TY - JOUR
T1 - A mammalian bromodomain protein, Brd4, interacts with replication factor C and inhibits progression to S phase
AU - Maruyama, Tetsuo
AU - Farina, Andrea
AU - Dey, Anup
AU - Cheong, Jae Hun
AU - Bermudez, Vladimir P.
AU - Tamura, Tomohiko
AU - Sciortino, Selvaggia
AU - Shuman, Jon
AU - Hurwitz, Jerard
AU - Ozato, Keiko
PY - 2002/9
Y1 - 2002/9
N2 - Brd4 belongs to the BET family of nuclear proteins that carry two bromodomains implicated in the interaction with chromatin. Expression of Brd4 correlates with cell growth and is induced during early G1 upon mitogenic stimuli. In the present study, we investigated the role of Brd4 in cell growth regulation. We found that ectopic expression of Brd4 in NIH 3T3 and HeLa cells inhibits cell cycle progression from G1 to S. Coimmunoprecipitation experiments showed that endogenous and transfected Brd4 interacts with replication factor C (RFC), the conserved five-subunit complex essential for DNA replication. In vitro analysis showed that Brd4 binds directly to the largest subunit, RFC-140, thereby interacting with the entire RFC. In line with the inhibitory activity seen in vivo, recombinant Brd4 inhibited RFC-dependent DNA elongation reactions in vitro. Analysis of Brd4 deletion mutants indicated that both the interaction with RFC-140 and the inhibition of entry into S phase are dependent on the second bromodomain of Brd4. Lastly, supporting the functional importance of this interaction, it was found that cotransfection with RFC-140 reduced the growth-inhibitory effect of Brd4. Taken as a whole, the present study suggests that Brd4 regulates cell cycle progression in part by interacting with RFC.
AB - Brd4 belongs to the BET family of nuclear proteins that carry two bromodomains implicated in the interaction with chromatin. Expression of Brd4 correlates with cell growth and is induced during early G1 upon mitogenic stimuli. In the present study, we investigated the role of Brd4 in cell growth regulation. We found that ectopic expression of Brd4 in NIH 3T3 and HeLa cells inhibits cell cycle progression from G1 to S. Coimmunoprecipitation experiments showed that endogenous and transfected Brd4 interacts with replication factor C (RFC), the conserved five-subunit complex essential for DNA replication. In vitro analysis showed that Brd4 binds directly to the largest subunit, RFC-140, thereby interacting with the entire RFC. In line with the inhibitory activity seen in vivo, recombinant Brd4 inhibited RFC-dependent DNA elongation reactions in vitro. Analysis of Brd4 deletion mutants indicated that both the interaction with RFC-140 and the inhibition of entry into S phase are dependent on the second bromodomain of Brd4. Lastly, supporting the functional importance of this interaction, it was found that cotransfection with RFC-140 reduced the growth-inhibitory effect of Brd4. Taken as a whole, the present study suggests that Brd4 regulates cell cycle progression in part by interacting with RFC.
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U2 - 10.1128/MCB.22.18.6509-6520.2002
DO - 10.1128/MCB.22.18.6509-6520.2002
M3 - Article
C2 - 12192049
AN - SCOPUS:0036724194
SN - 0270-7306
VL - 22
SP - 6509
EP - 6520
JO - Molecular and cellular biology
JF - Molecular and cellular biology
IS - 18
ER -