,
we show that Sp2 DNA binding activity and trans -activation are negatively regulated in mammalian cells. Taken together, our data indicate that Sp2 is functionally distinct
relative to other Sp family members and suggest that Sp2 may play a unique role in cell physiology. |
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Bibliography: | ObjectType-Article-2 SourceType-Scholarly Journals-1 ObjectType-Feature-1 content type line 23 ObjectType-Article-1 ObjectType-Feature-2 |
ISSN: | 0021-9258 1083-351X |
DOI: | 10.1074/jbc.M313589200 |