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Molecular and functional characterisation of E2F-5, a new member of the E2F family
V Buck1, K E Allen, T Sørensen
1Laboratory of Eukaryotic Molecular Genetics, MRC National Institute for Medical Research, London, UK.
Abstract:
The transcription factor DRTF1/E2F is implicated in the control of cellular proliferation due to its interaction with key regulators of cell cycle progression, such as the retinoblastoma tumour suppressor gene product and related pocket proteins, cyclins and cyclin-dependent kinases. DRTF1/E2F DNA binding activity arises when a member of two distinct families of proteins, DP and E2F, interact as DP/E2F heterodimers. Here, we report the isolation and characterisation of a new member of the E2F family of proteins, called E2F-5. E2F-5 was isolated through a yeast two hybrid assay in which a 14.5 d.p.c. mouse embryo library was screened for molecules capable of binding to murine DP-1, but also interacts with all known members of the DP family of proteins. E2F-5 exists as a physiological heterodimer with DP-1 in the generic DRTF1/E2F DNA binding activity present in mammalian cell extracts, an interaction which results in co-operative DNA binding activity and transcriptional activation through the E2F site. A potent transcriptional activation domain, which functions in both yeast and mammalian cells and resides in the C-terminal region of E2F-5, is specifically inactivated upon pocket protein binding. Comparison of the sequence with other members of the family indicates that E2F-5 shows a greater level of similarity with E2F-4 than to E2F-1, -2 and -3. The structural and functional similarity of E2F-5 and E2F-4 defines a subfamily of E2F proteins.
Insights
Researchers identified E2F-5, a new transcription factor crucial for cellular proliferation. E2F-5 forms heterodimers with DP proteins, enhancing DNA binding and transcriptional activation, and is regulated by pocket proteins.
Area of Science:
- Molecular Biology
- Cell Biology
- Genetics
Background:
- The transcription factor DRTF1/E2F regulates cellular proliferation through interactions with cell cycle proteins.
- DRTF1/E2F DNA binding activity requires heterodimerization of DP and E2F family proteins.
Purpose of the Study:
- To isolate and characterize a novel member of the E2F family, designated E2F-5.
- To elucidate the interaction and functional properties of E2F-5 in the context of DRTF1/E2F activity.
Main Methods:
- Yeast two-hybrid assay using a 14.5 d.p.c. mouse embryo library to screen for DP-1 binding proteins.
- Characterization of E2F-5's interaction with DP family proteins and its DNA binding and transcriptional activation capabilities.
- Sequence comparison with known E2F family members.
Main Results:
- E2F-5 was identified and shown to interact with all known DP family proteins, particularly murine DP-1.
- E2F-5 forms physiological heterodimers with DP-1, contributing to DRTF1/E2F DNA binding activity and transcriptional activation.
- A C-terminal transcriptional activation domain in E2F-5 is inactivated by pocket protein binding.
- E2F-5 exhibits greater sequence similarity to E2F-4 than to E2F-1, -2, or -3.
Conclusions:
- E2F-5 is a novel E2F family member involved in regulating cellular proliferation.
- E2F-5 and E2F-4 form a distinct subfamily within the E2F protein family due to their structural and functional similarities.