The N-terminal domain of the Drosophila retinoblastoma protein Rbf1 interacts with ORC and associates with chromatin in an E2F independent manner.
BACKGROUND
The retinoblastoma (Rb) tumor suppressor protein can function as a DNA replication inhibitor as well as a transcription factor. Regulation of DNA replication may occur through interaction of Rb with the origin recognition complex (ORC).


RESULTS
We characterized the interaction of Drosophila Rb, Rbf1, with ORC. Using expression of proteins in Drosophila S2 cells, we found that an N-terminal Rbf1 fragment (amino acids 1-345) is sufficient for Rbf1 association with ORC but does not bind to dE2F1. We also found that the C-terminal half of Rbf1 (amino acids 345-845) interacts with ORC. We observed that the amino-terminal domain of Rbf1 localizes to chromatin in vivo and associates with chromosomal regions implicated in replication initiation, including colocalization with Orc2 and acetylated histone H4.


CONCLUSIONS
Our results suggest that Rbf1 can associate with ORC and chromatin through domains independent of the E2F binding site. We infer that Rbf1 may play a role in regulating replication directly through its association with ORC and/or chromatin factors other than E2F. Our data suggest an important role for retinoblastoma family proteins in cell proliferation and tumor suppression through interaction with the replication initiation machinery.
Ahlander
Joseph
J
Department of Molecular and Cellular Biology, University of Arizona, Tucson, Arizona, United States of America.
Chen
Xiao-Bo
XB
Bosco
Giovanni
G
D000818:Animals; D001665:Binding Sites; D002467:Cell Nucleus / Q000378:metabolism; D049109:Cell Proliferation; D002843:Chromatin / Q000737:chemistry / Q000378:metabolism; D004330:Drosophila / Q000378:metabolism*; D029721:Drosophila Proteins / Q000378:metabolism*; D050684:E2F Transcription Factors / Q000737:chemistry*; D031541:Fluorescence Resonance Energy Transfer; D006657:Histones / Q000737:chemistry; D008954:Models, Biological; D051738:Origin Recognition Complex; D038761:Photobleaching; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D016160:Retinoblastoma Protein; D014157:Transcription Factors / Q000378:metabolism*
D002843:Chromatin; D029721:Drosophila Proteins; D050684:E2F Transcription Factors; D006657:Histones; D051738:Origin Recognition Complex; C099965:Rbf protein, Drosophila; D016160:Retinoblastoma Protein; D014157:Transcription Factors