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Functional interaction between coactivators CBP/p300, PCAF, and transcription factor FKLF2
Chao-Zhong Song1, Kimberly Keller, Ken Murata
1Division of Medical Genetics, Department of Medicine, University of Washington, Seattle, WA 98195, USA.
Abstract:
The Sp1/KLF family of factors regulates diverse cellular processes, including growth and development. Fetal Krüppel-like factor (FKLF2) is a new member of this family. In this study, we characterized the coactivators involved in FKLF2 transcriptional activation. Our results show that both CBP/p300 and p300/CBP-associated factor (PCAF) enhance FKLF2 transcriptional activity. We demonstrate that the acetyltransferase activity of PCAF but not that of CBP/p300 is required for stimulating FKLF2 transcription activity. We further show that p300 and PCAF act cooperatively in stimulating FKLF2 transcriptional activation. FKLF2 interacts with both CBP and PCAF through specific domains, and CBP and PCAF acetylate FKLF2. Both CBP/p300 and PCAF stimulate FKLF2 DNA binding activity. The integrity of the acetyltransferase domain of PCAF but not that of CBP/p300 is required for stimulating FKLF2 DNA binding activity. These results demonstrate that CBP/p300 and PCAF stimulate FKLF2 transcriptional activity at least by enhancing its DNA binding. The acetyltransferase activities of CBP/p300 and PCAF play a distinct role in stimulating FKLF2 transcription and DNA binding.
Insights
Fetal Krüppel-like factor 2 (FKLF2) transcriptional activity is enhanced by coactivators CBP/p300 and PCAF. PCAF
Area of Science:
- Molecular Biology
- Gene Regulation
- Transcription Factors
Background:
- The Sp1/KLF family regulates crucial cellular processes.
- Fetal Krüppel-like factor 2 (FKLF2) is a recently identified member of this family.
- Understanding FKLF2's transcriptional regulation is key to deciphering its role in growth and development.
Purpose of the Study:
- To identify and characterize coactivators involved in FKLF2 transcriptional activation.
- To elucidate the specific roles of CBP/p300 and PCAF in FKLF2 function.
- To investigate the mechanism by which coactivators enhance FKLF2 activity.
Main Methods:
- Co-transfection assays to measure transcriptional activity.
- Western blotting and co-immunoprecipitation to assess protein interactions.
- In vitro acetylation assays to determine enzyme activity.
- Electrophoretic mobility shift assays (EMSAs) to evaluate DNA binding.
Main Results:
- Both CBP/p300 and PCAF significantly enhance FKLF2 transcriptional activity.
- PCAF's acetyltransferase activity is essential for FKLF2 transcription, while CBP/p300's is not.
- CBP/p300 and PCAF cooperate to boost FKLF2 transcriptional activation.
- FKLF2 interacts with CBP and PCAF, and is acetylated by both.
- Both coactivators enhance FKLF2 DNA binding, with PCAF's acetyltransferase domain being critical.
Conclusions:
- CBP/p300 and PCAF are key coactivators for FKLF2.
- Distinct roles of acetyltransferase activities of CBP/p300 and PCAF in FKLF2 transcription and DNA binding.
- Enhanced DNA binding is a primary mechanism for FKLF2 transcriptional stimulation by these coactivators.