Retinoblastoma/p107/p130 pocket proteins. Proteins dynamics and interactions with target gene promoters

Kristy R. Stengel, Chellappagounder Thangavel, David A. Solomon, Steve P. Angus, Yi Zheng, Erik S. Knudsen

Research output: Contribution to journalArticle

25 Citations (Scopus)

Abstract

The retinoblastoma (RB) tumor suppressor and its family members, p107 and p130, function by repressing E2F transcription factor activity to limit the expression of genes required for cell cycle progression. Traditionally, it is thought that the RB family proteins repress E2F target gene expression through complexing with E2F at gene promoters. However, whereas chromatin immunoprecipitation experiments have demonstrated p107 and p130 at E2F-responsive promoters, RB chromatin association has not been reliably observed. Here we used green fluorescent protein-tagged proteins to rigorously explore the mechanism of RB-mediated transcriptional repression relative to its p107 and p130 family members. The use of live cell fluorescent imaging demonstrated that RB, p107, and p130 exhibit similar nuclear dynamics. Although these findings suggest a similar engagement with nuclear structures, chromatin immunoprecipitation approaches with multiple independent antibodies failed to detect the association of RB with target gene promoters. However, by employing antibodies directed against green fluorescent protein, we could utilize the same antibody to assess RB, p107, and p130 engagement. This approach demonstrated RB association with target gene promoters in a fashion analogous to p107 and p130. Extension of this technology demonstrated that direct RB phosphorylation disrupts promoter association to regulate transcription. Thus, RB is associated with promoters in a manner similar to p107/p130 and that association is modulated by phosphorylation during cell cycle progression.

Original languageEnglish (US)
Pages (from-to)19265-19271
Number of pages7
JournalJournal of Biological Chemistry
Volume284
Issue number29
DOIs
StatePublished - Jul 17 2009

Fingerprint

Retinoblastoma
Genes
Association reactions
Chromatin
Phosphorylation
Green Fluorescent Proteins
Proteins
Antibodies
Cells
E2F Transcription Factors
Chromatin Immunoprecipitation
Transcription
Gene expression
Tumors
cdc Genes
Retinoblastoma Protein
Imaging techniques
Cell Cycle
Experiments
Technology

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Molecular Biology

Cite this

Stengel, K. R., Thangavel, C., Solomon, D. A., Angus, S. P., Zheng, Y., & Knudsen, E. S. (2009). Retinoblastoma/p107/p130 pocket proteins. Proteins dynamics and interactions with target gene promoters. Journal of Biological Chemistry, 284(29), 19265-19271. https://doi.org/10.1074/jbc.M808740200

Retinoblastoma/p107/p130 pocket proteins. Proteins dynamics and interactions with target gene promoters. / Stengel, Kristy R.; Thangavel, Chellappagounder; Solomon, David A.; Angus, Steve P.; Zheng, Yi; Knudsen, Erik S.

In: Journal of Biological Chemistry, Vol. 284, No. 29, 17.07.2009, p. 19265-19271.

Research output: Contribution to journalArticle

Stengel, KR, Thangavel, C, Solomon, DA, Angus, SP, Zheng, Y & Knudsen, ES 2009, 'Retinoblastoma/p107/p130 pocket proteins. Proteins dynamics and interactions with target gene promoters', Journal of Biological Chemistry, vol. 284, no. 29, pp. 19265-19271. https://doi.org/10.1074/jbc.M808740200
Stengel, Kristy R. ; Thangavel, Chellappagounder ; Solomon, David A. ; Angus, Steve P. ; Zheng, Yi ; Knudsen, Erik S. / Retinoblastoma/p107/p130 pocket proteins. Proteins dynamics and interactions with target gene promoters. In: Journal of Biological Chemistry. 2009 ; Vol. 284, No. 29. pp. 19265-19271.
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