Regulation of retinoblastoma protein activity by the c terminal domain

Research output: Contribution to journalArticle

Abstract

Purpose. The retinoblastoma protein (Rb) is a iumor suppressor that regulates cell proliferation by controlling cellular transition from Gl to S phase of the cell cycle. The A-B pocket of Rb forms a transcriptional represser motif which is regulated through phosphorylation of the C-terminal of Rb. We examined the mechanism of C-terminal regulation of the A-B pocket. Methods. Transient iransfections of plasmids expressing GAL4-R5 or LexA-Rb fusion proteins in C33A (Rb-t cells, and detection of transcriptional activity using a reporter plasmid containing the chloramphenico! acetyttransferase (CAT) gene, SV40 promoter, and GAL4 or LexA binding sites. Results. Overexpression of the A-B pocket as a GAL4 or LexA fusion protein resulted in repression of CAT activity. Coexpression of the C-terminal domain on a separate protein blocked this transcriptional repression. Conclusions. Since the C-terminal can block transcriptional repression by the A-B pocket when expressed on a separate protein, the mechanism of C-terminal regulation may depend on interdomain interactions, rather than intramolecular conformational changes.

Original languageEnglish
JournalInvestigative Ophthalmology and Visual Science
Volume38
Issue number4
StatePublished - Dec 1 1997
Externally publishedYes

Fingerprint

Retinoblastoma Protein
Plasmids
Protein C
S Phase
Cell Cycle
Proteins
Binding Sites
Phosphorylation
Cell Proliferation
Genes

ASJC Scopus subject areas

  • Ophthalmology

Cite this

Regulation of retinoblastoma protein activity by the c terminal domain. / William Harbour, J.; Luo, R. X.

In: Investigative Ophthalmology and Visual Science, Vol. 38, No. 4, 01.12.1997.

Research output: Contribution to journalArticle

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N2 - Purpose. The retinoblastoma protein (Rb) is a iumor suppressor that regulates cell proliferation by controlling cellular transition from Gl to S phase of the cell cycle. The A-B pocket of Rb forms a transcriptional represser motif which is regulated through phosphorylation of the C-terminal of Rb. We examined the mechanism of C-terminal regulation of the A-B pocket. Methods. Transient iransfections of plasmids expressing GAL4-R5 or LexA-Rb fusion proteins in C33A (Rb-t cells, and detection of transcriptional activity using a reporter plasmid containing the chloramphenico! acetyttransferase (CAT) gene, SV40 promoter, and GAL4 or LexA binding sites. Results. Overexpression of the A-B pocket as a GAL4 or LexA fusion protein resulted in repression of CAT activity. Coexpression of the C-terminal domain on a separate protein blocked this transcriptional repression. Conclusions. Since the C-terminal can block transcriptional repression by the A-B pocket when expressed on a separate protein, the mechanism of C-terminal regulation may depend on interdomain interactions, rather than intramolecular conformational changes.

AB - Purpose. The retinoblastoma protein (Rb) is a iumor suppressor that regulates cell proliferation by controlling cellular transition from Gl to S phase of the cell cycle. The A-B pocket of Rb forms a transcriptional represser motif which is regulated through phosphorylation of the C-terminal of Rb. We examined the mechanism of C-terminal regulation of the A-B pocket. Methods. Transient iransfections of plasmids expressing GAL4-R5 or LexA-Rb fusion proteins in C33A (Rb-t cells, and detection of transcriptional activity using a reporter plasmid containing the chloramphenico! acetyttransferase (CAT) gene, SV40 promoter, and GAL4 or LexA binding sites. Results. Overexpression of the A-B pocket as a GAL4 or LexA fusion protein resulted in repression of CAT activity. Coexpression of the C-terminal domain on a separate protein blocked this transcriptional repression. Conclusions. Since the C-terminal can block transcriptional repression by the A-B pocket when expressed on a separate protein, the mechanism of C-terminal regulation may depend on interdomain interactions, rather than intramolecular conformational changes.

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