TY - JOUR
T1 - Regulation of the Chlamydomonas cell cycle by a stable, chromatin-associated retinoblastoma tumor suppressor complex
AU - Olson, Bradley J.S.C.
AU - Oberholzer, Michael
AU - Li, Yubing
AU - Zones, James M.
AU - Kohli, Harjivan S.
AU - Bisova, Katerina
AU - Fang, Su Chiung
AU - Meisenhelder, Jill
AU - Hunter, Tony
AU - Umena, James G.
PY - 2010/10
Y1 - 2010/10
N2 - We examined the cell cycle dynamics of the retinoblastoma (RB) protein complex in the unicellular alga reinhardtii that has single homologs for each subunit-RB, E2F, and DP. We found that Chlamydomonas RB (encoded by MAT3) is a cell cycle-regulated phosphorprotein, that E2F1-DP1 can bind to a consensus E2F site, and that all three proteins interact in vivo to form complex that can be quantitatively immunopurified. Yeast two-hybrid assays revealed the formation of a ternary complex between MAT3, DP1, and E2F1 that requires a C-terminal motif in E2F1 analogous to the RB binding domain of plant and animal E2Fs. We examined the abundance of MAT3/RB and E2F1-DP1 in highly synchronous cultures and found that they are synthesized and remain stably associated throughout the cell cycle with no detectable fraction of free E2F1-DP1. Consistent with their stable association, MAT3/RB and DP1 are constitutively nuclear, and MAT3/ RB does not require DP1-E2F1 for nuclear localization. In the nucleus, MAT3/RB remains bound to chromatin throughout the cell cycle, and its chromatin binding is mediated through E2F1-DP1. Together, our data show that E2F-DP complexes can regulate the cell cycle without dissociation of their RB-related subunit and that other changes may be sufficient to convert RB-E2F-DP from a cell cycle repressor to an activator.
AB - We examined the cell cycle dynamics of the retinoblastoma (RB) protein complex in the unicellular alga reinhardtii that has single homologs for each subunit-RB, E2F, and DP. We found that Chlamydomonas RB (encoded by MAT3) is a cell cycle-regulated phosphorprotein, that E2F1-DP1 can bind to a consensus E2F site, and that all three proteins interact in vivo to form complex that can be quantitatively immunopurified. Yeast two-hybrid assays revealed the formation of a ternary complex between MAT3, DP1, and E2F1 that requires a C-terminal motif in E2F1 analogous to the RB binding domain of plant and animal E2Fs. We examined the abundance of MAT3/RB and E2F1-DP1 in highly synchronous cultures and found that they are synthesized and remain stably associated throughout the cell cycle with no detectable fraction of free E2F1-DP1. Consistent with their stable association, MAT3/RB and DP1 are constitutively nuclear, and MAT3/ RB does not require DP1-E2F1 for nuclear localization. In the nucleus, MAT3/RB remains bound to chromatin throughout the cell cycle, and its chromatin binding is mediated through E2F1-DP1. Together, our data show that E2F-DP complexes can regulate the cell cycle without dissociation of their RB-related subunit and that other changes may be sufficient to convert RB-E2F-DP from a cell cycle repressor to an activator.
UR - http://www.scopus.com/inward/record.url?scp=78649506206&partnerID=8YFLogxK
U2 - 10.1105/tpc.110.076067
DO - 10.1105/tpc.110.076067
M3 - Article
AN - SCOPUS:78649506206
SN - 1040-4651
VL - 22
SP - 3331
EP - 3347
JO - Plant Cell
JF - Plant Cell
IS - 10
ER -