Dual mechanisms for the inhibition of E2F binding to RB by cyclin- dependent kinase-mediated RB phosphorylation

Erik S. Knudsen, Jean Y J Wang

Research output: Contribution to journalArticle

218 Citations (Scopus)

Abstract

The growth suppression function of RB is dependent on its protein binding activity. RB contains at least three distinct protein binding functions: (i) the A/B pocket, which binds proteins with the LXCXE motif; (ii) the C pocket, which binds the c-Abl tyrosine kinase; and (ii) the large A/B pocket, which binds the E2F family of transcription factors. Phosphorylation of RB, which is catalyzed by cyclin-dependent protein kinases, inhibits all three protein binding activities. We have previously shown that LXCXE binding is inactivated by the phosphorylation of two threonines (Thr821 and Thr826), while the C pocket is inhibited by the phosphorylation of two serines (Set807 and Ser811). In this report, we show that the E2F binding activity of RB is inhibited by two sets of phosphorylation sites acting through distinct mechanisms. Phosphorylation at several of the seven C-terminal sites can inhibit E2F binding. Additionally, phosphorylation of two serine sites in the insert domain can inhibit E2F binding, but this inhibition requires the presence of the RB N-terminal region. RB mutant proteins lacking all seven C-terminal sites and two insert domain serines can block Rat-1 cells in G1. These RB mutants can bind LXCXE proteins, c-Ab1, and E2F even after they become phosphorylated at the remaining nonmutated sites. Thus, multiple phosphorylation sites regulate the protein binding activities of RB through different mechanisms, and a constitutive growth suppressor can be generated through the combined mutation of the relevant phosphorylation sites in RB.

Original languageEnglish (US)
Pages (from-to)5771-5783
Number of pages13
JournalMolecular and Cellular Biology
Volume17
Issue number10
StatePublished - Oct 1997

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Cyclin-Dependent Kinases
Phosphorylation
Protein Binding
Serine
E2F Transcription Factors
Mutant Proteins
Threonine
Growth
Protein-Tyrosine Kinases
Proteins
Mutation

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics
  • Cell Biology

Cite this

Dual mechanisms for the inhibition of E2F binding to RB by cyclin- dependent kinase-mediated RB phosphorylation. / Knudsen, Erik S.; Wang, Jean Y J.

In: Molecular and Cellular Biology, Vol. 17, No. 10, 10.1997, p. 5771-5783.

Research output: Contribution to journalArticle

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