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PMID: 10891495 Published · ppublish English Journal Article Research Support, U.S. Gov't, P.H.S.

Establishment of irreversible growth arrest in myogenic differentiation requires the RB LXCXE-binding function.

Molecular and cellular biology ·Vol. 20 ·No. 15 ·2000-08-00 ·Pages 5571-80

Chen TT, Wang JY

Abstract

The crystal structure of the A-B domain of RB has defined the binding pocket for the LXCXE peptide motif. Using the crystal structure as a guide, we have inactivated the LXCXE-binding pocket by replacing N757 with Phe [to obtain RB(N757F)]. RB(N757F) does not bind to viral oncoproteins but retains the ability to bind and inhibit E2F. RB(N757F) is less effective than the wild-type RB [RB(WT)] in repressing E2F-regulated transcription, and its repression activity is not affected by trichostatin A, an inhibitor of histone deacetylases. However, RB(N757F) is as effective as RB(WT) in suppressing cell growth. Interestingly, RB(N757F) cannot establish an irreversible growth arrest in differentiated myocytes. Differentiated myocytes with RB(WT) become refractory to serum. By contrast, differentiated myocytes with RB(N757F) undergo DNA synthesis and phosphorylate RB(N757F) in response to serum, despite a high level of p21Cip1 expression. Mutation of the phosphorylation sites in RB(N757F) rescued its defect and allowed myocytes to permanently withdraw from the cell cycle. These results demonstrate that it is possible to inactivate the LXCXE-binding pocket without compromising the overall integrity of RB. Moreover, the LXCXE-binding pocket is dispensable for the intrinsic growth suppression function of RB. However, the LXCXE-binding function is essential for RB to establish the serum-refractory state in differentiated myocytes.

MeSH Terms
Amino Acid Motifs Animals Base Sequence Binding Sites Carrier Proteins Cell Cycle Proteins Cell Differentiation/genetics Cell Division/genetics Cells, Cultured Cyclin-Dependent Kinase Inhibitor p16/metabolism Cyclin-Dependent Kinase Inhibitor p21 Cyclins/metabolism DNA-Binding Proteins E2F Transcription Factors Histone Deacetylase 1 Histone Deacetylases/metabolism Humans Mice Molecular Sequence Data Muscle, Skeletal/cytology,physiology MyoD Protein/metabolism Point Mutation Retinoblastoma Protein/genetics,metabolism Retinoblastoma-Binding Protein 1 Transcription Factor DP1 Transcription Factors/metabolism
Chemicals
Arid4a protein, mouse CDKN1A protein, human Carrier Proteins Cdkn1a protein, mouse Cell Cycle Proteins Cyclin-Dependent Kinase Inhibitor p16 Cyclin-Dependent Kinase Inhibitor p21 Cyclins DNA-Binding Proteins E2F Transcription Factors MyoD Protein Retinoblastoma Protein Retinoblastoma-Binding Protein 1 Transcription Factor DP1 Transcription Factors HDAC1 protein, human Histone Deacetylase 1 Histone Deacetylases
Authors & Affiliations
2 authors, click to expand affiliations / ORCID
Chen T T
Department of Biology and the Cancer Center, University of California, San Diego, La Jolla, California 92093-0322, USA.
Wang J Y
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Article Info
Journal
Molecular and cellular biology
Abbr.
Mol Cell Biol
ISSN
0270-7306
DOI
10.1128/MCB.20.15.5571-5580.2000
Published
2000-08-00
Pages
5571-80
Language
English
Region
United States
NLM ID
8109087
PMCID
PMC86015
Subset
IM
Grants
NCI NIH HHS · R01 CA058320 · United States
NCI NIH HHS · CA58320 · United States
External Links
PubMed source DOI: 10.1128/MCB.20.15.5571-5580.2000 PMC Full Text View journal details
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